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Genome-wide Investigation of Cellular Functions for tRNA NucleusCytoplasm Trafficking in the Yeast Saccharomyces cerevisiae DISSERTATION Presented in Partial Fulfillment of the Requirements for the Degree Doctor of Philosophy in the Graduate School of The Ohio State University By Hui-Yi Chu Graduate Program in Molecular, Cellular and Developmental Biology The Ohio State University 2012 Dissertation Committee: Anita K. Hopper, Advisor Stephen Osmani Kurt Fredrick Jane Jackman Copyright by Hui-Yi Chu 2012 Abstract In eukaryotic cells tRNAs are transcribed in the nucleus and exported to the cytoplasm for their essential role in protein synthesis. This export event was thought to be unidirectional. Surprisingly, several lines of evidence showed that mature cytoplasmic tRNAs shuttle between nucleus and cytoplasm and their distribution is nutrient-dependent. This newly discovered tRNA retrograde process is conserved from yeast to vertebrates. Although how exactly the tRNA nuclear-cytoplasmic trafficking is regulated is still under investigation, previous studies identified several transporters involved in tRNA subcellular dynamics. At least three members of the β-importin family function in tRNA nuclear-cytoplasmic intracellular movement: (1) Los1 functions in both the tRNA primary export and re-export processes; (2) Mtr10, directly or indirectly, is responsible for the constitutive retrograde import of cytoplasmic tRNA to the nucleus; (3) Msn5 functions solely in the re-export process. In this thesis I focus on the physiological role(s) of the tRNA nuclear retrograde pathway. One possibility is that nuclear accumulation of cytoplasmic tRNA serves to modulate translation of particular transcripts. To test this hypothesis, I compared expression profiles from non-translating mRNAs and polyribosome-bound translating mRNAs collected from msn5Δ and mtr10Δ mutants and wild-type cells, in fed or acute amino acid starvation conditions. Microarray data revealed that several amino acid biosynthetic pathways, including the sulfur assimilation, arginine ii biosynthesis, and leucine biosynthesis pathways, are primary targets of the tRNA trafficking processes. I confirmed the microarray data by both Northern and Western blot analyses. Levels of all tested target proteins involved in such amino acid biogenesis pathways are down-regulated when the tRNA nuclear import or re-export is disrupted. The steady state levels of target total RNAs are similar between wild-type cells and tRNA trafficking defective mutants. The data suggest that the reduction of target proteins most likely results from translation defects of the target mRNAs. This study provides information that tRNA nuclear-cytoplasmic dynamics is connected to amino acid biosynthesis via control at the level of translation. iii This thesis is dedicated to my family and the memory of my dear mother. iv Acknowledgements First and foremost I want to thank my advisor Anita K. Hopper, for her warm encouragement and advice. From her I have learned how to enjoy the research life and open my mind for every possibility. She is always patient, optimistic, and enthusiastic, even during difficult moments. I appreciate all her supports and contributions of time, ideas, and funding in my Ph.D pursuits. When I lose my confidence, she always believed me more than myself. I thank her wide knowledge and logical way of thinking, which made this thesis possible. I am grateful for the excellent role model which she provided as a woman scientist. It is my honor to be one of her students. I am acknowledged my thesis committee, Professor Stephen Osmani, Professor Kurt Fredrick, and Professor Jane Jackman, for their interests, professional comments, and personal encouragements. I am also grateful that my previous advisor in the Nationwide Children Hospital, Dr. Akihira Otoshi, for his giving me the chance coming to the United States. I want to thank the director of MCDB graduate program, Professor David Bisaro, for his understanding and decision of accepting me, which made me get the opportunity to join in this wonderful lab. Being part of the Hopper lab would definitely become a beautiful memory in my life. It was my pleasure to work with such nice lab members. I thank our post-docs Rebecca, for her good ideas and always willing to help us whenever we have questions, v and Emily, for her nice communication skills. Thanks to Greetchen, Nripesh, Tsung-Po, Hsiao-Yun (Ivy), Jingyan, and Marina. When I became too serious or too panic, they provided laughs and supports. When I had successful experiments, they shared the joyful moments. My life in the OSU was fulfilling and the learning experience was most comprehensive in my study journey. As a hearing impaired foreigner, I sincerely thank Cindy Knecht and Andrea Crago from the disability service that was provided by the OSU and the Professional Reporters, Inc. Their amazing real-time transcribing work truly helped me overcome the double barriers of disability and language. Thank to OSU provided such a friendly campus. I thank people in the department of Molecular Genetics, in particular Professor Jim Hopper, Professor Paul Herman, and their lab members. In these years of joys and frustrations, thanks for all the supports my friends in Columbus. Thanks to Rosa, Keith and their dog Bailey, for their generosity and warm supports. Thanks to I-Ju, Liang-Chun, and Yuh-Ying, for banding together over food and life. I also thank Kuo-Fang and Shu-Hao, studying together with them was a different and great experience. Also thanks to many old friends in Taiwan, they keep caring about me no matter the distance. Finally, and most importantly, thanks to my parents for their nurture and education. Their unconditional love made me brave to face the challenges and was the motivation of my pursuits. Thanks to my sister and brother, and their families, for their understanding and supports. And last, but not least, to Yuan-Sheng, thank you for the love and consistent being there for me. vi Vita March 1978 ....................................................Born−Taipei, Taiwan 2000...............................................................B.S. Biology, National Taiwan Normal University, Taiwan 2003................................................................M.S. Genetics, National Yang-Ming University, Taiwan 2006 to present ..............................................Graduate Research Associate, Molecular, Cellular, and Developmental Biology Program, The Ohio State University Publication Lai, Y.-S., Murali, S., Ju, H. Y., Wu, M. F., Guo, I. C., Chen, S. C., Fang, K. and Chang, C. Y. Two iridovirus-susceptible cell lines established for kidney and liver of grouper, Epinephelus awoara, and partial characterization of grouper iridovirus. Journal of fish disease 23, 379-338, 2000. vii Lai, Y.-S., Murali, S., Chiu, H. C., Ju, H. Y., Lin, Y. S., Chen, S. C., Guo, I. C., Fang, K. and Chang, C. Y. Preparation of yellow grouper nervous necrosis virus in a new nodavirussusceptible cell lines from yellow grouper, Epinephelus awoara brain tissue. Journal of fish disease 24,299-309, 2001. Chu, H. Y. Phenotypic Analysis of the Aurora-A Transgenic Mice. Master Thesis. Institute of Genetics, National Yang Ming University, July, 2003. Chu, H. Y. and Otoshi, A. Cloning and functional analysis of hypothalamic homeobox gene Bsx1a and its isoform, Bsx1b., Mol Cell Biol. 27(10), 3743-9, 2007. Li, C.C., Chu, H.Y., Yang, C.W., Chou, C.K. and Tsai, T.F. Aurora-A overexpression in mouse liver causes p53-dependent premitotic arrest during liver regeneration. Mol Cancer Res. 7(5), 678-88, 2009. Chu, H. Y. and Hopper, A. K. Genome-wide investigation of the cellular functions for tRNA nucleus-cytoplasm trafficking in S. cerevisiae. (in prep.) Fields of Study Major Field: Molecular, Cellular and Developmental Biology viii Table of Contents Abstract ............................................................................................................................... ii Acknowledgements ............................................................................................................. v Vita .................................................................................................................................... vii Fields of Study ................................................................................................................. viii Table of Contents ............................................................................................................... ix List of Tables ................................................................................................................... xiv List of Figures .................................................................................................................. xvi Chapter 1 Introduction.................................................................................................. 1 1.1 Nucleocytoplasmic transport ................................................................................ 1 1.2 tRNA biology in yeast .......................................................................................... 5 1.2.1 The life of tRNA: biogenesis, maturation, and turnover ........................... 5 1.2.2 tRNA subcellular dynamics..................................................................... 10 1.3 Amino acid stress responses ............................................................................... 12 1.4 Microarray analysis ............................................................................................ 14 1.4.1 Microarray general applications and advanced usage for translational profiling ................................................................................................... 14 1.4.2 Data analysis............................................................................................ 16 1.5 Aim of this study ................................................................................................ 21 ix Chapter 2 Materials and Methods.............................................................................. 22 2.1 Yeast growth conditions and strain construction ............................................... 22 2.1.1 Media and growth conditions .................................................................. 22 2.1.2 Yeast strain construction ......................................................................... 23 2.1.3 Growth assay ........................................................................................... 25 2.2 Polysome profiles and RNA isolation ................................................................ 25 2.2.1 Polysome profiles .................................................................................... 25 2.2.2 Quantification of polysome profiles ........................................................ 27 2.2.3 RNA extraction........................................................................................ 28 2.3 Microarray data analysis .................................................................................... 28 2.3.1 Pre-processing of raw data ...................................................................... 29 2.3.2 Identification of significant differentially-expressed genes .................... 29 2.3.3 Hierarchical clustering of significantly changed target transcripts ......... 30 2.4 Northern blot analysis ........................................................................................ 30 2.4.1 Preparation of DNA probes ..................................................................... 31 2.5 Western blot analysis ......................................................................................... 31 2.5.1 Protein extraction .................................................................................... 31 2.5.2 Western blot and quantification .............................................................. 32 2.6 Cis-regulatory element analyses of targets......................................................... 33 2.6.1 Codon bias and amino acid usage ........................................................... 33 Chapter 3 Genome-wide investigations of cellular functions for tRNA nuclearcytoplasmic trafficking in regulation of translation .................................................... 43 x 3.1 Introduction ........................................................................................................ 43 3.2 Results ................................................................................................................ 48 3.2.1 Polysome profiles analysis ...................................................................... 48 3.2.2 Microarray data pre-processing ............................................................... 49 3.2.3 Identification of significant differentially-expressed genes (DEGs) ....... 57 3.2.4 Transcriptional control and RNA stability of target mRNAs are not affected .................................................................................................... 60 3.2.5 Verification of microarray data by western blot analyses ....................... 61 3.2.6 tRNA subcellular trafficking affects translation regulation of amino acid biosyntheses transcripts ........................................................................... 77 3.2.7 Cis-regulatory elements analysis of tRNA trafficking targets ................ 85 3.3 Discussion .......................................................................................................... 86 Chapter 4 Transcriptional analysis of mtr10Δ cells .................................................. 96 4.1 Introduction ........................................................................................................ 96 4.2 Results ................................................................................................................ 97 4.2.1 Microarray analysis of transcription profiles in msn5∆ cells .................. 98 4.2.2 Microarray analysis of transcription profiles in mtr10∆ cells ................. 98 4.2.3 Confirmations of selected target mRNA expressions by Northern blot analysis .................................................................................................. 104 4.2.4 Phenotypic analyses by growth assays .................................................. 105 4.3 Discussion ........................................................................................................ 112 Chapter 5 Summary and future directions ............................................................. 115 xi APPENDIX A: Tested possibilities for mechanism of down-regulated translation of the target mRNAs involved in the Met, Arg, and Leu biosynthetic pathways ..................... 117 APPENDIX B: Transcripts with decreased translation activity index in fed mtr10Δ cells ......................................................................................................................................... 121 APPENDIX C: Transcripts with increased translation activity index in fed mtr10Δ cells ......................................................................................................................................... 129 APPENDIX D: Transcripts with changed translation activity index in mtr10Δ cells in amino acid starvation condition ...................................................................................... 136 APPENDIX E: Transcripts with decreased translation activity index in fed msn5Δ cells ......................................................................................................................................... 138 APPENDIX F: Summary tables of functional catalogue analysis for affected transcripts in mtr10Δ cells in fed or amino acid starvation conditions............................................. 140 APPENDIX G: Down-regulated genes in mtr10Δ cells in fed condition ...................... 145 APPENDIX H: Up-regulated genes in mtr10Δ cells in fed condition ........................... 157 APPENDIX I: Down-regulated genes in mtr10Δ cells in amino acid starvation condition ......................................................................................................................................... 178 APPENDIX J: Up-regulated genes in mtr10Δ cells in amino acid starved condition ... 212 APPENDIX K: Summary tables of functional catalogue analysis for affected transcripts in msn5Δ cells in fed or amino acid starvation conditions.............................................. 251 APPENDIX L: Down-regulated genes in msn5Δ cells in fed condition........................ 256 APPENDIX M: Up-regulated genes in msn5Δ cells in fed condition ........................... 264 xii APPENDIX N: Down-regulated genes in msn5Δ cells in amino acid starvation condition ......................................................................................................................................... 275 APPENDIX O: Up-regulated genes in msn5Δ cells in amino acid starvation condition290 APPENDIX P: R codes for microarray data analysis .................................................... 302 References ....................................................................................................................... 326 xiii List of Tables Table 2. 1. Yeast strains used in this study .................................................................... 34 Table 2. 2. Oligonucleotides used in this study ............................................................. 38 Table 3. 1. Summary tables of numbers of differentially expressed genes in mtr10Δ or msn5Δ cells in fed or starved conditions. ......................................................................... 67 Table 4. 1. Numbers of significant differentially transcriptionally expressed genes from total mRNA samples of msn5Δ or mtr10Δ cells in fed or amino acid starvation conditions.. ...................................................................................................................... 103 Table 4. 2. GO term enrichment analysis and FunCat analysis of genes with significant changed in mtr10Δ cells in fed condition........................................................................ 107 Table 4. 3. GO term enrichment analysis and FunCat analysis of genes with significant changed in mtr10Δ cells in acute amino acid starved condition.. ................................... 108 Table F. 1. FunCat analysis of down-regulated transcripts in mtr10Δ cells in fed condition. ........................................................................................................................ 140 Table F. 2. FunCat analysis of up-regulated transcripts in mtr10Δ cells in fed condition. ......................................................................................................................................... 141 xiv Table F. 3. FunCat analysis of down-regulated transcripts in mtr10Δ cells in amino acid starved condition. ............................................................................................................ 142 Table F. 4. FunCat analysis of up-regulated transcripts in mtr10Δ cells in amino acid starved condition. ............................................................................................................ 143 Table K. 1. FunCat analysis of down-regulated transcripts in msn5Δ cells in fed condition. ........................................................................................................................ 251 Table K. 2. Up-regulated transcripts in msn5Δ cells in fed condition. ......................... 252 Table K. 3. FunCat analysis of down-regulated transcripts in msn5Δ cells in amino acid starvation condition......................................................................................................... 254 Table K. 4. FunCat analysis of up-regulated transcripts in msn5Δ cells in amino acid starvation condition......................................................................................................... 255 xv List of Figures Figure 1. 1. Ran Cycle and Ran-dependent nuclear export and import. ........................... 4 Figure 1. 2. Summary of tRNA processing pathways.. .................................................... 8 Figure 1. 3. Probe and probesets. .................................................................................... 19 Figure 2. 1. Illustration of quantification method. .......................................................... 27 Figure 3. 1. Current model of tRNA subcellular movement in yeast.. ........................... 46 Figure 3. 2. Flowchart of experimental designs.. ............................................................ 47 Figure 3. 3. Polysome profiles of wild-type, mtr10Δ, and msn5Δ cells collected from fed or 30-min amino acid starved conditions.. ........................................................................ 51 Figure 3. 4. Polysome profiles of wild-type, los1Δ, and los1Δ msn5Δ cells collected from fed or 30-min amino acid starved condition. ........................................................... 52 Figure 3. 5. Quantification of polysome profiles of tRNA trafficking mutants in fed (AC) or amino acid starved (D-F) conditions. ...................................................................... 54 Figure 3. 6. Summary of microarray experiments.. ........................................................ 55 Figure 3. 7. Global assessment of microarray raw data set from fractionated RNA samples by using principle component analysis (PCA). ................................................... 56 Figure 3. 8. Box plot of raw microarray data. ................................................................. 64 xvi Figure 3. 9. Box plot of RMA normalized microarray data............................................ 65 Figure 3. 10. Volcano plot showing differential expressed genes selected by P/NP index compared from mtr10Δ to wild-type cells. ....................................................................... 66 Figure 3. 11. Heat map of hierarchical clustering results of gene expression profiles from translational DEGs in mtr10Δ cells versus wild-type cells in fed condition............ 68 Figure 3. 12. Heat map of hierarchical clustering results of translational DEGs from msn5Δ cells compared to wild-type cells in fed condition................................................ 69 Figure 3. 13. Venn diagram of DEGs which were decreased in translation activity index in both mtr10Δ and msn5Δ cells. ...................................................................................... 70 Figure 3. 14. Amino acid biosynthesis pathways affected in tRNA trafficking mutants. ........................................................................................................................................... 72 Figure 3. 15. Expression profile of MET3 transcript among all microarray samples. .... 74 Figure 3. 16. Northern blot analysis of total RNA collected from wild-type and tRNA trafficking mutants in fed condition. ................................................................................. 75 Figure 3. 17. Growth assay of tRNA trafficking mutants on SC, SC-arginine and SCmethionine plates. ............................................................................................................. 76 Figure 3. 18. Growth assays of cells with Met22-3HA fusion protein.. ......................... 79 Figure 3. 19. Western blot analysis of endogenously 3HA- or GFP- tagged target proteins.. ............................................................................................................................ 81 Figure 3. 20. Western blot analysis of protein levels in tRNA trafficking mutants harboring recovery plasmids.. ........................................................................................... 82 xvii Figure 3. 21. Immunoblot analyses of Met3-, Arg3-, and Leu4-3HA fusion proteins in wild-type cells and dhh1Δ pat1Δ cells. ............................................................................. 83 Figure 3. 22. Heat map of hierarchical clustering of codon occurrence frequency for DEGs with decreased translation activity index in mtr10Δ cells. ..................................... 84 Figure 4. 1. PCA analysis of transcription profiles of total mRNAs from wild-type, msn5Δ, and mtr10Δ cells in fed or acute amino acid starvation condition.. ................... 101 Figure 4. 2. Volcano plots showing differential expressed genes at transcriptional level in mtr10Δ cells versus wild-type cells.. .......................................................................... 102 Figure 4. 3. Venn diagram of differentially expressed genes from mtr10Δ cells relative to wild-type cells in fed and amino acid starved conditions. .......................................... 109 Figure 4. 4. Northern blot analysis of total mRNAs collected from wild-type, mtr10Δ, and msn5Δ cells grown in fed condition. ........................................................................ 110 Figure 4. 5. Growth assay to assess the function of Mtr10 in DNA damage response. 111 Figure A. 1. Growth assay of mutants defective in tRNA export and modifications.. . 118 Figure A. 2. Growth assay of mutants defective in tRNA import and modifications... 119 Figure A. 3. Western blot analysis of Arg3-3HA in tRNA modification mutants. ...... 120 xviii CHAPTER 1 Introduction Eukaryotic cells contain a variety of lipid bilayer membrane-surrounded organelles. One of these organelles, the nucleus, is the genetic information center that controls biological activities. An important issue in eukaryotic cell biology concerns the coordination of cellular process via spatial and temporal regulation. The nucleus communicates with the cytoplasm via nuclear pores through which macromolecules flow. DNA contains the genetic information which is transcribed into RNAs and translated into proteins. Transfer RNA (tRNA) is one of three main forms of RNAs. tRNAs function by delivering amino acids to the protein synthesis machinery in the cytoplasm. Nuclear-cytoplasmic cross-talk involving tRNAs is of particular interest to our laboratory. To investigate such questions, we employ the powerful genetic tool, the budding yeast Saccharomyces cerevisiae, which has been broadly used for biological research, as our model system. 1.1 Nucleocytoplasmic transport The traffic between nucleus and cytoplasm occurs through nuclear pores. Nuclear pores are composed of nuclear pore complexes (NPCs) which are organized into channel structures that cross the inner and outer nuclear membranes. NPCs allow diffusion of 1 small molecules (< 40 kDa). In contrast, macromolecules (large proteins or ribonucleoproteins [RNPs]) contain specific sequences (nuclear export sequences, NES; or nuclear localization sequences, NLS) that are recognized and actively translocated in and out of the NPCs by transport receptors in an energy dependent manner (for review, see Aitchison and Rout, 2012). In the S. cerevisiae genome, there are 14 nuclear transport receptors that are members of karyopherin β family (Stewart, 2003). According to the direction in which they move their cargos, the receptors are referred to as exportins (nucleus to cytoplasm) or importins (cytoplasm to nucleus). The process of nuclear transport is controlled by the small GTPase, Ran. There are two forms of Ran, the GTP-bound form, which is primarily located in the nucleus, or the GDP-bound form, which is primarily located in the cytoplasm (Kuersten et al., 2001). This asymmetric distribution results from the distinct subcellular localizations of the Ran cycle regulators, the cytoplasmic GTPase-activating protein (RanGAP, encoded by RNA1) and the nuclear guanine nucleotide exchange factor (RanGEF, encoded by RCC1), which are responsible for the conversion of Ran-GTP and Ran-GDP states (top panel of Figure 1. 1) (for reviews, see Nakielny and Dreyfuss, 1999; Rodriguez et al., 2004). The exportin receptors bind Ran-GTP and the cargo in the nucleus, and the complex moves via the nuclear pore to the cytoplasm where the complex disassociates upon Ran-GTP hydrolysis to Ran-GDP by RanGAP (middle panel of Figure 1. 1). Ran-GDP returns to the nucleus by the aid of NTF2 (Ribbeck et al., 1998), and is converted to Ran-GTP form again by RanGEF. The importin receptors bind the cargo (directly or adaptor-mediated) in the cytoplasm in a Ran-GTP independent manner. The 2 importin-cargo complex translocates to the nucleus where importin encounters Ran-GTP, and this leads to the release of cargo and importin returns to the cytoplasm for next cycle (bottom panel of Figure 1. 1) (for review, see Chook and Süel, 2011). As the molecular size of tRNA is ~25 kDa which is theoretically able to leave the nucleus by diffusion, its export requires the Ran pathway and the functions of karyopherin. 3 Figure 1. 1. Ran Cycle and Ran-dependent nuclear export and import. 4 1.2 tRNA biology in yeast 1.2.1 The life of tRNA: biogenesis, maturation, and turnover tRNA biogenesis tRNA is an essential adaptor molecule composed 73 to 93 nucleotides that fold into Lshaped tertiary structures. tRNAs function to translate genetic information encoded in mRNAs to proteins by delivering the appropriate amino acids during translation at the ribosome. In yeast tRNA transcription occurs in the nucleolus, a sub-compartment in the nucleus (Thompson et al., 2003). tRNA genes (tDNAs) are transcribed by RNA polymerase III (Pol III), which recognizes the promoter sequences within tRNA genes (Dieci et al., 2007). Transcriptional control of tRNAs is regulated by Pol III via the master regulator, Maf1 (for reviews, see Ciesla and Boguta, 2008; Willis and Moir, 2007). tRNA processing Since the initial tRNA transcripts contain extra nucleotides at 5’and 3’ ends and some of tRNA precursors have intervening sequences (IVS, or introns) between exons, the tRNA transcripts need to be processed to mature sized molecules. tRNAs are also highly modified in post-transcriptional steps. The processing events, including removal of 5’ and 3’ termini, CCA addition, and intron splicing, generally occur in a specific order. The leader sequence is cleaved from pre-tRNAs to generate a mature 5’ end by the ribonucleoprotein endonuclease (RNase P). RNase P is composed of a RNA molecule and protein subunits (for reviews, see Esakova and Krasilnikov, 2010; Walker and Engelke, 2006). The 3’ end of pre-tRNA is processed by removal of the trailer sequence 5 and the subsequent addition of CCA nucleotides after the residue N73. The former reaction is catalyzed by the yeast endoribonuclease tRNase Z (Trz1) and exonuclease Rex1, which is balanced by the presence of Lhp1 (yeast La protein, which is a chaperon required for tRNA maturation) (for reviews, see Nakanishi and Nureki, 2005; Phizicky and Hopper, 2010). Addition of nucleotides C74, C75, and A76 is required for subsequent amino acid attachment. This reaction is catalyzed by the enzyme tRNA nucleotidyltransferase, encoded by CCA1 in yeast. CCA1 is an essential gene, but there is a temperature-sensitive mutant of this gene, cca1-1. Cells with this mutation cca1-1 have a phenotype that is associated with incompletely-processed tRNAs. Interestingly, yeast Cca1 is an isozyme which has three ATGs. The multiple transcripts result in the production of three proteins that are differentially distributed to nucleus, cytoplasm, and mitochondria (Chen et al., 1992b; Wolfe et al., 1994). Among the total of 286 nuclear tRNA genes in yeast, 59 encode introns. Introns are located between the 3’ end of the anticodon after nucleotide 37 and the loop 1. Intron lengths vary from 14 to 60 nucleotides. The intron-containing tRNAs belong to 10 different tRNA families (for reviews, see Hopper and Phizicky, 2003; Abelson et al., 1998; Hani and Feldmann, 1998). tRNA intron removal is catalyzed by the heterotetrameric splicing endonuclease complex (SEN). Although the SEN complex is essential, a previous study demonstrated that the intron of one tRNA, tRNATrp(CCA), is unnecessary in translation and cell growth (Mori et al., 2011). In yeast, the pre-tRNA splicing reaction includes three steps. First, the intron-containing tRNAs are cleaved at both ends of the intron by SEN. Second, the subsequently generated two halves are 6 ligated by tRNA ligase Trl1 and 2’-phosphate is formed at the splice junction. Removal of the 2’ phosphate at the splice junction is catalyzed by the 2’ phosphotransferase Tpt1 (for review, see Hopper and Phizicky, 2003). SEN is functionally conserved from yeast to vertebrates; however, its subcellular localization is not. The work from Yoshihisa showed that the yeast SEN is located at mitochondria surface (Yoshihisa et al., 2003), but vertebrate SEN is in the nucleus (Paushkin et al., 2004). A recent study reveals that SEN possesses a novel function unrelated to splicing that requires all four subunits to be located on the mitochondria (Dhungel and Hopper, 2012). tRNA modification To date, more than 100 post-transcriptional modified nucleotides have been described in tRNAs (Cantara et al., 2011). Although the biological function(s) of many tRNA modifications are not completely understood, some of them have been shown to play a role in stabilizing tRNA tertiary structure, facilitating tRNA folding, and preventing translation frameshift to increase the accuracy of protein synthesis (for review, see Gustilo et al., 2008). tRNA modification enzymes locate at different subcellular compartments, including nucleolus, nucleoplasm, the inner nuclear membrane, the cytoplasm, and mitochondria (for review, see Hopper and Phizicky, 2003). 7 Figure 1. 2. Summary of tRNA processing pathways. tRNA modification pathways are not included. 8 tRNA aminoacylation Before binding to ribosomes, tRNAs are aminoacylated, or charged, at their 3’ end of the CCA sequence. Charging is catalyzed by aminoacyl tRNA synthetases (aaRS). There are 20 aaRSs, each corresponding to a single amino acid. Since there are multiple tRNAs for given amino acids, one enzyme generally recognize more than one tRNA substrate. Although the classical view was that tRNA aminoacylation solely occurs in the cytoplasm, several aminoacyl tRNA synthetases are located in the nucleus in vertebrates and yeast, and tRNA charging has been shown to occur in the nucleus (Lund and Dahlberg, 1998; Sarkar and Hopper, 1998). tRNA turnover/degradation As tRNAs are long-lived relative to mRNAs and are recycled for multiple rounds of tRNA charging, it is not surprising that there are at least two quality control pathways that function in repairing or degrading damaged tRNAs (for reviews, see Hopper et al., 2010; Phizicky and Hopper, 2010; Thompson and Parker, 2009). The yeast TRAMP complex, that includes Trf4/5, Air1/Air2, and Mtr4, was the first discovered mechanism to monitor the integrity of pre-tRNA during tRNA biogenesis (Kadaba et al., 2004; Kadaba et al., 2006). This pathway recognizes hypomodified nuclear tRNAs, such as tRNAiMet at m1A58, caused by mutation of Trm6. Substrates of the TRAMP pathway are poly-adenylated and subjected to degradation from their 3’ end by the nuclear exosome. Other studies demonstrated that additional tRNA substrates, for example, pre-tRNA transcript (LaCava et al., 2005), tRNAAla with a structural defect (Vanacova et al., 2005), 9 U6 small nuclear RNA, or 3’ truncated 5S rRNA (Kadaba et al., 2006), are also checked by this pathway. In contrast to the TRAMP pathway which is restricted to the nucleus, the second surveillance mechanism, the rapid tRNA decay (RTD) pathway, is located in both the nucleus and the cytoplasm. The RTD pathway acts on mature tRNAs from the 5’ end (for review, see Phizicky and Hopper, 2010). Cells with defective Trm8 and Trm4, which normally modify tRNAs at m7G46 and m5C49, respectively, exhibit elevated temperaturesensitive growth, caused by the rapid degradation of hypomodified tRNAVal(AAC) by the RTD pathway (Alexandrov et al., 2006). Furthermore, tRNA substrates with different modification defects are also RTD targets (Chernyakov et al., 2008). The same study reported a similar rate of tRNA substrate degradation after addition of transcription inhibitor thiolutin, showing that the degradation targets mature tRNAs, rather than pretRNAs (Chernyakov et al., 2008). Two 5’−3’ exonucleases were identified to function in the RTD pathway by suppressor analysis, Rat1, which locates in the nucleus, and Xrn1, which locates in the cytoplasm. Both of them require Met22 activity since a MET22-null mutant leads to accumulation of the substrate pAp that subsequently inhibits the activities of Xrn1 and Rat1 (Chernyakov et al., 2008; Dichtl et al., 1997). 1.2.2 tRNA subcellular dynamics Transfer RNAs must exit the nucleus to the cytoplasm to function in protein synthesis; therefore, it has been thought for a long time that tRNAs move one-way into the cytoplasm. However, surprisingly, it was subsequently learned that cytoplasmic mature 10 tRNAs re-visit the nucleus, a process referred to as retrograde tRNA nuclear import (Shaheen and Hopper, 2005; Takano et al., 2005). Imported cytoplasmic tRNAs return back to the cytoplasm via a process termed tRNA re-export (Shaheen and Hopper, 2005; Takano et al., 2005; Whitney et al., 2007). This retrograde process is conserved from yeast to vertebrates (Barhoom et al., 2011; Shaheen et al., 2007; Zaitseva et al., 2006). In vertebrate cells, tRNA nuclear export is mediated by Exportin-t (Exp-t), a member of the Ran-binding karyopherins (Arts et al., 1998a; Kutay et al., 1998). The orthologue of Exp-t in fungi, Xpo-t or Los1, is so far the only known tRNA exporter able to bind to end-processed intron-containing tRNAs in a Ran-GTP dependent manner. The crystal structure showed that Xpo-t from S. pombe binds tRNAs directly by contacting the acceptor arm, TψC and D loops (Cook et al., 2009). The data support the early observations that Exp-t prefers tRNAs with 5’ and 3’ matured ends but has no preference for intron-containing or intron-less tRNAs. (Arts et al., 1998b; Lipowsky et al., 1999; Lund and Dahlberg, 1998). In yeast, los1Δ cells are viable, suggesting that there are tRNA export pathway(s) in addition to Los1 (Hurt et al., 1987). Exp-5 (Msn5 in yeast) was identified as a tRNA exporter and functions in parallel with Los1. However, there must exist additional export pathway because los1Δ msn5Δ mutant is still viable (Takano et al., 2005). Exportin-5 (Exp-5), or yeast orthologue Msn5, functions in exporting double strand miRNA in mammalian cells (Bohnsack et al., 2004; Gwizdek et al., 2003; Lund et al., 2004; Yi et al., 2003) and plants (Bollman et al., 2003). In yeast, Msn5 exports phosphorylated nuclear proteins to cytoplasm such as transcription factors Pho4 and Far1 11 (Blondel et al., 1999; Kaffman et al., 1998). The role of Msn5 in tRNA subcellular dynamics is solely in re-exporting back to the cytoplasm, as msn5Δ cells accumulate mature tRNAs but not precursor tRNAs in the nucleus (Murthi et al., 2010). Since it has been shown that the nuclear import process is constitutive, but tRNA nuclear accumulation is dependent on nutrient availability, it has been proposed that tRNA nuclear-cytoplasmic trafficking is most likely regulated at the re-export process in response to nutrient availability (Murthi et al., 2010). Spliced tRNAs are found to accumulate in the nucleus upon nutrient starvation (Hurto et al., 2007; Shaheen and Hopper, 2005; Takano et al., 2005; Whitney et al., 2007). One β-importin member, Mtr10 (vertebrate TRN-SR2), is responsible for tRNA nuclear accumulation upon nutrient deprivation and for the constitutive retrograde nuclear transport, directly or indirectly (Murthi et al., 2010). Relative to other members, there is limited understanding of the mechanism by which Mtr10 functions in tRNA nucleocytoplasmic dynamics. It is not known if Mtr10 binds tRNAs directly or adaptor mediated, and it is conceivable that Mtr10 functions in tethering tRNAs in the nucleus, therefore further studies on Mtr10 are necessary. 1.3 Amino acid stress responses Yeast cells rapidly respond to changing environments by reprogramming gene expression. Several studies have described the changes in global gene expression when the availability of nutrient, such as amino acids, carbon source, or phosphate, are limited (Castelli et al., 2011; Gasch et al., 2000; Kuhn et al., 2001; Natarajan et al., 2001; Ogawa 12 et al., 2000). Amino acid starvation is of particular interest in this work. It is known that amino acid deprivation results in global translation repression, which has been thought to inhibit general protein synthesis by which cells can save energy. Moreover, amino acid stress also causes induced translation of the central positive transcription regulator Gcn4. Gcn4 is a transcription activator that stimulates the transcription of several amino acid biosynthesis pathways upon amino acid deprivation (for review, see Hinnebusch, 2005). Signaling pathways involved in translation regulation for amino acid-starved cells are well-studied (for reviews, see Gasch and Werner-Washburne, 2002; Wilson and Roach, 2002). Uncharged tRNAs increase when cells are deprived for amino acids which subsequently activates the protein kinase Gcn2. Gcn2 phosphorylates the translation initiation factor eIF2-α, which prevents the regulatory circuit of eIF2 and initiation complex formation. Phosphorylation of eIF2α subsequent leads to general translation inhibition. Interestingly, although the global translation is inhibited, starvation of amino acids, purine, or glucose, activates the translation of Gcn4 mRNA (for review, see Hinnebusch, 2005). Gcn4 is regulated at multiple levels. There are four small uORFs in the upstream sequence of the Gcn4 transcript. They serve to negatively regulate translation of Gcn4. Under nutrient starvation, the ribosome bypasses the uORFs and thus derepresses the translation of Gcn4 in a Gcn2-dependent manner (Hinnebusch, 1997). Moreover, there are also several reports demonstrating that the derepression of Gcn4 can also be regulated in a Gcn2-independent fashion (for review, see Hinnebusch, 2005). Finally, Gcn4 can be regulated at the level of protein stability. The half-life of Gcn4 is 2 minutes when cells are grown in rich media, however, in cells deprived of amino acids 13 the half-life of Gcn4 increased to 10 minutes. Phosphorylation of Gcn4 by cyclin dependent kinase Pho85 is required before it is degraded by Rad6 and Cdc34 (Kornitzer et al., 1994; Shemer et al., 2002). 1.4 Microarray analysis 1.4.1 Microarray general applications and advanced usage for translational profiling Microarray technology is a very powerful approach to examine gene expression levels on large or genome-wide scale. Arrays are created by thousands of probes robotically spotted and immobilized on microscope glass slides or silicon chips (Affymetrix). The probes can be DNA, cDNA, or oligonucleotides, and they are designed to hybridize to specific fluorophore-labeled targets in samples. Thus the abundance of targets can be detected and quantified based on fluorescence signal levels. There are two basic types of arrays for expression analysis, dual channel (twocolor) and single channel (one-color). Two-color microarray (also called cDNA or spotted array) was first developed by the Patrick Brown laboratory (Schena et al., 1995); these arrays are generated by spotting cDNA probes on to the solid supports. The experiment design typically compares two biological samples, for example, reference (control) versus drug-treatment, and is thus measuring the relative abundance of targets between samples. The two samples are usually labeled with two different dyes, such as Cy3 (green) and Cy5 (red), and then mixed and hybridized to one array. During image analysis, the ratio of the two dye intensities is calculated for each spot and thus represents 14 the relative amount of each target. The advantages of this technology are that the probes can be customized for only a set of genes of interest (for example, immune-response related genes, Kim et al., 2005) and the cost is relatively inexpensive. Single-channel arrays, or high-density arrays, contain short-oligonucleotides (for example, in Affymetrix GeneChip®, probes are 25 nucleotides in length), these are designed to match part of each single dye-labeled target and thus are referred as “probeset”. These oligonucleotides are produced directly on silicon chips by photolithography technique (Fodor et al., 1991). A large number of different probesets can be synthesized on a single chip, (for example, the Affymetrix Human Genome U133 Plus 2.0 Array is comprised of more than 54,000 probesets, http://www.affymetrix.com/) and thus it is also called high-density array. This array generally provides “absolute” intensity of each labeled target, so two chips are needed to compare the expression level of a gene from two biological conditions. To date, many microarray experiments are utilized for various applications. Different experimental designs provide different kinds of information. In the last few years, a growing number of studies combined technology of translation status and microarray analysis to analyze translation profiles in a genome-wide scale under many different conditions, such as environmental stress (Arava et al., 2003; Blais et al., 2004; Kuhn et al., 2001; Shenton et al., 2006; Smirnova et al., 2005). The sucrose gradient sedimentation of mRNA is affected by its ribosome occupancy and that is an indicator of translational status as ribosome binding (initiation), moving along the coding region (elongation), and falling off (termination). Thus, the distribution pattern of ribosome 15 becomes an important indicator for analyzing global translation profiles or translational status of genes of interest. Different organization of ribosome association on an mRNA can reflect its translation efficiency; for example, high occupancy on polysomes of a particular transcript usually correlates with high efficiency of translation and may generate high levels of protein products (Tuller et al., 2007). In this study, Affymetrix GeneChip arrays (Yeast Genome 2.0 arrays) are used, so the experiment and data analysis are specific to these arrays. RNA was extracted from the biological samples, converted to cDNA, and then labeled with Biotin. The labeled cDNAs were hybridized to the surface of the arrays, followed by washing and processing. The arrays were scanned by a laser to generate the digital image files and the raw intensity data were obtained. 1.4.2 Data analysis After scanning the hybridized array an image file is created (.dat). Software provided by Affymetrix, GeneChip Operating System (GCOS), computes the signal intensity from image file and subsequently creates .cel files. The files need further processing before being subjected to statistic analysis and selection of differentially expressed genes. Although commercial software from microarray companies is costly, there are other open-source programs in various platforms that are freely available. These programs include Significance Analysis of Microarrays (SAM, which can be executed in Excel, Tusher et al., 2001), http://www.bioconductor.org/), Bioconductor and (Gentleman GenePattern 16 (Reich et al., 2004, et al., 2006, http://www.broadinstitute.org/cancer/software/genepattern/). In this project all of the microarray data analyses were executed by employing the Bioconductor in R environment or the GenePattern web-interface platform. Bioconductor uses the R statistical programming language and provides various packages not only for microarray analysis, but also for sequence data and other high-throughput data assays (554 software packages while writing the thesis). There are also several diagnostic plots that can be utilized to visualize the data set and to assess data quality for each step of data analysis. For example, box plots and principle component analysis (PCA) can be applied as a general assessment of microarray data. Scatter plots and volcano plots provide comparison results of two samples. GenePattern is another useful platform for genomic analysis, which offers many tools to analyze large-scale data in web-based interface or the user’s computer. There is no standard procedure for analyzing microarray data. Therefore, each step of analyzing decisions leads to non-identical results. Generally, the pipeline of microarray data analysis includes pre-processing of raw data, identification of significant differential genes, visualization, and interpretation. Each of the steps is briefly described below and the details are written with R language in Appendix P. Pre-processing On Affymetrix GeneChips, there are many probes (short oligonucleotides), referred as a probeset, designed for targeting a single gene (Figure 1. 3). The .cel file contains signal intensity data of each probe. Therefore, it is necessary to process the intensities of those 17 probesets into gene expression values for each target. This step is called normalization. In addition, there are other factors that must be considered such as technical artifacts. Thus the purpose of normalization is not only to process expression values, but also to remove these artificial effects and to correct backgrounds among chips. In this procedure, the cdf (chip description file) file, which is provided from Affymetrix, is required because it offers the identity of the probe information. There are several methods to perform normalization. One of them, Robust Multiarray Average (RMA), available in a package called “affy” from Bioconductor, is currently most commonly used for microarray normalization (Bolstad et al., 2003; Irizarry et al., 2003). RMA is an algorithm and the assumption of RMA is that usually only a small percentage of genes are altered in different conditions. Thus RMA uses quantile normalization and median polish (an alternative way to get median value) to calculate the expression values by using all probesets (the log2-converted perfect-match intensities) across all chips. In this way it provides sensitive detection of differential expressed genes with statistical tests. Identification of differential expressed genes The main purpose of microarray analysis is to identify the genes which are differentially expressed in different conditions or treatments. Numerous statistical strategies are designed and developed for selecting those expression-changed genes; most of them use variants of t-test to take sample size and variations into account. Since the traditional multiple tests is not appropriate for the genome-wide scale data, several alternative methods for permutation of p-value are developed for more accurate selection of genes of 18 interest, for example, the Bonferroni correction or false discovery rate (FDR). In this project, the package of “Linear Models for Microarray data analysis” (limma) in Bioconductor was used to select significant differentially expressed genes (DEGs). Employing limma analysis, DEGs can be ranked by both fold-change and Bonferroniadjusted p-value (Smyth, 2004). Once the list of significant differentially expressed genes is created, the data can be used for a variety of following analyses, such as hierarchical clustering and pattern recognition. Figure 1. 3. Probe and probesets. Hierarchical clustering When significant differentially expressed genes are selected, the next challenge is how to focus on real genes of interest. One approach is clustering, which is widely used in microarray analysis. Based on various algorithms, the produced clusters can have 19 differences in properties. For example, hierarchical, k-means, and self-organizing map (SOM), etc., are common methods for analyzing microarray data (Sherlock, 2000). The hierarchical clustering model is generated based on distance connectivity. Briefly, the algorithm calculates the distance of each gene expression value and returns groups of genes or samples with similar expression profiles. Additionally, there are various ways to measure distances and to cluster groups. Moreover, the input files can be normalized expression values from genome-wide array without statistical analysis or only the expression profiles from differential expression genes. Therefore, each choice in measurement and in clustering would give different results. In this project, the hierarchical analysis and the visualization of the results were generated from GenePattern. GO term enrichment analysis and FunCat Other approaches to focus on genes of interest are by employing the Gene Ontology (GO) enrichment analysis bin/amigo/term_enrichment?session_id=) (http://amigo.geneontology.org/cgior MIPS FunCat analysis (Functional Catalogue, http://mips.helmholtz-muenchen.de/proj/funcatDB/search_main_frame.html) (Ruepp et al., 2004). GO is a biological database that provides information of gene products within defined categories (for review, see Rhee et al., 2008). FunCat provides such information in a different way. Using similar concepts like clustering, GO term enrichment analysis and FunCat group genes associated with similar phenotypes and compare the occurrence frequency with genome frequency by statistical analysis. In this work both tools are performed to gain more insight of the gene list. 20 1.5 Aim of this study The major goal of this study is to investigate the potential role(s) of tRNA nuclearcytoplasmic trafficking in translation regulation, globally and specifically, by analyzing the polysome profiles and by employing microarrays of mRNAs that are or are not associated with polysomes. I compared the translation profiles by microarray analysis of mRNAs isolated from wild-type cells to mRNAs isolated from cells defective in tRNA nuclear import (mtr10Δ) or re-export (msn5Δ) grown in fed condition or acutely starved by removal of all amino acids (Chapter 3). In addition, since little is known about the importer Mtr10, a minor aim of this work was to analyze transcriptional profiles of mtr10Δ cells, which could provide insights into its cellular function(s) (Chapter 4). 21 CHAPTER 2 Materials and Methods 2.1 Yeast growth conditions and strain construction Most Saccharomyces cerevisiae strains employed in this study were derived from haploid BY4742 (MATα his3Δ1 leu2Δ0 lys2Δ0 ura3Δ0, from Open Biosystems (Winzeler et al., 1999) or are otherwise indicated. All of the strains are listed in Table 2. 1. 2.1.1 Media and growth conditions Yeast cells were grown in either YEPD (yeast extract/peptone medium with 2 % glucose/dextrose) or SC medium (Synthetic Complete defined medium; Difco yeast nitrogen base without amino acids, supplemented with amino acids, adenine [0.04 g/L], uracil [0.04 g/L], and 2 % glucose as carbon source) at 23°C or 30°C in an air shaker with speed of 220 rpm. Amino acids in SC medium included L-arginine (0.02 g/L), Laspartic acid (0.1 g/L), L-glutamic acid (0.1 g/L), L-histidine (0.02 g/L), L-isoleucine (0.06 g/L), L-leucine (0.06 g/L), L-lysine (0.03 g/L), L-methionine (0.02 g/L), Lphenylalanine (0.05 g/L), L-serine (0.375 g/L), L-threonine (0.2 g/L), L-tryptophan (0.02 22 g/L), L-tyrosine (0.05 g/L), L-valine (0.15 g/L). For amino acid starvation experiments, pre-warmed SC medium without addition of amino acids was used. 2.1.2 Yeast strain construction Oligonucleotides DNA oligonucleotides were synthesized by SIGMA-ALDRICH. The sequences of all employed oligonucleotides are in Table 2. 2. High efficiency yeast transformation Double deletion and genomic C-terminal 3HA- or GFP-tag strains were generated by introducing PCR fragments with selection marker using a Lithium acetate (LiOAc) transformation method (Chen et al., 1992a; Longtine et al., 1998). Yeast cells were grown overnight in 50 ml of rich YEPD media until OD600 reached 1-3. Cells were harvested, washed twice with 1X TE/LiOAc solution (1M LiOAc and TE buffer containing 10 mM Tris-HCl, pH7.4, and 1mM of EDTA, pH8.0, served as 10X stocks), and then resuspended in 0.7-1 ml of 1X TE/LiOAc solution. For each reaction, 100 µl of yeast competent cells were then mixed with 10 µl of PCR products of replacement cassettes after ethanol-precipitated or column-purified, and 10 µl of denatured salmon-sperm DNA, followed with addition of 650 µl transformation solution (ratio of 60% PEG: LiOAc: TE: H2O as 7:1:1:1). The mixtures were incubated at 23°C or 30°C in an air shaker for 1 hr, followed by heat-shock in a 42°C water bath for 15 min. Cells were then collected by centrifugation for 10 seconds and then plated onto selective solid media and incubated for 23 2 or more days. Colonies were then picked and grown on new selective solid media for a second round of selection. For drug-selective media, such as Hygromycin B, cells were grown in YEPD media for 6 hr or overnight prior to plating. Strain construction To select deletion strains, yeast cells were mixed with an appropriate DNA replacement cassette, natMX4, hphMX4 (Goldstein and McCusker, 1999), or Kleuyveromyces lactis LEU2 (Gueldener et al., 2002), followed by high efficiency transformation, and grown on solid media containing the appropriate drugs: YEPD + clonNAT (100 mg/L; Werner BioAgents, Jena, Germany), YEPD + hygromycin B (300 mg/L, Calbiochem, La Jolla, CA), or SC-Leu, respectively. For construction of genomic C-terminal 3HA- or GFP-tag strains, yeast cells were incubated with knock-in cassette (pFA6a-3HA-His3 or pFA6aGFP-His3) and were grown and selected on the SC-his solid media. Preparation of yeast genomic DNA For deletion strains, yeast cells were confirmed by PCR analysis of genomic DNA with primers complementary with an upstream region of the target gene and with selection marker. To confirm C-terminal tagging, primers complementary with middle of the particular coding region and with epitope were used. To extract genomic DNA, one colony (or more than one) with size of approximately 1 mm was picked from plates, transferred into a microfuge tube containing 100 µl of zymolyase digestion solution, and incubated at 37°C for 30 min. 1/10 volume of 10% SDS and 2 volumes of 24 phenol/chloroform were added followed by vigorously shaking. After centrifugation, supernatants were transferred and the nucleic acids were precipitated with 2 volumes of ethanol and 1/10 volume of 3M sodium acetate in -80°C. DNA pellets were then collected, washed by 70 % ethanol, and dissolved in 20 µl of sterile H2O. 2.1.3 Growth assay Yeast cells were cultured overnight in YEPD medium prior to the growth assay. Cells were adjusted to same starting OD600 and then 10-fold serial dilutions in sterile H2O were prepared. 5 µl aliquots of each dilution was spotted on the indicated solid media, such as SC, SC-met, or SC-arg, followed by incubated at indicated temperatures for 2 or more days and then photographed. 2.2 Polysome profiles and RNA isolation For microarray analysis, wild-type (BY4742), msn5Δ, and mtr10Δ cells were used; for polysome profile analysis, additional tRNA retrograde mutants, los1Δ and los1Δ msn5Δ, were used. 2.2.1 Polysome profiles One liter of cell culture was grown in synthetic complete media at 30°C in an air shaker to early logarithmic phase (A600 around 0.35). The culture was divided into two bottles; after centrifuging (~ 2,700 g, room temperature, 5 min) the cultures were resuspended with 500 ml of 30°C pre-warmed SC and SC-aa (SC medium lacks all amino acids) 25 medium, respectively, and the two cultures were returned to the shaker for 30 min. Cycloheximide was added to each culture to final concentration 100 µg/ml for two minutes before cultures were harvested. Cultures were cooled on ice less than 5 min, and cells were harvested and washed with 20 ml of cold freshly-prepared lysis buffer (20 mM Tris-HCl with pH 8.0, 140 mM KCl, 1.5 mM MgCl2, 0.5 mM dithiothreitol, 100 µg/ml cycloheximide, 1 mg/ml heparin). After the culture was washed, the cells were transferred to 15 ml conical tubes and then resuspended in 750 µl of lysis buffer. ~350 µl of cold acid-washed glass beads (0.4-0.6 mm diameter, Thomas Scientific) were added and then cells were disrupted by vortexing in cold room with 10 cycles of 30 seconds burst and 30 seconds ice. The clear crude lysate was obtained after centrifugation at ~ 12,600 g for 5 min at 4°C. 5 µl of lysate was transferred into 995 µl of water for A260 measurement to estimate the RNA concentration. Twenty A260 units of cell extract with the addition of Triton X-100 to final concentration 1 % were loaded onto an 11ml gradient (Polyallomer tube, Beckman Coulter) composed with five layers of 10%, 20%, 30%, 40%, and 50% sucrose (w/v) in gradient buffer (20 mM Tris-HCl with pH 8.0, 140 mM KCl, 5 mM MgCl2, 0.5 mM dithiothreitol, 100 µg/ml cycloheximide, 0.5 mg/ml heparin). Gradients were centrifuged in SW41 Ti rotor (Beckman Coulter) at 39,000 rpm for 2.5 hr in 4°C. Gradient was fractionated into 14 tubes (~0.9 ml) using ISCO UA-6 collection system (Brandel, Gaithersburg, MD) with UV254 recorder. During the process of fractionation polysome profiles were monitored and traced continuously. 26 2.2.2 Quantification of polysome profiles The scanned polysome profiles were analyzed using an area measurement tool in Adobe Acrobat 8 Professional version. Statistical significant analyses were conducted in Microsoft Excel with two tail t test. Figure 2. 1 shows how each peak was selected in the polysome profiles in fed or starved conditions. In fed condition, the areas of 40S and 80S peaks are measured separately. This sum of 40S and 80S area represented non-translating pool. In amino acid starved cells, there was a large increase in the non-translating region and usually above the range of UV254 detector. Therefore, the area of 40S + 80S was estimated by projecting (the shading region of right panel in Figure 2. 1). Profiles shown in Figure 2. 1 are representatives from more than three repeated experiments. Fed Starved Figure 2. 1. Illustration of quantification method. 27 2.2.3 RNA extraction For microarray experiments, polysomal RNA isolation procedures were similar as previously described (Arava et al., 2003; Coller and Parker, 2005). Briefly, fractions 5 to 8 (non-polysomal RNA samples) and 10 to 14 (polysomal RNA samples) were pooled together in 30 ml glass tubes. Fractions were then mixed with 2-fold volumes (2V) of 8M Guanidine-HCl; the RNAs were then precipitated in 100% ethanol. The nucleic acid pellets were obtained by centrifugation using JA-17 rotor (~13,800 g, for 20 min at 4°C). The pellets were washed once with 85% ethanol, dissolved in 400 µl of TE buffer, and then transferred into 2 ml microfuge tube following by phenol/chloroform extraction. After centrifugation (~ 12,600 g for 10 min at 4°C), the supernatant was transferred to a new 1.5 ml microfuge tube and 200 µl of 7.5 M LiCl (to final concentration 2.5M) was added to precipitate the RNAs after stored at -20°C overnight. RNA samples were collected at ~ 12,600 g for 20 min at 4°C and they were washed with 85% ethanol followed with dissolving in sterile H2O. To remove LiCl, RNA samples were precipitated again by ethanol/NaOAc and resuspended in 7 to 15 µl of nuclease-free water. Unfractionated RNA samples were directly extracted from the lysates for “total RNA”. Concentrations of RNA samples were measured by A260. 2.3 Microarray data analysis The quality of total, polysomal, or non-polysomal RNAs from BY4742, msn5Δ, and mtr10Δ cells grown in fed or amino acid starvation conditions were estimated using an Agilent Bioanalyzer. Microarray experiments were then performed using Affymetrix 28 GeneChip Yeast Genome 2.0 arrays according to manufacturer’s instructions by the OSU microarray shared resource (http://www.osuccc.osu.edu/microarray/). Data pre- processing and further statistical analysis were conducted using the free open-source software R (version 2.7.2 or later, R development Core Team, 2011) (http://www.rproject.org) and Bioconductor suite (Gentleman et al., 2004) supplemented with packages of Affy (Irizarry et al., 2003), Limma (linear models for microarray data) (Smyth, 2004), and others, as described in the programming codes in Appendix P. Microarray raw and normalized data set will be deposited in the GEO database. 2.3.1 Pre-processing of raw data In this project, the procedures of pre-processing included raw intensity data (.cel files) imported into R, removal of S. pombe probes, Robust Multiarray Average (RMA) normalization, and filtering outliers. After RMA normalization, expression intensities of each target were simultaneously transformed to a single log2 value, which was then utilized for calculation of the translational activity index (P/NP) using the equation log2 P – log2 NP. Distinct processed dataset from each mutant under fed or starvation were then generated (mtr10Δ versus wild-type in fed or in starvation conditions; msn5Δ versus wild-type in fed or in starvation conditions). Codes are in Appendix P. 2.3.2 Identification of significant differentially-expressed genes Each dataset was subjected to fit the linear model using the limma program according to instructions. Significant differentially expressed genes were selected by setting cutoff 29 conditions, such as log2 (fold-change) and adjusted p-value, and were highlighted in volcano plots. Codes are in Appendix P. These selected genes were then annotated and exported for further applications. In this study, hierarchical clustering, gene ontology enrichment analysis, functional categories were used. 2.3.3 Hierarchical clustering of significantly changed target transcripts When differentially-expressed targets were selected, their expression profiles from P, NP, and T, were extracted for heatmap creation. Hierarchical clustering analysis and generation of heatmaps were conducted using web-based tool in the GenePattern platform (by using the modules of HierarchicalClustering and HierarchicalClusteringViewer) (Reich et al., 2006) with the parameters of pearsoncorrelation in row distance, no column clustering, and pair-wise complete linkage. (http://www.broadinstitute.org/cancer/software/genepattern/). 2.4 Northern blot analysis Total RNA (12 µg in 6 µl) samples were denatured (for each sample, denaturing buffer contained 3 µl of 10X MOPS buffer, 5.3 µl of 37 % formaldehyde, 15 µl of de-ionized formamide, 1 µl of 1mg/ml ethidium bromide) at 65°C for 15 min, following by chilling on ice and addition of loading dye (75 % glycerol and 0.2 % bromophenol blue in RNA denature buffer). Denatured RNA samples were separated by 1.2 % agarose formaldehyde gels in 1X MOPS buffer subjected to electrophoresis at 50-80 V for approximately 5-6 hr. RNAs were then capillary transferred to Hybond N membrane (GE 30 Healthcare) using 20X SSC, following by UV cross-linking to the membranes. The blot was hybridized at 42°C with purified radiolabeled DNA probes. Blots were washed with wash buffer (2X SSC, 0.05 % SDS). The blots were then analyzed using Typhoon Trio variable Mode Imager (GE Healthcare) and ImageQuant 5.0 (Molecular Dynamics). 2.4.1 Preparation of DNA probes DNA probes (length between 300 to 800 base pair) used for Northern blots were generated by PCR with genomic DNA from wild-type cells or with plasmids from the yeast genomic tiling collection (Jones et al., 2008, OpenBiosystems). DNA probes were randomly radiolabeled with random primers (Invitrogen), α–32P-dCTP (PerkinElmer, Massachusetts, USA), dATP, dTTP, and dGTP, by DNA polymerase Klenow (Fermentas, Maryland, USA). Labeled probes were purified by eluting from G-50 column (GE Healthcare). The purified probes were then denatured in boiled water for 5 min and added into hybridization buffer. All of the oligonucleotides used in this study are listed in Table 2. 2. 2.5 Western blot analysis 2.5.1 Protein extraction Whole cell protein extracts were prepared from 15 ml cell cultures grown at 30°C to a density of 0.35-0.45 OD600. Cells were washed once with cold water and disrupted in 100 µl lysis buffer (50 mM Tris-HCl with pH7.4, 150 mM NaCl, 25 mM EDTA, 1 % Triton X-100, 0.5 % SDS, 10 mM PMSF, 1X protease inhibitor [Calbiochem, cocktail set IV]) 31 with approximately 50-70 µl of acid-washed glass beads. Cleared cell extracts were obtained after centrifugation at ~ 12,600 g for 5 min at 4°C. Protein concentrations were determined from 1 µl of extracts using the Bradford method (Bio-Rad). Protein extracts were then denatured in 4X SDS sample loading buffer (0.2 M Tris-HCl with pH6.8, 8 % SDS, 0.4 % BPB, 40 % Glycerol, 57.2 mM 2-ME) in boiling water for 5 minutes. 2.5.2 Western blot and quantification Depending on the individual endogenous target protein intracellular levels, total amounts of 5 to 20 µg of protein extracts from each strain were resolved on 10% SDS-PAGE with running buffer (0.025 M Tris, 0.192 M glycine, 0.1 % SDS). Immunoblotting was employed using a semi-dry transfer apparatus (BioRad Trans-Blot) at 15 V for 1 hr in transfer buffer (30 mM glycine, 0.037 % SDS, 20 % methanol, 48 mM Tris-HCl, pH 8.0). The blots were probed with primary antibodies in 5 % non-fat milk in 1X TBST buffer (20 mM Tris-HCl, pH 7.4, 120 mM NaCl, 0.1 % Tween-20). 3HA or GFP fusion proteins, Pgk1, or Kar2, were detected using horseradish peroxidase (HRP)-conjugated electrochemiluminescence (ECL) technology (Pierce, Rockford, IL, USA) using primary antibodies: anti-HA, (Roche, Rat monoclone, 3F10, 1:1,500), anti-GFP (Roche, mouse polyclone, 1:1,500), anti-Pgk1 (a kind gift from Paul Herman lab; rabbit polyclone, 1: 15,000), anti-Kar2 (Santa Cruz Biotechnology, rabbit polyclone, y-115, sc-33630, 1: 5,000). Quantification of protein signals were performed by ImageJ open source software (http://rsbweb.nih.gov/ij/). 32 2.6 Cis-regulatory element analyses of targets Sequences of upstream and downstream of target genes were retrieved from web-based resource (Regulatory Sequence Analysis Tools, RSAT, http://rsat.ulb.ac.be/rsat/) (Thomas-Chollier et al., 2011; Thomas-Chollier et al., 2008; van Helden, 2003). The output text file was later utilized for further analyses of codon bias usage and amino acid usage. 2.6.1 Codon bias and amino acid usage For investigating the codon bias and amino acid usage, the open reading frame sequences of translational differentially expressed genes (DEGs) in fed mtr10Δ cells were extracted from the table supplemented with the frequencies of each codon in each gene by the generous help of Dr. Thomas Begley (State University of New York in Albany) to compile the algorithm. Z score for each transcript, calculated as described previously (Begley et al., 2007). The data were imported into GenePattern for generation of hierarchical heat map. 33 Table 2. 1. Yeast strains used in this study 34 Strain Genotype Source BY4742 MATα his3Δ1 leu2Δ0 lys2Δ0 ura3Δ0, wild-type Winzeler et al., 1999 los1Δ MATα his3Δ1 leu2Δ0 lys2Δ0 ura3Δ0 los1::kanMX4 Winzeler et al., 1999 msn5Δ MATα his3Δ1 leu2Δ0 lys2Δ0 ura3Δ0 msn5::kanMX4 Winzeler et al., 1999 los1Δmsn5Δ MATα his3Δ1 leu2Δ0 lys2Δ0 ura3Δ0 los1::kanMX4 msn5::hphMX4 This study mtr10Δ MATα his3Δ1 leu2Δ0 lys2Δ0 ura3Δ0 mtr10::natMX4 This study BY4741 MATa his3Δ0 leu2Δ0 met15Δ0 ura3Δ0, wild-type Winzeler et al., 1999 dhh1ko1 MATa his3Δ0 leu2Δ0 met15Δ0 ura3Δ0 dhh1::hphMX4 Hurto and Hopper, 2011 pat1ko8 MATa his3Δ0 leu2Δ0 met15Δ0 ura3Δ0 pat1::natMX4 Hurto and Hopper, 2011 pat1dhh1-11 MATa his3Δ0 leu2Δ0 met15Δ0 ura3Δ0 pat1::natMX4 dhh1::hphMX4 Hurto and Hopper, 2011 mtr10Δ MATa his3Δ0 leu2Δ0 met15Δ0 ura3Δ0 mtr10::natMX4 Murthi et al., 2010 HYC163 MATα BY4742 MET22-3HA::His3MX6 This study HYC171 MATα los1Δ MET22-3HA::His3MX6 This study HYC179 MATα msn5Δ MET22-3HA::His3MX6 This study HYC186 MATα los1Δ msn5Δ MET22-3HA::His3MX6 This study HYC192 MATα mtr10Δ MET22-3HA::His3MX6 This study HYC261 MATα his3Δ1 leu2Δ0 lys2Δ0 ura3Δ0 MET22::natMX4 This study HYC296 MATα his3Δ1 leu2Δ0 lys2Δ0 ura3Δ0 trm8::Kl.leu2 trm4::natMX4 This study HYC348 MATα his3Δ1 leu2Δ0 lys2Δ0 ura3Δ0 trm8::Kl.leu2 trm4::kanMX4 met22:natMX4 This study HYC337 MATα his3Δ1 leu2Δ0 lys2Δ0 ura3Δ0 msn5::kanMX4 trm8::Kl.leu2 trm4::natMX4 This study HYC335 MATα his3Δ1 leu2Δ0 lys2Δ0 ura3Δ0 los1::kanMX4 trm8::Kl.leu2 trm4::natMX4 This study HYC341 MATα his3Δ1 leu2Δ0 lys2Δ0 ura3Δ0 los1::kanMX4 msn5::hphMX4 trm8::Kl.leu2 trm4::natMX4 This study 34 (continued.) Table 2. 1. (continued.) 35 Strain Genotype Source HYC375 This study HYC354 MATα his3Δ1 leu2Δ0 lys2Δ0 ura3Δ0 mtr10:: natMX4 trm8::Kl.leu2 trm4::kanMX4 MATα BY4742 MET2-3HA::His3MX6 HYC364 MATα los1Δ MET2-3HA::His3MX6 This study HYC359 MATα msn5Δ MET2-3HA::His3MX6 This study HYC367 MATα los1Δ msn5Δ MET2-3HA::His3MX6 This study HYC370 MATα mtr10Δ MET2-3HA::His3MX6 This study HYC479 MATα BY4742 MET3-3HA::His3MX6 This study HYC483 MATα los1Δ MET3-3HA::His3MX6 This study HYC486 MATα msn5Δ MET3-3HA::His3MX6 This study HYC489 MATα los1Δ msn5Δ MET3-3HA::His3MX6 This study HYC492 MATα mtr10Δ MET3-3HA::His3MX6 This study HYC495 MATα BY4742 MET14-3HA::His3MX6 This study HYC500 MATα los1Δ MET14-3HA::His3MX6 This study HYC504 MATα msn5Δ MET14-3HA::His3MX6 This study HYC508 MATα los1Δ msn5Δ MET14-3HA::His3MX6 This study HYC514 MATα mtr10Δ MET14-3HA::His3MX6 This study HYC560 MATα BY4742 CYS4-3HA::His3MX6 This study HYC565 MATα los1Δ CYS4-3HA::His3MX6 This study HYC570 MATα msn5Δ CYS4-3HA::His3MX6 This study HYC574 MATα los1Δ msn5Δ CYS4-3HA::His3MX6 This study HYC578 MATα mtr10Δ CYS4-3HA::His3MX6 This study HYC1162 MATα BY4742 ARG3-3HA::His3MX6 This study 35 This study (continued.) Table 2. 1. (continued.) 36 Strain Genotype Source HYC619 MATα los1ΔARG3-3HA::His3MX6 This study HYC1165 MATα msn5Δ ARG3-3HA::His3MX6 This study HYC630 MATα los1Δ msn5Δ ARG3-3HA::His3MX6 This study HYC1167 MATα mtr10Δ ARG3-3HA::His3MX6 This study HYC1171 MATα BY4742 ARG5,6-GFP(S65T)::His3MX6 This study HYC636 MATα los1Δ ARG5,6-GFP(S65T)::His3MX6 This study HYC1174 MATα msn5Δ ARG5,6-GFP(S65T)::His3MX6 This study HYC638 MATα los1Δ msn5Δ ARG5,6-GFP(S65T)::His3MX6 This study HYC1175 MATα mtr10Δ ARG5,6-GFP(S65T)::His3MX6 This study HYC1012 MATa his3Δ0 leu2Δ0 met15Δ0 ura3Δ0 pat1ko8 ARG3-3HA::His3MX6 This study HYC1017 MATa his3Δ0 leu2Δ0 met15Δ0 ura3Δ0 pat1ko8 MET3-3HA::His3MX6 This study HYC1041 MATa his3Δ0 leu2Δ0 met15Δ0 ura3Δ0 mtr10::natMX4 MET3-3HA::His3MX6 This study HYC1057 MATa his3Δ0 leu2Δ0 met15Δ0 ura3Δ0 mtr10::natMX4 ARG3-3HA::His3MX6 This study HYC1078, BY4741 MATa his3Δ0 leu2Δ0 met15Δ0 ura3Δ0 ARG3-3HA::His3MX6 This study HYC1082, BY4741 MATa his3Δ0 leu2Δ0 met15Δ0 ura3Δ0 MET3-3HA::His3MX6 This study HYC1086, dhh1 MATa his3Δ0 leu2Δ0 met15Δ0 ura3Δ0 ARG3-3HA::His3MX6 This study HYC1090, dhh1 MATa his3Δ0 leu2Δ0 met15Δ0 ura3Δ0 MET3-3HA::His3MX6 This study HYC1094, dhh1pat1 MATa his3Δ0 leu2Δ0 met15Δ0 ura3Δ0 ARG3-3HA::His3MX6 This study HYC1098, dhh1pat1 MATa his3Δ0 leu2Δ0 met15Δ0 ura3Δ0 MET3-3HA::His3MX6 This study HYC1185, BY4742 (HYC1171 derivative) ARG5,6-GFP(S65T)::His3MX6 [pRS426 URA3] This study HYC1186, BY4742 (HYC1171 derivative) ARG5,6-GFP(S65T)::His3MX6 [Trz1-MORF URA3] This study HYC1195, tyw1 MATα his3Δ1 leu2Δ0 lys2Δ0 ura3Δ0 tyw1::kanMX4 ARG3-3HA::His3MX6 This study HYC1201, BY4742 MATα his3Δ1 leu2Δ0 lys2Δ0 ura3Δ0 ARG3-3HA::His3MX TRM5-GFP::kanMX4 This study 36 (continued.) Table 2. 1. (continued) Strain Genotype Source HYC1210, BY4742 This study HYC1360, BY4742 MATα his3Δ1 leu2Δ0 lys2Δ0 ura3Δ0 ARG3-3HA::His3MX TRM5-NESGFP::kanMX4 MATα his3Δ1 leu2Δ0 lys2Δ0 ura3Δ0 mtr10::natMX4 ARG3-3HA::His3MX TRM5-GFP::kanMX4 MATα his3Δ1 leu2Δ0 lys2Δ0 ura3Δ0 mtr10::natMX4 ARG3-3HA::His3MX TRM5-NES-GFP::kanMX4 MATα BY4742 LEU4-3HA::His3MX6 HYC1360, los1msn5 MATα los1Δ msn5Δ LEU4-3HA::His3MX6 This study HYC1374, mtr10 MATα mtr10Δ LEU4-3HA::His3MX6 This study HYC1381, BY4741 MATa BY4741 LEU4-3HA::His3MX6 This study HYC1378, dhh1pat1 MATa dhh1Δ pat1Δ LEU4-3HA::His3MX6 This study HYC1380, dhh1 MATa dhh1Δ LEU4-3HA::His3MX6 This study HYC1383, mtr10 MATa mtr10Δ LEU4-3HA::His3MX6 This study HYC1231, mtr10 HYC1232, mtr10 37 37 This study This study This study Table 2. 2. Oligonucleotides used in this study Name Purpose Sequence (5’→3’) Source mtr10 deletion F (::nat or ::hph) mtr10 deletion R (nat or hph) TGAGGATATACTAAGGATTAACTTGCGTTATGGACAACTTACAG GTATCTGCAGGTCGACGGATCCCCGG CCATCCAACAAACGCCCTAATCCTTCCTTCCCTCACCTTTCTGTA ATCCCTTGAGTGGATCTGATATCGA WHIT47 mtr10 upstream CATCGGCACTATTCACAGGAA WHIT48 mtr10 downstream GGCATATCCTGATGTACTCAG Shaheen and Hopper, 2005 Shaheen and Hopper, 2005 Shaheen and Hopper, 2005 Shaheen and Hopper, 2005 msn5 deletion F (::nat or ::hph) msn5 deletion R (::nat or ::hph) CGTTGATTGGAAGAAAAGTAATGGATTCCACAGGCGCTTCTCAG ATTGTTACAGCTGAAGCTTCGTACGC CAGACCCACATTAAAACGCTTGATTATATGCATATTTACCGGCT GCCGACTGTGGATCTGATATCATCGA HY01 F msn5 upstream CCATTGAACAGAGGTGCTGGAG This study HY02 R msn5 downstream GGTGTATGCACGTACCTCTTAC This study HY06 F mtr10 Midstream TTGGTAGATATACCGAATGGACAG This study HY07 R mtr10 Midstream ACTCAATATCAATCGAACTCTGCAC This study HY25 F met22 DNA probe generation TCCTATTGACGGAACCAAGG This study HY26 R met22 DNA probe generation CCTCCAGCTTCATGGACAAT This study HY33 F met3 DNA probe generation TAGAGCCCACAGGGAGTTGACTG This study HY34 R met3 DNA probe generation GGTCTTGGTGGGTTGGATTCTC This study HY37 F met2 DNA probe generation CTAGTGCTTGAGTCTGGCGTGGT This study HY38 R met2 DNA probe generation GACATACGTGCAGCCGATAGTCC This study HY43 F met14 DNA probe generation CGCAAGGCATTGAGAAAACAGG This study HY45 F Met28 DNA probe generation CATTGCTGCTGGTTAGCAGTGG This study WHIT37 WHIT38 WHIT59 WHIT60 38 38 Michael Whitney Michael Whitney (continued.) Table 2. 2. continued. Purpose Sequence (5’→3’) Source HY46 R Met28 DNA probe generation CTACCTGCCCATGTTCCGTCTCT This study HY73 F Met22 C-term tag HY74 R Met22 C-term tag HY75 R confirm 3HA tag GCGCCTCAGCACTGAGCAGCG This study TPL010 R confirm GFP tagg TTGTGACCATTAACATCACCATCT Tsung-Po Lai HY83 F Trm8 upstream GCTAAGTGGGTTGGGTTTGGTTCGTACG This study HY84 R Trm8 downstream CGGAAGAGCTTGCGGCTCAAATAACCTGG This study HY85 F Trm4 deletion (::natMX4 or ::kanMX4) CACTTCCTTTTATCTACACTGTAATCCGAAGAATACACTATAAG GCTGGCTAGAAGccaggtcgacggatccccgg GCCTTTTAATAATATACATTTACTTTACAGTGGAGGGGATAAGA AACATGATAACTATCAATTAGCAGCGCcgtggatctgatatcatcgatgaattcga gc 39 Name GTTATAGCGTCAAGTGGCCCACGCGAGTTACATGACTTGGTGGT GTCTACATCATGCGATGTCATTCAGTCAAGAAACGCCcggatccccgg gttaattaa CACATACACACATATATATATGTACTCATATATTTATGTCTATCA ATAAAGTAAAATATATGTTATgaattcgagctcgtttaaac This study This study This study HY86 R Trm4 deletion (::natMX4 or ::kanMX4) HY87 F Trm4 upstream GCGGTGCTGAAGTAACGAAACCGCGCGC This study HY88 R Trm4 downstream CCGCAGGTCTTTCGCAATTTATACCTTGGGTC This study HY89 F Trm8 midstream GGCTGTGGATTCGGTGGGTTGATGATAG This study HY90 F Trm4 midstream GTCGAAGATAGCACTGAGGCGGC This study HY91 F Trm8 deletion (::Kl.leu2) HY92 R Trm8 deletion (::Kl.leu2) HY93 F Met22 deletion (::natMX4 or ::hphMX4) HY94 R Met22 deletion (::natMX4 or CAGCAGTTCCCATAGGATAAAATTTTCAAGCGTTTATTGTTAAG CTGAAAGCCAAGCCcgccagctgaagcttcgtac GTATATGTGGTAAATTGTTCTAGTTATACATCTATGTTACAATAT GGCTGGCGccttcacttgcatctatccgttc GTTAGTAAGTAAGAAGTTTAAAGACAACTCAGAAGACATCAGC ACTTTACTCTGGCATTGGAAAGAGAATTATTGGccaggtcgacggatccc cgg GTACTCATATATTTATGTCTATCAATAAAGTAAAATATATGTTAT 39 This study This study This study This study This study (continued.) Name Purpose Sequence (5’→3’) Source ::hphMX4) TTAGGCGTTTCTTGACTGAATGACATCGCcgtggatctgatatcatcgatgaatt cgagc HY95 F Met22 upstream CAAGAGGCTCCTTGAGGGCATTCAAG This study HY96 R Met22 downstream CGGTCAGAATCCTGCTGTATTTTCTTGTCTCAC This study HY97 F Met22 midstream CCCTTGGTTCCGTCAATAGGATCCAAACACC This study GGTAACGATGAAACGAAGACGTCTGTCTTTGGTGAGGCCGAAG AAGTTACCAACTGGcggatccccgggttaattaa GCTGTTTGTCTATATATAAATATAGATATAGATATACATGTACTG GTTTATCTATGTTATGCCgaattcgagctcgtttaaac This study 40 HY106 F Met2 C-term tagging HY107 R Met2 C-term tagging HY110 F Met2 upstream CCGCTTCGTTGTACAACCTACCTG This study HY111 F Arg3 DNA probe generation GGTGCCCAACCGATGTTTTTAGG This study HY112 R Arg3 DNA probe generation CGCACCGTTTCTCTCAGCAAC This study HY117 F Met3 C-term tagging HY118 R Met3 C-term tagging HY119 F Met14 C-term tagging HY120 R Met14 C-term tagging HY133 F Arg3 C-term tagging HY134 R Arg3 C-term tagging HY135 F Arg5,6 DNA probe generation CGCGCTTATTGGTGCTAGAGGT This study HY136 R Arg5,6 DNA probe generation CCTTGAAACCACTGCCCAACAT This study HY137 F Arg5,6 C-term tagging GCAAAATATTAATCTTGCTATGGGTTATGGAGAGTATGCTGGTA This study CCTATTTCACATATTGTACAAAAAGTTGTCCTATTCTTGGAAGAC AATGGCTTTTTTGTATTTcggatccccgggttaattaa GCAAATCTATTTATTTTGCGCGGTCGATCATGAATTTTGCCCTAC TTTTGAGATGGGgaattcgagctcgtttaaac CGGTTGAAGAATGTGCTACCATTATTTATGAGTACTTAATCAGT GAAAAAATCATCCGTAAGCATTTGcggatccccgggttaattaa CCTCGAATAAATATGTTCTATATTATATATATACATCTTTTATAT ATCATTAAATGTACAGgaattcgagctcgtttaaac GCAGCTATGTCTGCCATTGATATCTTTGTTAATAATAAAGGTAAT TTCAAGGACTTGAAAcggatccccgggttaattaa GCAAAATAATCAATGTATATCATTATTCATGCATCTATATCTGTA TTTATATATTAGTTATTAAGgaattcgagctcgtttaaac 40 This study This study This study This study This study This study This study (continued.) Table 2. 2. (continued.) Name Purpose Sequence (5’→3’) Source TCCCTGAAAATAAAATTATTGGTGTCcggatccccgggttaattaa CGTTATTTAATGGATATATATATATTATATATTTATATACGTTAA TGTCTCATGTGACTGAGCTGCAGgaattcgagctcgtttaaac GCCGTTATCACTGATGGCTTGAAACCAATCCATATCGTTACTAA GATGGATTTACTGAGCTACTTAGCAcggatccccgggttaattaa GAGAACGGTGCAATTGAATAGGAAAGGAATGACGGATTTTGCT TCTATGTTTGCTTTTATTTGAAGCgaattcgagctcgtttaaac HY138 R Arg5,6 C-term tagging HY144 F Cys4 C-term KI HY145 R Cys4 C-term KI HY146 F Cys4 midstream conf. GACTCTTCAAAGCTGGAGGCTTCGACG CWC15_DNA probe generation CWC15_DNA probe generation CAGACCACAGTTAGAAGCAAGAAGCGG GTCATATCGTTAATGTATCCGGACGCTG This study HY205 F LCD1_DNA probe generation CAGATCTGAAATCCTTGATAAGCGGTAGTTGGTGG This study HY206 R LCD1_DNA probe generation CCAAGTTCGGGGAGTCTTGTCCCACGTACC This study HY207 F RAD9_DNA probe generation GGGGATGCTGTTACCTTTGATGGAAATGAGTACG This study HY208 R RAD9_DNA probe generation CCAGTGTAGTGTAGGCCACCCCAACGCC This study CTCCTGTGGCTAACATACCCAATAACGCGC This study GACCCGTGGCTGCGAGCGAGTATATCCGAG This study HY203 F HY204 R 41 HY213 F HY214 R GTO3_DNA probe generation GTO3_DNA probe generation This study This study This study This study This study HY219 F ACT1 DNA probe generation CCGGTGATGGTGTTACTCACGTCGTTCC This study HY220 F ACT1 DNA probe generation GACCTTCATGGAAGATGGAGCCAAAGCGGTG This study HY252 F MTH1 DNA probe generation CCCCCTCTACTGTGCACACGCAACTAAC This study HY253 R MTH1 DNA probe generation GGTATTATGCTTGGTGGGGGCATGTCCGGTC This study HY267 F Trm5 midstream GGCGCCACTGATACCATCCAAATGCCATGGGTGC This study HY275 R Trm5 C-term KI CATCTCGTTTCTTTTTCTAATGGTCGCCTTTTTGCCTTTTTCCTCC Ohira and Suzuki, 41 (continued.) Table 2. 2. continued. Name Purpose Sequence (5’→3’) Source CACGGCgaattcgagctcgtttaaacTTTTCGACACTGGATGGCGGCGTT AGTA gcacccaccaaaccgatgtattgcgcgagtttccaacttccagctaacgtactaccaccgcttgagag acttactcttgatggtcgacggatccccggg 2011 Ohira and Suzuki, 2011 Ohira and Suzuki, 2011 42 HY276 F Trm5 C-term KI (NES-GFP) HY277 F Trm5 C-term KI (GFP) ccgatgtattgcgcgagtttccaacttccagctaacgtaggtcgacggatccccggg HY308 F Sps2 DNA probe generation CTGATACTGCATTAACCTCCATCG This study HY309 R Sps2 DNA probe generation CGCCTCTTATGATCGAACGACCGG This study HY312 F LEU4 C-term KI HY313 R LEU4 C-term KI HY314 F LEU4 midstream GCGTGCTCAAGGTGAAACTCAATGGAGAATCCC This study HY325 R LEU4 DNA probe generation TCGGAGACACCTACACCCCATTTGTAGGCC This study CTGGTGATGTGTCCATTCCATCTTTGGCCGAGGTCGAAGGTAAG AATGCTGCGGCATCTGGCTCTGCAcggatccccgggttaattaa CCGTGCTTCTAGTAATTATATGGTTAAAAAAAAAGGAAAGGAA GTAAATAAATAAGTATAGAAATAAATAGAAGCGgaattcgagctcgtttaa ac 42 This study This study CHAPTER 3 Genome-wide investigations of cellular functions for tRNA nuclear-cytoplasmic trafficking in regulation of translation 3.1 Introduction In eukaryotic cells, tRNAs are transcribed in the nucleus but function in protein synthesis in the cytoplasm. For decades, tRNAs were thought to travel only unidirectionally, nucleus to cytoplasm. However, it is now known that tRNAs actively shuttle between the nucleus and the cytoplasm (Shaheen and Hopper, 2005; Takano et al., 2005; Whitney et al., 2007), and this bi-directional movement is conserved in vertebrate cells (Barhoom et al., 2011; Shaheen et al., 2007; Zaitseva et al., 2006). Cytoplasmic mature tRNAs rapidly accumulate in the nucleus when cells are nutrient-deprived as observed by fluorescence in situ hybridization. Upon re-feeding, the nuclear previously-cytoplasmic tRNAs return to the cytoplasm to participate in translation (Hurto et al., 2007; Shaheen and Hopper, 2005; Whitney et al., 2007). 43 In the current working model, at least three nuclear transporters (all belong to βimportin family) function in tRNA nucleus-cytoplasm subcellular dynamics: Los1, Mtr10, and Msn5 (Figure 3. 1). Los1 functions in the initial export of intron-containing and intron-less tRNAs, and also in the re-export of previously cytoplasmic mature tRNAs back to the cytoplasm. Mtr10, directly or indirectly, is responsible for the retrograde movement of cytoplasmic tRNAs to the nucleus. Little is known about the biological roles of Mtr10 and one goal of this study is also to investigate the transcriptional expression profiles of mtr10Δ cells, which will be addressed in Chapter 4. Msn5 can export intron-less tRNAs and appears to function solely in tRNA re-export (Hopper et al., 2010; Murthi et al., 2010). There are still many questions regarding the tRNA trafficking process. For example, not all the exporters/importers have been identified (for reviews, see Hopper, 2006; Hopper et al., 2010; Hopper and Shaheen, 2008). Although it appears that tRNA re-export is regulated by nutrient status of the cell and the regulation is coordinated with P-body formation (Hurto and Hopper, 2011), none of the details of the molecular regulation have been uncovered. Whitney et al. demonstrated that the PKA pathway is required for the tRNA nuclear accumulation and this is independent of Gcn2 (Whitney et al., 2007). Perhaps the most important question “Why does cytoplasmic tRNA travel back to the nucleus” remains completely unresolved. Previous studies showed that tRNA accumulate in the nucleus when cells are nutrient stressed or when tRNA is damaged, implicating the potential roles of tRNA retrograde pathway in response to stress or in tRNA quality control (Feng and Hopper, 2002; Sarkar et al., 1999). 44 Based on the essential role that tRNAs serve as adaptors to deliver amino acids for polypeptides synthesis during translation in the cytoplasm, in this chapter we focus on the impacts of tRNA nuclear-cytoplasmic dynamics on translation regulation in the yeast, Saccharomyces cerevisiae. In this work, we demonstrate that tRNA nuclear-cytoplasmic movement plays a role in regulating translation of mRNAs encoding proteins involved in several amino acid biosynthetic pathways, including sulfur assimilation (methionine), arginine, and leucine. Thus, tRNA nuclear-cytoplasm trafficking is involved in cellular metabolism/physiology. 45 Figure 3. 1. Current model of tRNA subcellular movement in yeast. The proposed model of tRNA nuclear-cytoplasmic trafficking includes three steps. 1. Primary export. The primary tRNA transcripts are end-processed and exported to the cytoplasm by the tRNA exportin Los1. 2. Constitutive retrograde import. After maturation processes (including intron-splicing on the mitochondria, base modifications, and aminoacylation), Mtr10 is responsible for the import of the cytoplasmic mature tRNAs to the nucleus. 3. Re-export. The previously-cytoplasmic tRNAs are re-exported from the nucleus to the cytoplasm by the functions of Los1 and Msn5. 46 Figure 3. 2. Flowchart of experimental designs. For polysome profiles, more than three repeats were performed. For microarray samples, two biological repeats were performed and RNAs were extracted from indicated pools and hybridized to two Affymetrix chips. 47 3.2 Results This section includes results of polysome profiles analyses, microarray data of translational activity, followed by verification and sequence analyses. Figure 3. 2 illustrates the steps of experiments for polysome profile analysis and microarray sample collection. 3.2.1 Polysome profiles analysis Previous studies showed that tRNAs accumulate in the nucleus when cells are nutrient starved (Hurto et al., 2007; Shaheen and Hopper, 2005; Whitney et al., 2007). Amino acid or glucose deprivation also causes translation inhibition (Ashe et al., 2000; Holmes et al., 2004). Therefore, we proposed that cells regulate the translation machinery by controlling tRNA availability via its subcellular distribution. If it is true, we expected that the global translation should be inhibited in cells with defects in tRNA export pathways, such as in msn5Δ, los1Δ, and los1Δ msn5Δ mutants in which tRNAs constitutively accumulated in the nucleus. Furthermore, in cells which fail to accumulate tRNAs in the nucleus the global translation in nutrient starvation, such as mtr10Δ cells, translation should not be inhibited. To test the hypothesis, we analyzed the polysome profiles of wild type, mtr10Δ, msn5Δ, los1Δ, and los1Δ msn5Δ cells grown in fed or acute amino acid deprivation conditions (Figure 3. 3 and Figure 3. 4). As anticipated, in wild-type cells there was a major shift from polysomes to the non-translating region when cells were nutrient deprived (Figure 3. 3). Surprisingly, in general, the polysome profiles from cells with defects in the tRNA trafficking pathways 48 were similar in fed or amino acid starvation condition, but three differences were demonstrated. First, quantifications of polysome profiles from fed msn5Δ and los1Δ msn5Δ cells revealed that there were statistically significant increases in the ratio of P/NP relative to that in wild-type cells (NP, non-polysomal region, represented the region of 40S+80S, Figure 3. 5). Since the polysome region did not exhibit large changes, the decreases of NP region accounted for the increases in P/NP ratio. Second, the polysome region of fed mtr10Δ cells was smaller, suggesting a translation defect. Third, in starved mtr10Δ cells the NP region was smaller and polysomal region was larger, suggesting the rate of ribosome runoff in mtr10Δ cells is slower. This suggested that mtr10Δ cells were defective in repressing translation when responding to nutrient deprivation (Figure 3. 5). Possible explanations are addressed in the Discussion section. In summary, the similar polysome profiles of tRNA export mutants suggested that the global translation is not affected when tRNAs are accumulated in the nucleus, and this led us to employ microarray experiments to identify the specific targets. 3.2.2 Microarray data pre-processing To investigate whether the tRNA trafficking process regulates translation of particular mRNAs, we performed microarray analysis on the distribution of polysome-bound (P, translating) and non-polysome bound (NP, not-translating) mRNAs from wild-type, msn5Δ, and mtr10Δ cells grown in fed or acute amino acid starvation conditions (summary of the microarray experiments is cartooned in Figure 3. 6). After microarray experiments, all of the raw intensity data files (*.cel files) from fractioned (P or NP) and 49 unfractionated (total, T) RNA samples were generated. These files were imported together into the platform R with Bioconductor (Gentleman et al., 2004; R development Core Team, 2011). A diagnostic plot, principle component analysis (PCA) plot was utilized to roughly assess the similarity of microarray data. Figure 3. 7 exhibited that expression profiles of msn5Δ cells are similar to those from wild-type cells whereas mtr10Δ profiles displayed larger differences. Prior to the normalization, an optional procedure of masking S. pombe probes was performed because Affymetrix Yeast 2.0 GeneChips, contain two yeast species genomes, S. cerevisiae and S. pombe. In addition, this step can decrease background noise. As mentioned in Introduction, it is required to process the intensity data from probesets into gene expression values by normalization. We used the function of Robust Multiarray Average (RMA) (Irizarry et al., 2003) in the affy package. It is an algorithm that has been commonly used for microarray normalization (Ashe et al., 2000; Shenton et al., 2006; Smirnova et al., 2005; Vyas et al., 2009). Box plots of microarray data before and after RMA normalization were shown in Figure 3. 8 and Figure 3. 9. After a step of filtering low signals, a log2-value matrix with 36 columns (RNA samples) and 5684 rows (genes) was used for calculation of P/NP ratio and subsequently subject to statistical analysis to select differential expressed genes. 50 Figure 3. 3. Polysome profiles of wild-type, mtr10Δ, and msn5Δ cells collected from fed or 30-min amino acid starved conditions. Twenty A260 units of cell lysate were loaded and separated on 10% to 50% sucrose gradient. Polysome traces were recorded by UV254 detector during fractionation and are shown as representatives from more than five experiments. Cartoons represent the phenotypes of tRNA distribution. 51 Figure 3. 4. Polysome profiles of wild-type, los1Δ, and los1Δ msn5Δ cells collected from fed or 30-min amino acid starved condition. Twenty A260 units of cell lysate were loaded and separated on 10% to 50% sucrose gradient. Polysome traces were recorded by UV254 detector during fractionation and are shown as representatives from more than five experiments. Cartoons represent the phenotypes of tRNA distribution. 52 D. 0.6 WT B. mtr10∆ msn5∆ n=8 0 n=8 0.2 n=17 0.4 los1∆ los1∆ msn5∆ WT E. Fed 1.2 * * NP/T NP/T 1 0.8 0.6 0.4 0.2 0 WT C. mtr10∆ msn5∆ los1∆ 1.4 1.2 1 0.8 0.6 0.4 0.2 0 los1∆ msn5∆ los1∆ los1∆ msn5∆ los1∆ los1∆ msn5∆ * WT mtr10∆ msn5∆ F. Fed los1∆ Starved los1∆ msn5∆ 2 Starved 2.5 * * * 2 P/NP 1.5 P/NP mtr10∆ msn5∆ n=8 0.8 1.4 1.2 1 0.8 0.6 0.4 0.2 0 n=8 * n=5 * P/T * n=5 1 n=11 P/T 1.2 Starved n=11 Fed n=18 A. 1 0.5 1.5 1 0.5 0 0 WT mtr10∆ msn5∆ los1∆ los1∆ msn5∆ WT 53 mtr10∆ msn5∆ Figure 3. 5. Quantification of polysome profiles of tRNA trafficking mutants in fed (AC) or amino acid starved (D-F) conditions. NP represents the region of 40S and 80S in the polysome profile; P represents the region of disome and polysome in the profile; T represents the area of whole profile. *: p-value 54 0.01 55 Figure 3. 6. Summary of microarray experiments. One RNA sample was hybridized to one Affymetrix yeast 2.0 GeneChip. 55 Figure 3. 7. Three dimensional view of result from principle component analysis (PCA). Global assessment of microarray raw data set from fractionated RNA samples was evaluated by using principle component analysis (Courtesy of microarray center of OSU). WT_fed_mono represents the samples that were collected from non-polysomal bound fractions of wild-type cells grown in fed condition. One block represents one microarray experiment. Same color represents biological repeats of same condition. 56 3.2.3 Identification of significant differentially-expressed genes (DEGs) In order to understand the impact on translational status of specific mRNAs when cells are defective in tRNA nuclear import or re-export pathway, we analyzed the translation activity index of each mRNA, which was obtained from the ratio of P/NP (Smirnova et al., 2005; Vyas et al., 2009). There are several indexes which have served as indicators of translation status. Generally, higher translational index correlates with higher protein level (Greenbaum et al., 2002; Tuller et al., 2007). The tRNA retrograde process was proposed to function in regulation of translation of a subset of transcripts (Shaheen and Hopper, 2005; Whitney et al., 2007); therefore, we expected that aberrant tRNA nuclearcytoplasmic traffic could result in a re-distribution between P- and NP- bound population for such target mRNAs. Moreover, considering that Msn5 and Mtr10 also transport other protein cargos, we proposed that the transcripts that are affected commonly in mtr10Δ cells and msn5Δ cells are more likely the primary targets of tRNA trafficking. Thus, we paid more attention on the transcripts with changes in translational activity index affected in both msn5Δ and mtr10Δ cells. To identify such tRNA trafficking targets, a three-step approach was executed. First, the ratio of P/NP of each mRNA from the pre-processed matrix was calculated. Second, the translationally differentially expressed genes (DEGs) were selected separately from mtr10Δ cells or from msn5Δ cells versus wild-type cells in fed or starved condition by using the statistic method of linear model for microarray data (Limma) (Smyth, 2004). In this work, translationally DEGs were selected when adjusted p-values were smaller than 0.05 and 1.5 fold change (log2 FC = 0.6) from each mutant relative 57 to wild-type under each condition. The numbers of statistically DEGs are summarized in Table 3. 1. Detail lists are in Appendix B to Appendix E. Expression profiles from these genes were selected and imported to GenePattern website for generation of hierarchical heat map (Figure 3. 11 from fed mtr10Δ and Figure 3. 12 from fed msn5Δ). We also performed the same statistical analyses of “total” RNA samples; with this information we can check transcriptional effects by selecting potential targets with changes in P/NP ratio and no or little changes in their “total” expression levels. For details of transcriptional analyses, see Chapter 4. The translation profiling is unchanged in msn5-depleted cells under fed or amino acid starvation condition Because in msn5Δ cells tRNAs constitutively accumulate in the nucleus, we expected a large change in its translation profile. However, the numbers of translationally differentially expressed genes in msn5Δ cells were few or none in fed or starved condition (see summary table of number of translationally differentially expressed genes in Table 3. 1). This result was consistent with the data from polysome profiles (Figure 3. 5), which showed that there was no large difference between msn5Δ and wild-type cells. The unchanged translational profiles of msn5Δ cells in fed or starved condition might be due to two, or more, possibilities. First, tRNA nuclear accumulation is uncoupled with translation. Second and more likely, the Los1 and unknown tRNA re-export pathways are sufficient to compensate or maintain translation. Further interpretations are in the Discussion. 58 Normal responses to amino acid deprivation at translation level in tRNA import mutant Considering that in mtr10-null cells tRNAs fail to accumulate in the nucleus upon nutrient stress (Hurto et al., 2007; Shaheen and Hopper, 2005) and that a defect of translation repression was demonstrated (Figure 3. 3), we anticipated a subset of transcripts were translationally changed in starved mtr10Δ cells. However, very few transcripts that were translationally changed in starved mtr10Δ cells (Table 3. 1), which suggested that tRNA nuclear retrograde transport process is not regulating translation in response to nutrient availability. Furthermore, this correlates to the observation that tRNA nuclear import is a constitutive process (Murthi et al., 2010). A group of transcripts is translationally down-regulated in both mtr10Δ cells and msn5Δ cells On the basis of the opposite phenotypes in tRNA subcellular distribution between msn5Δ and mtr10Δ cells, we attempted to identify transcripts which display opposite translation expression patterns (i.e., up-regulated in mtr10Δ cells and down-regulated in msn5Δ cells, or vise versa, when compared to wild-type cells); however, such transcripts were not identified. More surprisingly, we identified a group of transcripts that was downregulated in translation (with decreased P/NP ratio) in both msn5Δ and mtr10Δ cells under fed condition. Furthermore, by employing the web-based tool which provides a survey of the functional distribution of genes of interests, MIPS functional catalogue (Ruepp et al., 2004), we found that the group of the affected transcripts was functionally 59 overrepresented in methionine/sulfur and arginine amino acid biosynthesis (Figure 3. 13). In addition, the translational indexes of transcripts encoding other amino acid biosynthesis pathways, such as LEU4, SER33, and LYS5, were also decreased in mtr10Δ cells (Figure 3. 11). Our data indicated that tRNA subcellular dynamics has a unique role in regulating translation of a subset of mRNAs encoding enzymes involved in several amino acid biogenesis pathways, including methionine, arginine, leucine, serine, and lysine (methionine, arginine, and leucine biosynthetic pathways are shown in Figure 3. 14). 3.2.4 Transcriptional control and RNA stability of target mRNAs are not affected According to the microarray expression profiles, mRNAs encoding proteins in the methionine and arginine biosynthetic pathways were translationally down-regulated in fed msn5Δ and mtr10Δ cells. The total expression levels of these targets in tRNA trafficking mutants were similar to that in wild-type cells from microarray data (for example, MET3 in (Figure 3. 15). To confirm the microarray results, we employed Northern blot analysis. The total RNAs were extracted from wild-type and tRNA trafficking mutant cells (msn5Δ, mtr10Δ, los1Δ, los1Δ msn5Δ) grown in SC media (fed condition). Since Los1 functions in tRNA primary export and re-export pathways, we anticipated that blocking nuclear export pathways by deleting both Los1 and Msn5 would further enhance the phenotypes of the potential targets, which would be shown in the next section. The Northern blots were hybridized to the DNA probes for target mRNAs, such 60 as ARG5,6, MET3, and LEU4 (Figure 3. 16). The result showed that the steady state levels of target mRNAs were not largely changed in all tested strains. In addition, another line of evidence supported that the transcriptional control of these target transcripts were not affected. When cells were amino acid deprived, translation of the master regulator Gcn4 is induced and it subsequently activates the transcription of amino acid biogenesisrelated genes. If the transcriptional control is defective, cells cannot grow in amino acid deprived conditions. Given this consideration, we performed a growth assay of tRNA nuclear-cytoplasmic shuttling mutants on SC solid media without methionine or arginine. Interestingly, those mutants were able to grow on SC-methionine or SC-arginine plates (Figure 3. 17), suggesting that the general amino acid transcriptional control is not affected. 3.2.5 Verification of microarray data by western blot analyses The Northern blot analysis and growth assays indicated that the steady state levels of mRNAs and transcriptional control are not affected in the mutants that are defective in tRNA subcellular trafficking. To confirm the results of the microarray analysis which indicated that these transcripts are changed at the translation level, we employed western analysis to determine the endogenous levels of protein targets. We introduced the 3HA or GFP epitope tags at the C-terminal of the endogenous locus to generate in-frame fusion of the Met, Arg, or Leu protein targets in wild-type cells and each tRNA trafficking mutant cells, including mtr10Δ, msn5Δ, los1Δ, and los1Δ msn5Δ. All of the fusion proteins were full length and functional by assessing cell growth on methionine (a 61 representative result is shown in Figure 3. 18) or arginine (not shown) depletion plates. We then examined the steady state levels of endogenous protein targets by immunoblots of whole-cell extracts obtained from normal SC media cultures. Correlated to their decreased translation activity indexes from microarray data, the expression levels of all tested proteins (Met2-3HA, Met3-3HA, Met14-3HA, Met223HA, Arg3-3HA, Arg5,6-GFP, and Leu4-3HA) were remarkably reduced or even not apparent in mtr10Δ cells (Figure 3. 19) while these target proteins had basal level expression in wild-type cells. In msn5Δ or los1Δ cells these protein levels were not largely changed. As mentioned earlier, it has been shown that Los1 and Msn5 function in parallel in the tRNA nuclear re-export process. Blocking tRNA re-export pathway only by deleting Msn5 would not be sufficient to cause significant result; this led to the prediction that further blocking tRNA re-export by deleting both Los1 and Msn5 would lead to significant decrease in target protein levels. In agreement with this expectation, the levels of Met, Arg, and Leu biosynthetic pathway proteins in los1Δ msn5Δ cells were synergistically decreased when compared to that in single deletion strains (Figure 3. 19). Importantly, this result also demonstrated that the growth defect of mtr10Δ cells was not accounted by the reduced level of target proteins because los1Δ msn5Δ cells grow as well as wild-type cells. Furthermore, most of tested proteins exhibited higher levels in los1Δ cells (Figure 3. 19). We reasoned that it was likely caused by up-regulated translation of Gcn4 mRNA, because it has been shown that translation of Gcn4 is up-regulated when unspliced tRNA accumulates in the nucleus in los1Δ cells (Qiu et al., 2000). 62 To further validate microarray data, Cys4-3HA fusion proteins were also tested as unchanged candidate in all tested cells (Figure 3. 19). Moreover, when we introduced the functional Los1, Msn5, or Mtr10 proteins back into individual mutants, the levels of Met3-3HA protein expression in tRNA trafficking mutants were mostly or partially recovered to basal level as in wild-type cells (Figure 3. 20). Collectively, the results from western blot analysis confirmed the microarray data that target mRNAs with lower translation activity indexed were poorly-translated. 63 X01_WT.fed.poly.CEL X02_WT.fed.poly.CEL X03_WT.fed.mono.CEL X04_WT.fed.mono.CEL X05_WT.starved.poly.CEL X06_WT.starved.poly.CEL X07_WT.starved.mono.CEL X08_WT.starved.mono.CEL X09_msn5.fed.poly.CEL X10_msn5.fed.poly.CEL X11_msn5.fed.mono.CEL X12_msn5.fed.mono.CEL X13_msn5.starved.poly.CEL X14_msn5.starved.poly.CEL X15_msn5.starved.mono.CEL X16_msn5.starved.mono.CEL X17_mtr10.fed.poly.CEL X18_mtr10.fed.poly.CEL X19_mtr10.fed.mono.CEL X20_mtr10.fed.mono.CEL X21_mtr10.starved.poly.CEL X22_mtr10.starved.poly.CEL X23_mtr10.starved.mono.CEL X24_mtr10.starved.mono.CEL X25_Wt.fed_total_c.CEL X26_Wt.fed.total.CEL X27_Wt.Starved_total_c.CEL X28_Wt.starved.total.CEL X29_msn5.fed.total.CEL X30_msn5.fed.total.CEL X31_msn5.starved.total.CEL X32_msn5.starved.total.CEL X33_mtr10.fed.total.CEL X34_mtr10.fed.total.CEL X35_mtr10.starved.total.CEL X36_mtr10.starved.total.CEL 6 8 10 log2 intensity 12 14 Raw data Figure 3. 8. Box plot of raw microarray data. 64 X01_WT.fed.poly.CEL X02_WT.fed.poly.CEL X03_WT.fed.mono.CEL X04_WT.fed.mono.CEL X05_WT.starved.poly.CEL X06_WT.starved.poly.CEL X07_WT.starved.mono.CEL X08_WT.starved.mono.CEL X09_msn5.fed.poly.CEL X10_msn5.fed.poly.CEL X11_msn5.fed.mono.CEL X12_msn5.fed.mono.CEL X13_msn5.starved.poly.CEL X14_msn5.starved.poly.CEL X15_msn5.starved.mono.CEL X16_msn5.starved.mono.CEL X17_mtr10.fed.poly.CEL X18_mtr10.fed.poly.CEL X19_mtr10.fed.mono.CEL X20_mtr10.fed.mono.CEL X21_mtr10.starved.poly.CEL X22_mtr10.starved.poly.CEL X23_mtr10.starved.mono.CEL X24_mtr10.starved.mono.CEL X25_Wt.fed_total_c.CEL X26_Wt.fed.total.CEL X27_Wt.Starved_total_c.CEL X28_Wt.starved.total.CEL X29_msn5.fed.total.CEL X30_msn5.fed.total.CEL X31_msn5.starved.total.CEL X32_msn5.starved.total.CEL X33_mtr10.fed.total.CEL X34_mtr10.fed.total.CEL X35_mtr10.starved.total.CEL X36_mtr10.starved.total.CEL log2 intensity After Remove probes and RMA normalized 14 12 10 8 6 4 Figure 3. 9. Box plot of RMA normalized microarray data. 65 4 3 0 1 2 -log10 p value 5 6 Volcano plot of limma -- P/NP_fed_mtr10 vs wt -3 -2 -1 0 1 2 3 4 diff(logFC) 3 2 0 1 -log10 p value 4 5 Volcano plot of limma -- P/NP_starved_mtr10 vs wt -2 -1 0 1 2 3 4 diff(logFC) Figure 3. 10. Volcano plot showing differential expressed genes selected by P/NP index compared from mtr10Δ to wild-type cells. Upper, fed condition; bottom, starved condition. 66 mtr10∆ vs. wild-type Fed P/NP a T Starved a Fed b Starved b Fed P/T c Starved c Up 93 2 335 605 0 102 Down 131 5 198 560 1 143 msn5∆ vs. wild-type Fed P/NP d T Starved d Fed e Starved e Fed P/T f Starved f Up 0 0 177 193 0 0 Down 12 0 119 258 0 0 Table 3. 1. Summary tables of numbers of differentially expressed genes in mtr10Δ or msn5Δ cells in fed or starved conditions. top, mtr10Δ cells vs. wild-type cells; bottom, msn5Δ cells vs. wild-type cells. P/NP, translation activity index, obtained from log2 Plog2 NP. T, total RNA, obtained from unfrationated RNA sample. P/T, index for analysis of ribosome occupancy, obtained from log2 P- log2 T. a: cutoff condition: adjusted pvalue < 0.05 with log2 FC 0.6 (Up-regulated) or condition: adjusted p-value < 0.001 with log2 FC 0.01 with log2 FC 0.6 or − 0.6; c: adjusted p-value < 0.6 or < − 0.6; d: adjusted p-value < 0.05 with log2 FC 0.6; e: adjusted p-value < 0.05 with log2 FC with log2 FC − 0.6 (Down-regulated); b: cutoff 0.6 or 0.6 or − 0.6. 67 0.6 or − − 0.6; f: adjusted p-value < 0.05 Figure 3. 11. Heat map of hierarchical clustering results of gene expression profiles from translational DEGs in mtr10Δ cells versus wild-type cells in fed condition. 68 Figure 3. 12. Heat map of hierarchical clustering results of translational DEGs from msn5Δ cells compared to wild-type cells in fed condition. 69 Figure 3. 13. Venn diagram of DEGs which were decreased in translation activity index in both mtr10Δ and msn5Δ cells. 70 71 71 Figure 3. 14. Amino acid biosynthesis pathways affected in tRNA trafficking mutants. A. Sulfur assimilation pathway. B. Arginine 72 biosynthesis pathway. C. Leucine biosynthesis pathway. Transcripts with decreased translation activity index were marked with green shadow. Targets confirmed by Western blot analyses are indicated with orange box. 72 73 73 Figure 3. 15. Expression profile of MET3 transcript among all microarray samples. The expression values were log2 and centered to the median of all values from all samples. WT_fed_poly represents RNA sample was collected from polysome-bound pool from wild-type cells in fed condition. WT_fed_mono represents RNA samples was collected from non-polysome bound pool 74 from wild-type cells in fed condition. WT_starved_poly represents RNA sample was collected from polysome-bound pool from wild-type cells in amino acid starvation condition. 74 Figure 3. 16. Northern blot analysis of total RNA collected from wild-type and tRNA trafficking mutants in fed condition. The blot was hybridized to α-32P-dCTP-labeled ARG5,6, MET3, or LEU4 DNA probes. ACT1 served as an internal loading control. 25S and 18S rRNAs were indicated in the bottom ethidium bromide-staining formaldehyde agarose gel. Each ratio was normalized to the expression values of ACT1 and compared to WT. 75 Figure 3. 17. Growth assay of tRNA trafficking mutants on SC, SC-arginine and SCmethionine plates. All tested cells were grown in rich liquid media overnight and adjusted to similar density before spotted on indicated solid plates and incubated at 30°C for two days. arg3Δ and met22Δ cells served as controls for SC-arginine and SC-methionine, respectively. 76 3.2.6 tRNA subcellular trafficking affects translation regulation of amino acid biosyntheses transcripts We attempted to study the roles of Mtr10 and Msn5, by analyzing transcripts translationally affected by both of Mtr10 and Msn5. However, it still could be argued that the reduced levels of proteins involved in the Met, Arg, and Leu biosynthetic pathways resulted from defects in protein transport, rather than tRNA subcellular dynamics. We sought to alter tRNA nuclear/cytoplasmic dynamics in an independent way. Dhh1 and Pat1 are not β-importin members and it has been shown that in dhh1Δ pat1Δ cells translation is repressed (Coller and Parker, 2005) and tRNAs fail to accumulate in the nucleus upon amino acid starvation (Hurto and Hopper, 2011). Both of these phenotypes exist for mtr10Δ cells and thus the tRNA subcellular trafficking is also aberrant in dhh1Δ pat1Δ cells. Therefore, we disrupted tRNA nuclear-cytoplasmic trafficking and assessed the levels of target proteins in dhh1Δ pat1Δ cells. As predicted, the levels of endogenous fusion proteins in Met, Arg, and Leu biosynthetic pathways were largely decreased in dhh1Δ pat1Δ cells (Figure 3. 21). Moreover, it appeared that Dhh1 played a more important role in regulating translation of the Met, Arg, and Leu biosynthetic pathway transcripts because the reduction was also observed in dhh1Δ cells, but not in pat1Δ cells. It is also worthy to notice that BY4741 and its isogenic mtr10Δ cells were utilized as isogenic controls for dhh1Δ pat1Δ cells, and their protein expression patterns were similar as which shown in BY4742 derivative strains. This result further provides the evidence that the reduced levels of enzymes in 77 Met, Arg, and Leu biosynthetic pathways can be reproduced in different mating type/background. 78 79 Figure 3. 18. Growth assays of cells with Met22-3HA fusion protein. WT, wild-type cells, as the parent strain for Met22-3HA knock-in, #6 and #12 represent candidate numbers. All of the parental strains and their derivatives were grown in liquid YEPD media overnight at 30°C. Cells were adjusted to similar density and followed by 10X serial dilution before spotting on the indicated solid media and incubated at 30°C or 37°C for two days. 79 80 80 Figure 3. 19. Western blot analysis of endogenously 3HA- or GFP- tagged target proteins. A. targets with decreased translation 81 activity index were poorly-translated in tRNA trafficking mutants. B. target with no change in translation index was also not changed in protein expression. Pgk1 and Nsp1 served as internal loading controls. Each ratio was obtained from normalizing to internal control and then comparing to WT. 81 Figure 3. 20. Western blot analysis of protein levels in tRNA trafficking mutants harboring recovery plasmids. Cells with Met3-3HA fusion proteins were transformed with pRS416 vector or MORF plasmids containing individual functional karyopherin and its promoter (courtesy from Ivy Huang). Cells harboring plasmids were then cultured in SC-uracil liquid media to early log phase. Met3-3HA proteins were determined by immunoblotting with α-HA antibody. Pgk1 served as internal loading control. 82 Figure 3. 21. Immunoblot analyses of Met3-, Arg3-, and Leu4-3HA fusion proteins in wild-type cells and dhh1Δ pat1Δ cells. Pgk1 and Kar2 served as internal loading controls. Each ratio was obtained from normalizing to internal control and then comparing to WT. 83 Figure 3. 22. Heat map of hierarchical clustering of codon occurrence frequency for DEGs with decreased translation activity index in mtr10Δ cells. 84 Taken together, our investigation of translational expression profiles by microarray analysis of the tRNA subcellular trafficking mutants showed that the translation of proteins responsible for methionine, arginine, and leucine biosynthesis pathways were defective, suggesting that tRNA trafficking process plays a role in regulating cell metabolism by influencing the translation of a subset of mRNAs. This is the first and novel connection between tRNA subcellular dynamics and cell physiological process. 3.2.7 Cis-regulatory elements analysis of tRNA trafficking targets Protein translation is generally regulated at the steps of translation initiation or the rate of elongation (Day and Tuite, 1998; McCarthy, 1998). Translation initiation is the ratelimiting step and can be regulated by the 5’ sequences or length of upstream region of the ORF, or by the secondary structure of the 5’ leader (Hinnebusch, 2005; Lawless et al., 2009; Tuller et al., 2009). We were unable to identify a common cis-acting motif from upstream and downstream sequences, except for Gcn4-responsive, methionine-, arginineresponse elements, among these primary target genes (data not shown, for reviews see Hinnebusch, 1988; Thomas and Surdin-Kerjan, 1997). Translation elongation can be affected by the number of rare codons (Letzring et al., 2010). We proposed that the target transcripts (mRNAs encode enzymes in Met, Arg, and Leu biosynthetic pathways) might have rarely used codons or amino acids, which can lead to inefficient translation and cause the reduced level of protein products. We analyzed the codon usage as well as amino acid usage for each target by comparing the 85 appearance frequencies of each codon or amino acid in targets open reading frames versus genome average occurrence. However, we were unable to document rare codon bias (Figure 3. 22) or particular amino acid usage patterns (not shown). The possible mechanisms for translation regulation will be further described in Discussion. 3.3 Discussion In this work we demonstrate that constitutive tRNA nuclear accumulation in cells with defective tRNA re-export (los1Δ, msn5Δ, and los1Δ msn5Δ) does not cause large global translation repression. There are several possible explanations. First, it is possible that the remaining cytoplasmic pool of tRNAs is sufficient to maintain the translation machinery. Second, and a non-exclusive, possibility is that the additional tRNA export pathway(s) is sufficient to transport tRNAs back to the cytoplasm, and that these pathways contribute to the generally normal translation. Finally, although we cannot rule out the impacts from additional tRNA export pathway, perhaps global translation is not coupled with tRNA subcellular distribution and, if this is the case, the mechanisms of translation inhibition and tRNA nuclear accumulation upon nutrient stress must be distinct. Thus, the translation repression upon nutrient stress is likely not resulted from nuclear accumulation of tRNA. Although there is no dramatic change, we still found a significant reduction in non-polysome region in the polysome profiles from msn5Δ and los1Δ msn5Δ cells. This result might be attributed to the combination of small translation defects of affected mRNAs (e. g. 10 % decrease of each mRNA), which are not selected in the microarray data analysis. 86 In contrast, polysomal region is decreased in mtr10Δ cells which are defective in tRNA nuclear import, suggesting a translation defect. This observation might be contributed from several factors. The mtr10Δ strain exhibits growth defect and this is likely due to impaired import of the essential mRNA export factor, Npl3, to the nucleus (Pemberton et al., 1997; Senger et al., 1998). Furthermore, Npl3 has been shown to associate with polysomes and act as a negative translation regulator, and Mtr10 is involved in the disassociation of Npl3 from polysome-bound mRNAs (Windgassen et al., 2004). Therefore, the translation defect might result from the cytoplasmic pool of Npl3 that are associated with polysomes. Taken together, the translation defect in mtr10Δ strain could be an ultimate outcome of several factors: growth defect, impaired nuclear import of Npl3, impaired tRNA nuclear import, and also likely the aberrant transport of unidentified cargos. Upon amino acid deprivation, mtr10Δ cells, which fail to accumulate tRNAs in the nucleus upon nutrient stress, are also defective in translation repression. This result is consistent with the model proposed previously (Hurto and Hopper, 2011). In this model, tRNA nuclear accumulation is a downstream event of the global translation repression in response to nutrient stress. If this is the case, the mutant with failure to repress translation upon nutrient stress should also not able to accumulate tRNA in the nucleus. In agreement with this prediction, in dhh1Δ pat1Δ cells, mutations which have been shown to block general translation repression in amino acid or glucose withdrawal condition (Coller and Parker, 2005; Holmes et al., 2004), tRNAs also fail to accumulate in the nucleus in amino acid or glucose starvation condition (Hurto and Hopper, 2011). 87 A defect of translation repression in response to amino acid starvation is observed in mtr10Δ cells, but microarray analysis of translational profiles from starved mtr10Δ cells does not correlate such change. We reason that the changes in translation of affected mRNAs are slight, so they are not selected by the cutoff conditions with large foldchange while compared to wild-type cells in the microarray analysis, but each slight change can be additive and ultimately affects the polysome profile. Surprisingly, we discover that the basal level of translation of a group of target mRNAs is reduced in cells defective in tRNA trafficking by mutations in tRNA retrograde import or re-export or independently by deleting Dhh1 and Pat1 proteins. The affected mRNAs encode enzymes involved in amino acid biosynthesis pathways, including sulfur assimilation (methionine), arginine, and leucine. In addition, Ser33 and Lys5 are also potential targets; however, the endogenous abundance of Ser33 and Lys5 proteins are too low to be assessed by western blot. According to previous results with reduced levels of enzymes in Met, Arg, and Leu biosynthetic pathways, we anticipate the protein levels of Ser33 and Lys5 should also decrease in tRNA trafficking mutants. If so, there are five amino acid biosynthesis pathways affected. Taken together, these results support that tRNA subcellular dynamics serves as a novel mechanism to regulate the translation of mRNAs involved in amino acid biosynthesis pathways. And this role also further correlates with the constitutive tRNA import process (Murthi et al., 2010). The mechanism(s) for how the affected enzymes are reduced is not clear. It appears that the reduction of these proteins is defective at translation regulation of the target mRNAs based on three observations from this work. First, the transcriptional 88 control and the steady state levels of the mRNAs involved in are generally not changed. Second, the levels of Met, Arg, and Leu biosynthetic enzymes are also decreased in dhh1Δ pat1Δ cells, in which cells mRNAs are stabilized. Third, the expression profiles of most target mRNAs showed that their decreased translation activity indexes (P/NP) are due to increasing in the NP values, suggesting a translation initiation defect of such mRNAs (Figure 3. 15). Although we do not exclude the possibility for enhanced protein turnover, it would be important to investigate whether and how the translation of the target mRNAs is affected. There is still much to learn about how signaling pathway(s) regulate tRNA nuclear-cytoplasmic dynamics, which may also provide a possible mechanism for regulating translation of target mRNAs. So far no evidence indicates that Dhh1 and Pat1 are tRNA transporters, and it was proposed that both Dhh1 and Pat1 are involved in a signal transduction pathway for tRNA subcellular trafficking (Hurto and Hopper, 2011). Considering the tRNA import process is constitutive and its impairment causes decreased basal levels of translation of mRNAs that encode Met, Arg, and Leu biosynthetic pathways, the signaling pathway must be active in normal condition. Moreover, given the nuclear accumulation of tRNAs in nutrient starvation stress, the signaling pathway must be responsive to nutrient availability and transmit the signal through Dhh1, Pat1, and Mtr10 into the nucleus. In agreement with this hypothesis, the amino acid responsive Gcn2-dependent signaling pathway is not involved in the tRNA subcellular movement because Gcn2-depleted cells do not prevent the nuclear accumulation of tRNAs (Whitney et al., 2007). The PKA pathway has been shown to be involved in the tRNA trafficking 89 (Whitney et al., 2007), but given the complexity of this pathway, as well as TOR and glucose derepression pathways, how exactly the signaling pathway(s) plays a role in tRNA nuclear-cytoplasmic trafficking needs further investigation (Whitney et al., 2007). It is unknown what the consequence(s) of reduced enzyme levels in the Met, Arg, and Leu biogenesis pathways is. One possibility is that reduction of such enzymes leads to decrease amino acid products. However, it is less likely according to several lines of evidence. First, no tRNA charging defects are observed in tRNA trafficking mutants (Whitney et al., 2007, and personal communication with Dr. Rebecca Hurto), suggesting that aminoacylation status of bulk tRNAs must be not affected in tRNA trafficking mutants. Second, uncharged tRNAs can activate the kinase Gcn2 and subsequently enhance translation of GCN4, the general regulator of amino acid biosynthesis, which activates the transcription of Gcn4-responsive genes. Evidence from transcription expression profiles of fed mtr10Δ and msn5Δ cells showed that Gcn4-responsive genes are not induced (data not shown). This result correlates to previous observation that aminoacylation status of tRNAs are not affected when tRNA nuclear-cytoplasmic trafficking is defective (Whitney et al., 2007). We still cannot completely rule out this possibility that the productions of amino acids, including Met, Arg, Leu, or more, production are reduced in the tRNA trafficking mutants and this reduction is complemented by amino acids from media. It is also likely that tRNA nuclear-cytoplasmic trafficking functions in maintaining the basal levels of these target enzymes to regulate intermediates of the Met, Arg, and Leu biosynthetic pathways, not to regulate the amino acid production per se. If 90 it is true, we can predict the intracellular levels of specific substrates of these target enzymes may be accumulated and/or enzyme products decreased in the Met, Arg, and Leu biosynthetic pathways in tRNA trafficking mutants. Since several amino acid biogenesis-related enzymes can be feedback regulated by their products (Cherest et al., 1973, for reviews, see Hinnebusch, 1988; Thomas and Surdin-Kerjan, 1997), it seems also possible that tRNA trafficking defects lead to changes in metabolic flux and that subsequently causes accumulation of amino acids. The excess amino acid products further inhibit the rate-limiting enzyme activities without affecting aminoacylation levels of tRNAs. If it is true, it will also be interesting to learn why sulfur assimilation and arginine biosynthesis pathways are repressed by regulating multiple proteins of the pathways, whereas the affected Leu, Ser, and Lys biosynthetic pathway proteins function at first step of each pathway (leucine biosynthesis pathway see Figure 3. 14, serine and lysine pathways are not shown). Therefore, extensive analysis of amino acid intermediates and metabolome in tRNA trafficking mutants may shed light on understanding the physiological functions of tRNA subcellular dynamics. How these specific mRNA targets are commonly regulated at translation level by tRNA nuclear-cytoplasmic trafficking? It is not clear. We are unable to identify common cis-regulatory elements among the ORFs within these down-regulated translating mRNAs by analyzing codon bias or amino acid usage bias. Moreover, other than Gcn4responsive elements, methionine-responsive element, and arginine-responsive element, we are not able to find other common regulatory motifs among the 5’ UTR and 3’ UTR regions from all of the target mRNAs. These results led us to propose that perhaps these 91 targets are regulated by secondary structure close to AUG or by uORF, which serves as negative translation regulator. Importantly, a previous study characterizing MET2 transcript provides a hint in which MET2 is implicated to be regulated at posttranscriptional level and there are two possible stem-loop-stem structures from upstream region to open reading frame (-9 to +61 and -80 to -44, Baroni et al., 1986; Forlani et al., 1991). Whether and how MET2 transcript is regulated via the secondary structure is still unknown. A recent publication has been shown that proteins synthesis can be initiated by non-canonical uORF in mammalian cells (Starck et al., 2012). Thus, investigation of secondary structure near start codon as well as identification of possible uORF (canonical and non-canonical) among the upstream sequences of these targets might provide possible regulatory mechanisms. Although we discovered a novel function for tRNA subcellular trafficking in cellular metabolism, its other potential functions are not excluded. For example, tRNA nuclear retrograde import might function in transporting damaged or hypomodified tRNAs back to the nucleus for quality control/proofreading. It has been shown that two tRNA surveillance systems exist in yeast cells, the TRAMP pathway, which locates in the nucleus, and the RTD pathway, which locates in the nucleus and the cytoplasm. (Alexandrov et al., 2006; Chernyakov et al., 2008; Kadaba et al., 2006). Since tRNA nuclear import is constitutive, transporting damaged cytoplasmic tRNAs for repair or turnover further supports this observation. Moreover, in trm8Δ trm4Δ cells tRNAVal(AAC) is hypomodified and degraded by the RTD pathway, which was proposed to cause the temperature sensitive growth at 37°C (Chernyakov et al., 2008). When we introduced 92 los1Δ into trm8Δ trm4Δ cells, we found this triple mutant is even more sensitive when grown at 30°C (Appendix A, Figure A. 1). We reason that it is likely because tRNA substrates are retained in the nucleus in cells lacking Los1, and this subsequently leads to increasing tRNA substrates degraded by endonucleases. A recent study showed that the tRNA nuclear-cytoplasmic trafficking is required for wybutosine (yW) base modification of tRNAPhe at position 37 (Ohira and Suzuki, 2011). G37 of tRNAPhe must be imported to the nucleus and modified by nuclear Trm5, and the product m1G37 of tRNAPhe needs to return cytoplasm for next modification to form yW. The yW formation is catalyzed by the four cytoplasmic enzymes, Tyw1-4, and is important to prevent translation frameshift (Kalhor et al., 2005; Noma et al., 2006). Therefore, interrupting tRNA nuclear import or re-export would generate the hypomodified tRNAPhe , which subsequently increase translation frameshift, and that might be the reason for reduced levels of proteins involved in Met, Arg, and Leu biosynthetic pathways in cells defective in tRNA nuclear import or export. And if this is the case, mislocation of the nuclear Trm5 to the cytoplasm may prevent the reduction. Furthermore, in tyw1Δ cells the levels of proteins responsible for Met, Arg, and Leu biosynthetic pathways should also decrease. However, our result revealed that mislocation of the nuclear Trm5 to the cytoplasm in mtr10Δ cells does not rescue the reduced protein levels (Appendix A, Figure A. 3). Also, Arg3-3HA (Appendix A, Figure A. 3) or Met3-3HA (data not shown) protein levels are unchanged in tyw1Δ cells. Therefore, hypomodified tRNAPhe is not the reason for down-regulation of translating amino acid biosynthetic mRNAs. However, there are other modifications whose 93 pathways are not yet discovered. This might also follow the tRNA nuclear-cytoplasmic trafficking and contribute to the regulation on the translation of the target mRNAs. It is also possible that the tRNA nuclear import functions for nuclear translation. Whether nuclear translation does occur in the eukaryotic cells has been argued for a long time. Recently it has been reported that nuclear translation occurs in the mammalian cells (David et al., 2012; Reid and Nicchitta, 2012). Others argued that their data do not support the nuclear translation (Dahlberg and Lund, 2012). Even if nuclear translation occurs in the mammalian cells, it may or may not occur in yeast. Thus, tRNA import may still leave a novel function in the nucleus. We seek to provide addition information about the cellular function of tRNA nucleocytoplasmic trafficking. For example, we would like to extend the analysis of cisregulatory elements by identifying possible uORFs or secondary structures in the upstream region of target mRNAs. Moreover, the mRNAs encode Met, Arg, and Leu biosynthetic pathways should be associated with the non-polysomal region in the Northern blot analysis of samples collected from tRNA trafficking mutants. Furthermore, since there is no large change in microarray analysis of msn5Δ cells and Los1 also functions in tRNA re-export, therefore, we expect a larger change in los1Δ msn5Δ cells by microarray analysis. There are more questions remaining unanswered. How do cells sense tRNA intracellular distribution? Are nuclear accumulated tRNAs functional? If so, what is the physiological role(s) of previously-cytoplasmic nuclear tRNA during nutrient starvation? The roles of tRNA nuclear-cytoplasmic trafficking in quality control and the newly 94 discovered regulation/maintenance of amino acid biosynthesis are still not able to explain why and how tRNAs are retained in the nucleus upon nutrient stress? More studies on how exporters are regulated during nutrient stress and which signaling pathway(s) is involved will be important to answer these questions. 95 CHAPTER 4 Transcriptional analysis of mtr10Δ cells 4.1 Introduction Mtr10 was first discovered in a genome-wide screen for polyadenylated mRNAs accumulation in the nucleus at 37°C. Thus, Mtr10 was suggested to function in mRNA nuclear export (Kadowaki et al., 1994). Later, it was reported to be a member of the karyopherin family that functions in nuclear import of an essential mRNA binding protein, Npl3. Npl3 functions in nuclear export of mRNAs and large ribosomal subunits (Hackmann et al., 2011; Lee et al., 1996; Stage-Zimmermann et al., 2000; Windgassen et al., 2004). Npl3 immunoprecipitates with Mtr10 and the complex only disassociates in the presence of Ran-GTP. Npl3 shuttles between the nucleus and the cytoplasm, and thus in the absence of Mtr10 as expected for a β-importin, Npl3 mislocalizes to the cytoplasm (Pemberton et al., 1997; Senger et al., 1998). Mtr10 is also required for the nuclear import of the RNA component of the yeast telomerase, TLC1 (Ferrezuelo et al., 2002; Gallardo et al., 2008). The nucleo-cytoplasmic trafficking of TLC1 RNA is required for telomerase biogenesis (Gallardo et al., 2008; 96 Teixeira et al., 2002). In wild-type cells TLC1 RNA is predominantly in the nucleus and it is mislocalized to the entire cell in mtr10Δ cells (Ferrezuelo et al., 2002). To date, the understanding of the physiological roles of Mtr10 is limited. It was proposed that there must be more unexplored cargos which depend on Mtr10 (Senger et al., 1998). In this study, we employed a genome-wide approach to determine the direct and indirect targets of Mtr10 by comparing the gene expression level of each transcript in mtr10Δ to wild-type cells in fed or amino acid starvation condition. We demonstrated that in addition to its previously known role to import Npl3, Mtr10 also likely functions in sporulation, DNA damage response, glutathione homeostasis, metal ion homeostasis, and lipid metabolism, directly or indirectly. 4.2 Results This section includes data analysis of transcription profiles from total mRNAs of msn5Δ (section 4.2.1) and mtr10Δ (section 4.2.2 and thereafter) cells compared to wild-type cells in fed or amino acid starvation conditions. Because the transcription profiles of msn5Δ cells were as expected from the large numbers of studies of this karyopherin (Bakhrat et al., 2008; Blondel et al., 1999; Bollman et al., 2003; DeVit and Johnston, 1999; Kaffman et al., 1998; Shimada et al., 2000; Willis and Moir, 2007), the results described in this section predominately focus on the transcriptional analyses of mtr10Δ cells. The data include the microarray analyses of the transcriptional profiles, the validations of selected transcripts, and functional analyses. 97 4.2.1 Microarray analysis of transcription profiles in msn5∆ cells Since Msn5 is known to export several nuclear phosphorylated transcription factors, such as Far1 and Pho4, to the cytoplasm in fed conditions (Blondel et al., 1999; Kaffman et al., 1998), we expected that msn5Δ cells would accumulate these cargos in the nucleus and therefore their target genes would have induced transcription. After employing statistical analysis of the transcription profiles from msn5Δ cells to the profiles from wild-type cells, we used the cutoff conditions with adjusted p-value 0.05 and log2 FC 0.6 or −0.6 to select differentially expressed genes. In msn5Δ cells, 177 genes were up-regulated and 119 genes were down-regulated in the fed condition; and 193 genes were up-regulated and 258 genes were down-regulated in acute amino acid starved condition (see Table 4. 1). All of the genes with statistically significant changes in expression are listed in Appendix L to Appendix O (msn5Δ vs. wild-type in fed or amino acid starved conditions). Most of the affected mRNAs are consistent with previous studies (Blondel et al., 1999; Springer et al., 2003). For example, the transcription of several Pho4responsive genes, such as PHO84, PHO89, PHO11, and PHO5, were activated in msn5Δ cells (Appendix M). Hence, in the following sections we focus on the transcriptional profiles from mtr10Δ cells. 4.2.2 Microarray analysis of transcription profiles in mtr10∆ cells Current understanding of the cellular roles of the importin Mtr10 is largely unclear. Since the results from analysis of the transcriptional profiles of msn5Δ cells confirmed its known roles, we proposed that analysis of transcriptional profiles of mtr10Δ cells could 98 provide further insights into the cellular functions of Mtr10. In general, the transcription profiles from mtr10Δ cells exhibited a relatively larger change as roughly assessed by PCA analysis (Figure 4. 1). We next performed statistical analysis of the microarray transcriptional profiles from total RNA samples of mtr10Δ cells compared to total RNA samples isolated from wild-type cells that were propagated in fed or acute amino acid starvation condition. Because ~10% of genome was affected in mtr10Δ cells (~900 genes affected in fed and ~1400 genes affected in starved condition) when using adjusted pvalue 0.05, we used a more stringent condition to select significant differentially expressed genes to narrow down the range of genes of interests (Table 4. 1). Total numbers of 533 genes from fed mtr10Δ cells and 1165 genes from starved mtr10Δ cells were selected using the cutoff condition as adjusted p-value 0.001 and log2 FC 0.6 or −0.6 (Figure 4. 2 and Table 4. 1). More details are described below. All of the genes with statistically significant changed in expression are listed in Appendix G to Appendix J (mtr10Δ vs. wild-type). Genes with expression changed in mtr10Δ cells under fed condition There are 335 genes that were up-regulated and 198 genes that were down-regulated in cells lacking Mtr10 (Table 4. 1). To understand the functions of the differential expressed genes, we employed GO term-enrichment analysis and FunCat analysis (complete result of FunCat analysis is in Appendix F). Among the 335 up-regulated genes, the top one over-represented function of each analysis was sexual reproduction (from GO term enrichment analysis, n=28) and zygospore development (from FunCat, n=21), 99 respectively (Table 4. 2). The results suggested that Mtr10, directly or indirectly, is involved in the regulation of zygote or spore formation. Among 198 down-regulated genes, 11 encoded proteins that function in metal ion homeostasis (top one from GO term enrichment analysis) and 22 encoded proteins that function in lipid metabolism (top one from FunCat analysis) (Table 4. 2). Taken together, the transcriptional profiling of fed mtr10Δ cells revealed several potential functions of Mtr10. 100 Figure 4. 1. PCA analysis of transcription profiles of total mRNAs from wild-type, msn5Δ, and mtr10Δ cells in fed or acute amino acid starvation condition. PCA analysis provided a general view of microarray data before statistical analysis. One ball represents one microarray experiment. 101 6 4 0 2 -log10 p value 8 Volcano plot of limma -- Tf_mtr10 vs wt -6 -4 -2 0 2 4 Diff (logFC) 6 4 0 2 -log10 p value 8 Volcano plot of limma -- Ts_mtr10 vs wt -6 -4 -2 0 2 Diff (logFC) Figure 4. 2. Volcano plots showing differential expressed genes at transcriptional level in mtr10Δ cells versus wild-type cells. Upper, in fed condition; bottom, in acute amino acid starved condition. Red dots represent up-regulated genes; blue dots represent downregulated genes. Cutoff values of adjusted p-value < 0.001 with log2 FC 0.6 or are shown in dashed lines in the plots. Y axis represents log10 of original p-value. 102 − 0.6 msn5Δ vs. WT mtr10Δ vs. WT Total.Fed a Total.Starved a Total.Fed b Up 177 193 335 605 Down 119 258 198 560 Total.Starved b Table 4. 1. Numbers of significant differentially transcriptionally expressed genes from total mRNA samples of msn5Δ or mtr10Δ cells in fed or amino acid starvation conditions. a: adjusted p-value < 0.05 with log2 FC with log2 FC 0.6 or 0.6 or − 0.6; b: adjusted p-value < 0.001 − 0.6. Genes changed in mtr10-deletion cells under starved condition By identifying genes with expression changes in amino acid starved mtr10Δ cells compared to amino acid starved wild-type cells, we may be able to discover “Mtr10dependent amino acid-deprived responsive” genes. If so, the data could provide information about the possible role of Mtr10 in response to environmental stress. In the transcriptional profiles from starved mtr10Δ cells, there were 605 genes that were upregulated and 560 genes that were down-regulated (Table 4. 1). Some of the affected genes were attributed to loss of Mtr10 per se, and thus appeared in both profiles from fed and starved conditions (the common affected genes in fed and starved conditions in Figure 4. 3). We identified numerous genes with affected expression only in starved condition. The functions of up-regulated genes included cell cycle, DNA repair, nuclear mRNA splicing, and protein folding (Table 4. 3). Interestingly, among starvation-only 103 down-regulated genes via identified a large numbers of genes encoded transporters. This result suggested that Mtr10 is likely involved in regulation of transporters in response to amino acid starvation. 4.2.3 Confirmations of selected target mRNA expressions by Northern blot analysis Genome-wide studies usually provide valuable information, but often produce errors or false-positive results. Hence, it is necessary to validate every gene of interest selected from microarray expression profiles by different methods, such as Northern blot analysis or real-time RT-PCR. To confirm the microarray data analysis of the transcriptional profile from mtr10Δ cells grown in fed condition, I performed the Northern analyses. The total mRNAs were collected from wild-type, mtr10Δ, and msn5Δ cells grown in SC media. In this experiment, msn5Δ cells served as an internal negative control. At least one target gene from each functional category was chose to be validated. Most of the targets from up-regulated expression profiles in mtr10Δ cells were confirmed: MTH1 (encodes a protein involved in negative regulator of glucose signaling transduction pathway), CWC15 (encodes a protein involved in mRNA splicing), RAD9 (encodes DNA-damage checkpoint protein), and LCD1 (encodes a protein essential in DNA integrity pathway) (Figure 4. 4) were all up-regulated. The expression profiles of SNF3 (encodes membrane glucose sensor) and DAL80 (encodes negative regulator of nitrogen degradation pathway) were tested but no signals were detected, likely due to their low expressions (data not shown). SPS2, SPR3, and SPR28, encode proteins 104 expressed during sporulation, were also tested. The expression level of SPS2 gene was not significantly increased in mtr10Δ cells (Figure 4. 4). In addition, the signals of SPR3 and SPR28 were too low to be detected by Northern analysis (data not shown). We also examined the expression levels of two candidate genes, GTO3 and TPO4, which were down-regulated by microarray analyses. GTO3 encodes a glutathione transferase and TPO4 encodes a polyamine transporter. The results of Northern analysis showed that the expression level of GTO3 was dramatically reduced in mtr10Δ cells relative to wild-type or msn5Δ cells and thus confirmed the microarray data (Figure 4. 4). In contrast, the expression level of TPO4 was unchanged in mtr10Δ cells when compared to wild-type cells (data not shown). In conclusion, by employing Northern blot analysis, the expression profiles of most tested targets from microarray analyses in fed mtr10Δ cells were confirmed. Mtr10 not only imports Npl3 but also is possibly involved in regulation of glucose signaling pathway, DNA damage checkpoint pathways, and glutathione homeostasis. Although further investigations are required, the microarray analysis of transcription profiles from mtr10Δ cells provides valuable information and hints of the physiological roles of Mtr10. 4.2.4 Phenotypic analyses by growth assays Since it appeared that the DNA damage-related genes were up-regulated in mtr10Δ cells (Figure 4. 4), we proposed that the DNA damage pathway was activated in mtr10Δ cells. If so, mtr10Δ cells might be more resistant to DNA damage than wild-type cells. To test this hypothesis, we performed growth assay of wild-type, mtr10Δ, msn5Δ, los1Δ, and 105 los1Δ msn5Δ cells, on solid media containing the mutagen 5-fluorouracil (5-FU) which induces DNA base excision repair pathway (Seiple et al., 2006), or methyl methanesulfonate (MMS) which causes DNA double strand breaks (Lundin et al., 2005) (Figure 4. 5). Since it was previously shown that los1Δ cells are sensitive to 5-FU (Gustavsson and Ronne, 2008), we utilized los1Δ strain as a positive control. We also examined other tRNA nucleocytoplasmic trafficking mutants, such as msn5Δ and los1Δ msn5Δ, to test their growth in the presence of these mutagens. Contrary to the expectation, the mtr10Δ strain was not resistant to either DNA damage agent. Conversely, the mtr10Δ cells were more sensitive (Figure 4. 5). These results indicated that both DNA repair pathways were defective in the absence of Mtr10 even through RAD9 and LCD1 mRNA levels were induced (Figure 4. 4). As expected, los1Δ cells were not viable in the presence of 5-FU, msn5Δ cells grew similarly as wildtype cells. These results suggest that Los1 and Msn5 do not function in parallel in response to DNA damage. Since 5-FU activates DNA base-excision pathway (Seiple et al., 2006) and los1Δ cells are sensitive to 5-FU (Gustavsson and Ronne, 2008), we hypothesized that Los1 functions in the DNA damage response. If this is the case, los1Δ cells would be also sensitive when DNA damage response alternatively induced by MMS. However, wild-type, msn5Δ, los1Δ, and los1Δ msn5Δ strains had similar growth on the MMS-containing solid media. The data suggest that these tRNA exportin pathways function independently for the DNA double-strand damage response. 106 107 Table 4. 2. GO term enrichment analysis and FunCat analysis of genes with significant changed in mtr10Δ cells in fed condition. Significant up-regulated or down-regulated expressed genes were analyzed and grouped with their GO terms or functional categories by using web-interface tools in GO or FunCat, respectively. Bold words represent the over-represented GO term or functional categories. Numbers of genes in each term or category are noted in parentheses. The p-value was obtained by comparing the occurrence frequency to background frequency. 107 108 Table 4. 3. GO term enrichment analysis and FunCat analysis of genes with significant changed in mtr10Δ cells in acute amino acid starved condition. Significant up-regulated or down-regulated expressed genes were analyzed and grouped with their GO terms or functional categories by using web-interface tools in GO or FunCat, respectively. Bold words represent the overrepresented GO term or functional categories. Numbers of genes in each term or category are noted in parentheses. The p-value was obtained by comparing the occurrence frequency to background frequency. 108 Figure 4. 3. Venn diagram of differentially expressed genes from mtr10Δ cells relative to wild-type cells in fed and amino acid starved conditions. 109 110 Figure 4. 4. Northern blot analysis of total mRNAs collected from wild-type, mtr10Δ, and msn5Δ cells grown in fed condition. ACT1 served as internal loading control. 25S and 18S rRNAs were also serving as internal loading controls, the bottom pictures were ethidium bromide-stained agarose gels. Each ratio was obtained from normalizing to ACT1 expression value and then compared to WT. 110 Figure 4. 5. Growth assay to assess the function of Mtr10 in DNA damage response. All of the strains were grown in liquid YEPD media overnight at 30°C and were adjusted to similar density followed with 10X serial dilution before spotting on the indicated solid media. 5-FU, 5-fluorouracil, a pyrimidine analog. MMS, an agent causes DNA double strand breaks. The concentrations of 5-FU and MMS were 15 µg/ml and 0.02 %, respectively, in SC media. The cells were incubated at indicated temperatures for 2 days. 111 4.3 Discussion Here we report that in mtr10Δ cells a large number of genes (about ~10% of the genome when we used the stringent conditions with adjusted p-value < 0.001 and log2 FC or 0.6 − 0.6) have affected expression from microarray analysis. Among the up-regulated genes, those encoding zygote development and sporulation are over-represented. In Northern analysis the expression level of one of these genes, SPS2, is not significantly increased. It is likely because its expression is too low to be detected. Therefore, the potential function of Mtr10, and/or its unknown cargo(s), in the regulation of sporulation may not be exclusive. Other than sporulation-related genes, we also confirmed the enhanced expression of DNA damage check point-related genes, RAD9 and LCD1, in fed mtr10Δ cells. However, the results of growth assay indicated that the mtr10Δ cells are highly sensitive to 5-FU and MMS and do not grow in the presence of these agents. It is unknown whether up-regulated expression of RAD9 and LCD1 are direct or indirect due to loss of Mtr10, but this misregulation may contribute to the defect of DNA damage response in mtr10Δ cells. Since the studies about Rad9 and Lcd1 mainly focus on the regulation of their phosphorylation states during DNA damage signaling pathway (Andrew, 1998; Rouse and Jackson, 2000), our findings provide a potentially different level of regulation. More interestingly, previous work showed that intron-containing tRNAs are retained in the nucleus after UV-induced DNA damage due to the translocation of Los1 from the nucleus to cytoplasm (Ghavidel et al., 2007). Although the regulation of this translocation and its biological significance are unclear, this observation leads to the idea that Mtr10 112 might play a role in the DNA damage signaling transduction pathway. If so, it might be directly regulated by the translocation of Mtr10 upon DNA damage (like Los1) or through Mtr10’s function in shuttling the unknown cargo(s) that function in the DNA damage signaling pathways. Northern analysis also confirmed that induction of expression levels of MTH1, which encodes a negative regulator in the glucose signaling pathway (for review, see Santangelo, 2006). When cells are grown in glucose-containing media, MTH1 expression is repressed by the Snf1-Mig1 glucose repression pathway (Kim et al., 2006). Strikingly, MTH1 not only de-represses in mtr10Δ cells but also in msn5Δ cells. This observation indicates that the Snf1-Mig1 glucose repression pathway is not fully functional in cells with defective in tRNA nuclear-cytoplasmic traffic. It is unknown whether the accumulating MTH1 transcripts leads to increased Mth1 protein and more phosphorylated Mth1. Because it has been shown that the regulation of tRNA subcellular distribution is glucose-dependent/specific (Whitney et al., 2007), our data provides supporting evidence that there might be a connection between the regulation of glucose signaling transduction pathway and tRNA subcellular movement. For down-regulated genes, we confirmed GTO3 reduced transcript levels. GTO3 encodes a glutathione transferase. This result provides another potential function for Mtr10 in glutathione homeostasis. In addition, Mtr10 may participate in two other cellular processes: metal ion homeostasis and lipid metabolism, according to the microarray analysis. It is particularly interesting to examine the expression levels of genes related to the lipid metabolism in mtr10Δ cells, because lipid homeostasis is 113 important for maintaining cell membrane and cell shape, and previous observations shows that mtr10Δ cells have abnormal long and shield-like shape (Murthi et al., 2010; Whitney et al., 2007). Therefore, although further investigations are needed, the transcriptional profiles provide explanations for the abnormal shape of mtr10Δ mutant cells. 114 CHAPTER 5 Summary and future directions In this study the data provide no evidence for the possibility that tRNA nuclear retrograde process functions as a stress signal upon nutrient deprivation from the results of polysome profiles and microarray data of starved tRNA trafficking mutants. But we discovered a novel connection between tRNA nuclear-cytoplasmic trafficking and translation of the Met, Arg, and Leu amino acid biosynthetic mRNAs. In cells with defects of tRNA trafficking, several enzymes involved in the Met, Arg, and Leu amino acid biosynthesis pathways are poorly produced, suggesting a role of tRNA nuclearcytoplasmic trafficking in maintaining the basal levels of translation of mRNAs encoding Met, Arg, and Leu biosynthetic pathways. Future direction It is unknown what are the consequence(s) caused by the reduced levels of the Met, Arg, and Leu biosynthetic enzymes. A high-throughput analysis of amino acid intermediates in Met, Arg, and Leu biosynthesis pathways by LC/MS might provide more insights in the ultimate outcomes in amino acid metabolism when tRNA nuclearcytoplasmic traffic is defective. Additionally, it is also interesting to study by which 115 mechanism that tRNA trafficking specifically regulates the translations of mRNAs involved in Met, Arg, and Leu biosynthetic pathways. On the other hand, more work is required to confirm the microarray data for the mtr10Δ mutant. In addition, further investigations on the consequences of the affected genes will contribute a greater understanding of the biological functions of Mtr10. In particular, what is the role of Mtr10 in DNA damage response pathways? Whether or not the glucose signaling pathways are misregulated in the mutants with defects of tRNA nuclear-cytoplasmic trafficking? If so, how these two pathways are coupled? 116 APPENDIX A: Tested possibilities for mechanism of down-regulated translation of the target mRNAs involved in the Met, Arg, and Leu biosynthetic pathways This section includes two results. First, in Figure A. 1 and Figure A. 2, I employed the growth assay to determine the temperature sensitive growth of tRNA modification mutants and its derivative strain with tRNA trafficking mutants. Second, we tested the hypothesis that mislocate the nuclear Trm5 to the cytoplasm might rescue the reduced levels of Arg3-3HA in cells with defective of tRNA import process (mtr10Δ) (Figure A. 3). In addition, we assessed Arg3-3HA protein levels in tRNA yW modification mutant (tyw1Δ) (Figure A. 3). The result showed that Arg3-3HA protein level was not rescued when mislocating Trm5 into the cytoplasm in mtr10Δ cells and Arg3-3HA protein was unchanged in tyw1Δ cells. Moreover, nuclear retention tRNAs by overexpressing Trz1MORF does not lead to reduced levels of Arg5,6-GFP. All together, several lines of evidence indicated that the reduced levels of target amino acid biosynthesis pathways do not result from tRNAPhe modification defect. 117 118 Figure A. 1. Growth assay of mutants defective in tRNA export and modifications. trm8Δ trm4Δ mutants combined with defective tRNA export processes (los1Δ, msn5Δ, los1Δ msn5Δ). trm8Δ trm4Δ-derived defective tRNA export mutants. All mutants were grown in liquid media overnight and density-adjusted cells were serial-diluted on SC solid media and incubated two or more days at indicated temperature. #1, #2, and #3 represented three different candidates. 118 119 Figure A. 2. Growth assay of mutants defective in tRNA import and modifications. All mutants were grown in liquid media overnight and density-adjusted cells were serial-diluted on SC solid media and incubated two or more days at indicated temperature. #1, #2, and #3 represented three different candidates. A. trm8Δ trm4Δ mutants combined with defective tRNA export processes (los1Δ, msn5Δ, los1Δ msn5Δ). B. trm8Δ trm4Δ mutants combined with defective of tRNA import process mtr10Δ. 119 Figure A. 3. Western blot analysis of Arg3-3HA in tRNA modification mutants. A. Endogenously GFP or NES-GFP tagged Trm5 was created in Wild-type or mtr10Δ cells that harboring Arg3-3HA fusion protein. The whole cell extracts were collected from early log phase of cell culture in SC media. Kar2 is internal loading control. B. Assessment of Arg3-3HA protein levels in tyw1Δ cells. Wild-type or mtr10Δ cells served as positive and negative controls. C. Assessment of Arg5,6-GFP in cells expressing vector or Trz1-MORF plasmid before and after galactose 3-hr induction. 120 APPENDIX B: Transcripts with decreased translation activity index in fed mtr10Δ cells The transcripts with decreased P/NP ratio (cutoff condition: log2 FC 0.6 and adjusted p-value 0.05) in mtr10Δ cells while compared to wild-type cells in fed condition are listed below and ordered according to log2 (fold-change). Transcripts with decreased P/NP ratio in fed mtr10Δ cells. log2 (mtr10[P f/NP.f]/ WT[Pf/N P.f]) Symbols ORF Description SUL1 YBR294W STR3 YGL184C BDH2 YAL061W PHM6 YDR281C MET2 YNL277W MET28 YIR017C NA YGR226C AAD14 YNL331C TPK1 YJL164C MET3 YJR010W High affinity sulfate permease; sulfate uptake is mediated by specific sulfate transporters Sul1p and Sul2p, which control the concentration of endogenous activated sulfate intermediates Peroxisomal cystathionine beta‐lyase, converts cystathionine into homocysteine; may be redox regulated by Gto1p Putative medium‐chain alcohol dehydrogenase with similarity to BDH1; transcription induced by constitutively active PDR1 and PDR3 Protein of unknown function, expression is regulated by phosphate levels L‐homoserine‐O‐acetyltransferase, catalyzes the conversion of homoserine to O‐acetyl homoserine which is the first step of the methionine biosynthetic pathway Basic leucine zipper (bZIP) transcriptional activator in the Cbf1p‐ Met4p‐Met28p complex, participates in the regulation of sulfur metabolism Dubious open reading frame, unlikely to encode a protein; not conserved in closely related Saccharomyces species; overlaps significantly with a verified ORF, AMA1/YGR225W Putative aryl‐alcohol dehydrogenase with similarity to P. chrysosporium aryl‐alcohol dehydrogenase; mutational analysis has not yet revealed a physiological role cAMP‐dependent protein kinase catalytic subunit; promotes vegetative growth in response to nutrients via the Ras‐cAMP signaling pathway; inhibited by regulatory subunit Bcy1p in the absence of cAMP; partially redundant with Tpk2p and Tpk3p ATP sulfurylase, catalyzes the primary step of intracellular sulfate activation, essential for assimilatory reduction of sulfate to sulfide, involved in methionine metabolism 121 adj.p.val ‐2.8202 0.0026 ‐2.4652 0.0024 ‐2.2714 0.0037 ‐2.0514 0.0171 ‐1.8123 0.0041 ‐1.7081 0.0050 ‐1.6090 0.0408 ‐1.5827 0.0127 ‐1.5650 0.0060 ‐1.4839 0.0060 log2 (mtr10[P f/NP.f]/ WT[Pf/N P.f]) Symbols ORF Description MMP1 YLL061W ‐1.4724 0.0050 NA YHR140W High‐affinity S‐methylmethionine permease, required for utilization of S‐methylmethionine as a sulfur source; has similarity to S‐ adenosylmethionine permease Sam3p Putative integral membrane protein of unknown function ‐1.4717 0.0288 FRE5 YOR384W ‐1.4654 0.0205 NA YBR184W ‐1.4519 0.0060 MET32 YDR253C ‐1.4489 0.0096 NA YFR032C‐B ‐1.4458 0.0171 QDR2 YIL121W ‐1.4424 0.0301 COX17 YLL009C ‐1.3702 0.0459 MET14 YKL001C ‐1.3027 0.0171 ARG3 YJL088W ‐1.2977 0.0060 SAP4 YGL229C ‐1.2958 0.0216 MHT1 YLL062C ‐1.2937 0.0108 MET16 YPR167C ‐1.2880 0.0112 NA YDR366C Putative ferric reductase with similarity to Fre2p; expression induced by low iron levels; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐throughput studies Putative protein of unknown function; YBR184W is not an essential gene Zinc‐finger DNA‐binding protein, involved in transcriptional regulation of the methionine biosynthetic genes, similar to Met31p Putative protein of unknown function; identified by gene‐trapping, microarray‐based expression analysis, and genome‐wide homology searching Multidrug transporter of the major facilitator superfamily, required for resistance to quinidine, barban, cisplatin, and bleomycin; may have a role in potassium uptake Copper metallochaperone that transfers copper to Sco1p and Cox11p for eventual delivery to cytochrome c oxidase; contains twin cysteine‐x9‐cysteine motifs Adenylylsulfate kinase, required for sulfate assimilation and involved in methionine metabolism Ornithine carbamoyltransferase (carbamoylphosphate:L‐ornithine carbamoyltransferase), catalyzes the sixth step in the biosynthesis of the arginine precursor ornithine Protein required for function of the Sit4p protein phosphatase, member of a family of similar proteins that form complexes with Sit4p, including Sap155p, Sap185p, and Sap190p S‐methylmethionine‐homocysteine methyltransferase, functions along with Sam4p in the conversion of S‐adenosylmethionine (AdoMet) to methionine to control the methionine/AdoMet ratio 3'‐phosphoadenylsulfate reductase, reduces 3'‐phosphoadenylyl sulfate to adenosine‐3',5'‐bisphosphate and free sulfite using reduced thioredoxin as cosubstrate, involved in sulfate assimilation and methionine metabolism Putative protein of unknown function ‐1.2866 0.0204 AMS1 YGL156W ‐1.2697 0.0112 NA YKL069W ‐1.2514 0.0366 SEO1 YAL067C ‐1.2394 0.0293 EMI2 YDR516C ‐1.2189 0.0416 NA YBL008W‐A ‐1.2071 0.0459 NA YLR361C‐A Vacuolar alpha mannosidase, involved in free oligosaccharide (fOS) degradation; delivered to the vacuole in a novel pathway separate from the secretory pathway Methionine‐R‐sulfoxide reductase, reduces the R enantiomer of free Met‐SO, in contrast to Ycl033Cp which reduces Met‐R‐SO in a peptide linkage; has a role in protection against oxidative stress Putative permease, member of the allantoate transporter subfamily of the major facilitator superfamily; mutation confers resistance to ethionine sulfoxide Non‐essential protein of unknown function required for transcriptional induction of the early meiotic‐specific transcription factor IME1; required for sporulation; expression is regulated by glucose‐repression transcription factors Mig1/2p Putative protein of unknown function; identified by fungal homology and RT‐PCR Putative protein of unknown function ‐1.1860 0.0429 SUL2 YLR092W High affinity sulfate permease; sulfate uptake is mediated by specific sulfate transporters Sul1p and Sul2p, which control the concentration of endogenous activated sulfate intermediates ‐1.1638 0.0252 122 adj.p.val log2 (mtr10[P f/NP.f]/ WT[Pf/N P.f]) Symbols ORF Description SPC3 YLR066W MET1 YKR069W ARG7 YMR062C HXT9 YJL219W RGI1 YER067W PAU17 YLL025W NAT5 YOR253W NA YDL114W SET4 YJL105W Subunit of signal peptidase complex (Spc1p, Spc2p, Spc3p, Sec11p), which catalyzes cleavage of N‐terminal signal sequences of proteins targeted to the secretory pathway; homologous to mammalian SPC22/23 S‐adenosyl‐L‐methionine uroporphyrinogen III transmethylase, involved in the biosynthesis of siroheme, a prosthetic group used by sulfite reductase; required for sulfate assimilation and methionine biosynthesis Mitochondrial ornithine acetyltransferase, catalyzes the fifth step in arginine biosynthesis; also possesses acetylglutamate synthase activity, regenerates acetylglutamate while forming ornithine Putative hexose transporter that is nearly identical to Hxt11p, has similarity to major facilitator superfamily (MFS) transporters, expression of HXT9 is regulated by transcription factors Pdr1p and Pdr3p Protein of unknown function involved in energy metabolism under respiratory conditions; protein abundance is increased upon intracellular iron depletion Protein of unknown function, member of the seripauperin multigene family encoded mainly in subtelomeric regions; YLL025W is not an essential gene Subunit of the N‐terminal acetyltransferase NatA (Nat1p, Ard1p, Nat5p); N‐terminally acetylates many proteins, which influences multiple processes such as the cell cycle, heat‐shock resistance, mating, sporulation, and telomeric silencing Putative protein of unknown function with similarity to acyl‐carrier‐ protein reductases; YDL114W is not an essential gene Protein of unknown function, contains a SET domain NA YJL218W NA YOR387C NA YIL046W‐A BNA4 YBL098W RTT102 YGR275W IRC15 YPL017C NA YJL163C GID7 YCL039W PAU5 YFL020C adj.p.val ‐1.1282 0.0431 ‐1.1043 0.0205 ‐1.1018 0.0205 ‐1.0957 0.0454 ‐1.0825 0.0308 ‐1.0771 0.0171 ‐1.0728 0.0454 ‐1.0609 0.0108 ‐1.0592 0.0330 Putative protein of unknown function, similar to bacterial galactoside O‐acetyltransferases; induced by oleate in an OAF1/PIP2‐dependent manner; promoter contains an oleate response element consensus sequence; non‐essential gene Putative protein of unknown function; regulated by the metal‐ responsive Aft1p transcription factor; highly inducible in zinc‐ depleted conditions; localizes to the soluble fraction Putative protein of unknown function; identified by expression profiling and mass spectrometry Kynurenine 3‐mono oxygenase, required for the de novo biosynthesis of NAD from tryptophan via kynurenine; expression regulated by Hst1p; putative therapeutic target for Huntington disease Component of both the SWI/SNF and RSC chromatin remodeling complexes, suggested role in chromosome maintenance; possible weak regulator of Ty1 transposition Microtubule associated protein; regulates microtubule dynamics; required for accurate meiotic chromosome segregation; null mutant displays large budded cells due to delayed mitotic progression, increased levels of spontaneous Rad52 foci Putative protein of unknown function ‐1.0499 0.0108 ‐1.0169 0.0293 ‐1.0125 0.0293 ‐1.0108 0.0498 ‐1.0074 0.0171 ‐0.9902 0.0301 ‐0.9795 0.0330 Protein of unknown function, involved in proteasome‐dependent catabolite inactivation of fructose‐1,6‐bisphosphatase; contains six WD40 repeats; computational analysis suggests that Gid7p and Moh1p have similar functions Member of the seripauperin multigene family encoded mainly in subtelomeric regions; induced during alcoholic fermentation; ‐0.9741 0.0424 ‐0.9672 0.0301 123 Symbols NA ORF AIM19 YMR105W‐ A YIL087C MER1 YNL210W MRPL49 YJL096W AVT6 YER119C DOG2 YHR043C AGE2 YIL044C YHC3 YJL059W NA YOL162W MET10 YFR030W MET22 YOL064C NSE4 YDL105W HUL4 YJR036C IRC24 YIR036C NA YGR204C‐A GSY1 YFR015C MSW1 YDR268W BUR6 YER159C NA YOR238W log2 (mtr10[P f/NP.f]/ WT[Pf/N P.f]) Description induced by low temperature and also by anaerobic conditions; negatively regulated by oxygen and repressed by heme Putative protein of unknown function adj.p.val ‐0.9640 0.0207 Putative protein of unknown function; the authentic, non‐tagged protein is detected in purified mitochondria in high‐throughput studies; null mutant displays reduced respiratory growth Protein with RNA‐binding motifs required for meiosis‐specific mRNA splicing; required for chromosome pairing and meiotic recombination Mitochondrial ribosomal protein of the large subunit ‐0.9508 0.0365 ‐0.9436 0.0301 ‐0.9428 0.0288 Vacuolar aspartate and glutamate exporter; member of a family of seven genes (AVT1‐7) related to vesicular GABA‐glycine transporters; involved in compartmentalizing acidic amino acids in response to nitrogen starvation 2‐deoxyglucose‐6‐phosphate phosphatase, member of a family of low molecular weight phosphatases, similar to Dog1p, induced by oxidative and osmotic stress, confers 2‐deoxyglucose resistance when overexpressed ADP‐ribosylation factor (ARF) GTPase activating protein (GAP) effector, involved in Trans‐Golgi‐Network (TGN) transport; contains C2C2H2 cysteine/histidine motif Vacuolar membrane protein involved in the ATP‐dependent transport of arginine into the vacuole and possibly in balancing ion homeostasis; homolog of human CLN3 involved in Batten disease (juvenile onset neuronal ceroid lipofuscinosis) Putative protein of unknown function; member of the Dal5p subfamily of the major facilitator family Subunit alpha of assimilatory sulfite reductase, which converts sulfite into sulfide Bisphosphate‐3'‐nucleotidase, involved in salt tolerance and methionine biogenesis; dephosphorylates 3'‐phosphoadenosine‐5'‐ phosphate and 3'‐phosphoadenosine‐5'‐phosphosulfate, intermediates of the sulfate assimilation pathway Nuclear protein that plays a role in the function of the Smc5p‐ Rhc18p complex Protein with similarity to hect domain E3 ubiquitin‐protein ligases, not essential for viability Putative benzil reductase;(GFP)‐fusion protein localizes to the cytoplasm and is induced by the DNA‐damaging agent MMS; sequence similarity with short‐chain dehydrogenase/reductases; null mutant has increased spontaneous Rad52p foci Putative protein of unknown function; identified by gene‐trapping, microarray‐based expression analysis, and genome‐wide homology searching Glycogen synthase with similarity to Gsy2p, the more highly expressed yeast homolog; expression induced by glucose limitation, nitrogen starvation, environmental stress, and entry into stationary phase Mitochondrial tryptophanyl‐tRNA synthetase ‐0.9385 0.0408 ‐0.9243 0.0205 ‐0.9228 0.0366 ‐0.8925 0.0293 ‐0.8915 0.0205 ‐0.8869 0.0301 ‐0.8845 0.0215 ‐0.8792 0.0205 ‐0.8737 0.0293 ‐0.8731 0.0301 ‐0.8700 0.0288 ‐0.8694 0.0252 ‐0.8661 0.0288 ‐0.8641 0.0207 ‐0.8601 0.0171 Subunit of a heterodimeric NC2 transcription regulator complex with Ncb2p; complex binds to TBP and can repress transcription by preventing preinitiation complex assembly or stimulate activated transcription; homologous to human NC2alpha Putative protein of unknown function; green fluorescent protein 124 Symbols ORF MET17 YLR303W ECM11 YDR446W PHO80 YOL001W FMN1 YDR236C CIR2 YOR356W HOR2 YER062C MET5 YJR137C PIG1 YLR273C NA YFR017C RRF1 YHR038W RDL2 YOR286W BNA3 YJL060W XYL2 YLR070C NA YDL218W CWC15 YDR163W PAU18 YLL064C LYS5 YGL154C log2 (mtr10[P f/NP.f]/ WT[Pf/N P.f]) Description adj.p.val (GFP)‐fusion protein localizes to the cytoplasm Methionine and cysteine synthase (O‐acetyl homoserine‐O‐acetyl serine sulfhydrylase), required for sulfur amino acid synthesis Non‐essential protein apparently involved in meiosis, GFP fusion protein is present in discrete clusters in the nucleus throughout mitosis; may be involved in maintaining chromatin structure Cyclin, interacts with cyclin‐dependent kinase Pho85p; regulates the response to nutrient levels and environmental conditions, including the response to phosphate limitation and stress‐dependent calcium signaling Riboflavin kinase, phosphorylates riboflavin to form riboflavin monophosphate (FMN), which is a necessary cofactor for many enzymes; localizes to microsomes and to the mitochondrial inner membrane Putative ortholog of human electron transfer flavoprotein dehydrogenase (ETF‐dH); found in a large supramolecular complex with other mitochondrial dehydrogenases; may have a role in oxidative stress response One of two redundant DL‐glycerol‐3‐phosphatases (RHR2/GPP1 encodes the other) involved in glycerol biosynthesis; induced in response to hyperosmotic stress and oxidative stress, and during the diauxic transition Sulfite reductase beta subunit, involved in amino acid biosynthesis, transcription repressed by methionine Putative targeting subunit for the type‐1 protein phosphatase Glc7p that tethers it to the Gsy2p glycogen synthase Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm and is induced in response to the DNA‐damaging agent MMS; YFR017C is not an essential gene Mitochondrial ribosome recycling factor, essential for mitochondrial protein synthesis and for the maintenance of the respiratory function of mitochondria Protein with rhodanese activity; contains a rhodanese‐like domain similar to Rdl1p, Uba4p, Tum1p, and Ych1p; overexpression causes a cell cycle delay; null mutant displays elevated frequency of mitochondrial genome loss Kynurenine aminotransferase, catalyzes formation of kynurenic acid from kynurenine; potential Cdc28p substrate Xylitol dehydrogenase, converts xylitol to D‐xylulose; expression induced by xylose, even though this pentose sugar is not well utilized by S. cerevisiae; null mutant has cell wall defect Putative protein of unknown function; YDL218W transcription is regulated by Azf1p and induced by starvation and aerobic conditions; expression also induced in cells treated with the mycotoxin patulin Non‐essential protein involved in pre‐mRNA splicing, component of a complex containing Cef1p; has similarity to S. pombe Cwf15p Protein of unknown function, member of the seripauperin multigene family encoded mainly in subtelomeric regions; identical to Pau6p Phosphopantetheinyl transferase involved in lysine biosynthesis; converts inactive apo‐form of Lys2p (alpha‐aminoadipate reductase) into catalytically active holo‐form by posttranslational addition of phosphopantetheine 125 ‐0.8522 0.0205 ‐0.8511 0.0293 ‐0.8458 0.0439 ‐0.8445 0.0281 ‐0.8429 0.0216 ‐0.8370 0.0248 ‐0.8342 0.0295 ‐0.8249 0.0248 ‐0.8211 0.0416 ‐0.8157 0.0366 ‐0.8092 0.0354 ‐0.8022 0.0301 ‐0.7997 0.0437 ‐0.7966 0.0366 ‐0.7961 0.0301 ‐0.7936 0.0408 ‐0.7917 0.0293 log2 (mtr10[P f/NP.f]/ WT[Pf/N P.f]) Symbols ORF Description HBN1 YCL026C‐B APQ12 YIL040W OSH6 YKR003W NA YGL117W Putative protein of unknown function; similar to bacterial nitroreductases; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm and nucleus; protein becomes insoluble upon intracellular iron depletion Protein required for nuclear envelope morphology, nuclear pore complex localization, mRNA export from the nucleus; exhibits synthetic lethal genetic interactions with genes involved in lipid metabolism Member of an oxysterol‐binding protein family with overlapping, redundant functions in sterol metabolism and which collectively perform a function essential for viability; GFP‐fusion protein localizes to the cell periphery Putative protein of unknown function MSK1 YNL073W YSW1 YBR148W RSM18 YER050C ODC2 YOR222W PYK2 YOR347C MGT1 YDL200C NA YGL185C SHC1 YER096W MAM33 YIL070C RTR2 YDR066C AIM45 YPR004C SAM37 YMR060C VPS30 YPL120W Mitochondrial lysine‐tRNA synthetase, required for import of both aminoacylated and deacylated forms of tRNA(Lys) into mitochondria and for aminoacylation of mitochondrially encoded tRNA(Lys) Protein required for normal prospore membrane formation; interacts with Gip1p, which is the meiosis‐specific regulatory subunit of the Glc7p protein phosphatase; expressed specifically in spores and localizes to the prospore membrane Mitochondrial ribosomal protein of the small subunit, has similarity to E. coli S18 ribosomal protein Mitochondrial inner membrane transporter, exports 2‐oxoadipate and 2‐oxoglutarate from the mitochondrial matrix to the cytosol for use in lysine and glutamate biosynthesis and in lysine catabolism Pyruvate kinase that appears to be modulated by phosphorylation; PYK2 transcription is repressed by glucose, and Pyk2p may be active under low glycolytic flux DNA repair methyltransferase (6‐O‐methylguanine‐DNA methylase) involved in protection against DNA alkylation damage Putative protein with sequence similarity to hydroxyacid dehydrogenases; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm Sporulation‐specific activator of Chs3p (chitin synthase III), required for the synthesis of the chitosan layer of ascospores; has similarity to Skt5p, which activates Chs3p during vegetative growth; transcriptionally induced at alkaline pH Acidic protein of the mitochondrial matrix involved in oxidative phosphorylation; related to the human complement receptor gC1q‐ R Protein of unknown function with high similarity to Rtr1p; exhibits genetic interactions with Rtr1p; green fluorescent protein (GFP)‐ fusion protein localizes to the cytoplasm; YDR066C is not an essential gene Putative ortholog of mammalian electron transfer flavoprotein complex subunit ETF‐alpha; interacts with frataxin, Yfh1p; null mutant displays elevated frequency of mitochondrial genome loss; may have a role in oxidative stress response Component of the Sorting and Assembly Machinery (SAM or TOB complex) of the mitochondrial outer membrane, which binds precursors of beta‐barrel proteins and facilitates their outer membrane insertion; contributes to SAM complex stability Subunit of phosphatidylinositol (PtdIns) 3‐kinase complexes I and II; Complex I is essential in autophagy and Complex II is required for vacuolar protein sorting; ortholog of the higher eukaryotic gene Beclin 1 126 adj.p.val ‐0.7903 0.0454 ‐0.7753 0.0293 ‐0.7748 0.0301 ‐0.7739 0.0256 ‐0.7704 0.0301 ‐0.7665 0.0301 ‐0.7607 0.0478 ‐0.7406 0.0293 ‐0.7392 0.0408 ‐0.7329 0.0301 ‐0.7248 0.0400 ‐0.7160 0.0365 ‐0.7148 0.0301 ‐0.6954 0.0437 ‐0.6932 0.0293 ‐0.6908 0.0478 ‐0.6876 0.0293 log2 (mtr10[P f/NP.f]/ WT[Pf/N P.f]) Symbols ORF Description NA YLR031W Putative protein of unknown function ‐0.6863 0.0326 YSC84 YHR016C ‐0.6811 0.0324 BDH1 YAL060W ‐0.6753 0.0431 IME1 YJR094C ‐0.6740 0.0431 JEN1 YKL217W ‐0.6735 0.0449 NA YDR248C ‐0.6716 0.0416 LEU4 YNL104C ‐0.6661 0.0330 CLD1 YGR110W ‐0.6656 0.0482 RSB1 YOR049C ‐0.6587 0.0454 TMT1 YER175C ‐0.6578 0.0358 COQ4 YDR204W ‐0.6484 0.0416 SPS19 YNL202W ‐0.6481 0.0425 NA YDR336W ‐0.6439 0.0301 TUM1 YOR251C ‐0.6330 0.0366 NA YHR112C ‐0.6309 0.0416 AIM20 YIL158W Actin‐binding protein involved in bundling of actin filaments and endocytosis of actin cortical patches; activity stimulated by Las17p; contains SH3 domain similar to Rvs167p NAD‐dependent (R,R)‐butanediol dehydrogenase, catalyzes oxidation of (R,R)‐2,3‐butanediol to (3R)‐acetoin, oxidation of meso‐ butanediol to (3S)‐acetoin, and reduction of acetoin; enhances use of 2,3‐butanediol as an aerobic carbon source Master regulator of meiosis that is active only during meiotic events, activates transcription of early meiotic genes through interaction with Ume6p, degraded by the 26S proteasome following phosphorylation by Ime2p Lactate transporter, required for uptake of lactate and pyruvate; phosphorylated; expression is derepressed by transcriptional activator Cat8p during respiratory growth, and repressed in the presence of glucose, fructose, and mannose Putative protein of unknown function; sequence similarity to bacterial and human gluconokinase; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm; upregulated by deletion of the RNAP‐II associated factor, PAF1 Alpha‐isopropylmalate synthase (2‐isopropylmalate synthase); the main isozyme responsible for the first step in the leucine biosynthesis pathway Mitochondrial cardiolipin‐specific phospholipase; functions upstream of Taz1p to generate monolyso‐cardiolipin; transcription increases upon genotoxic stress; involved in restricting Ty1 transposition; has homology to mammalian CGI‐58 Suppressor of sphingoid long chain base (LCB) sensitivity of an LCB‐ lyase mutation; putative integral membrane transporter or flippase that may transport LCBs from the cytoplasmic side toward the extracytoplasmic side of the membrane Trans‐aconitate methyltransferase, cytosolic enzyme that catalyzes the methyl esterification of 3‐isopropylmalate, an intermediate of the leucine biosynthetic pathway, and trans‐aconitate, which inhibits the citric acid cycle Protein with a role in ubiquinone (Coenzyme Q) biosynthesis, possibly functioning in stabilization of Coq7p; located on the matrix face of the mitochondrial inner membrane; component of a mitochondrial ubiquinone‐synthesizing complex Peroxisomal 2,4‐dienoyl‐CoA reductase, auxiliary enzyme of fatty acid beta‐oxidation; homodimeric enzyme required for growth and sporulation on petroselineate medium; expression induced during late sporulation and in the presence of oleate Putative protein of unknown function; sumoylated under stress conditions in a genome wide study; YDR336W is not an essential gene Rhodanese domain sulfur transferase, accepts persulfite from Nfs1p and transfers it to Uba4p in the pathway for 2‐thiolation of the wobble uridine base of tRNAs; also stimulates sulfur transfer by Nfs1p; may be mitochondrially localized Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm Putative protein of unknown function; overexpression causes a cell cycle delay or arrest; green fluorescent protein (GFP)‐fusion protein localizes to the vacuole; null mutant displays elevated frequency of mitochondrial genome loss ‐0.6267 0.0408 127 adj.p.val log2 (mtr10[P f/NP.f]/ WT[Pf/N P.f]) Symbols ORF Description SWD3 YBR175W SNX41 YDR425W STB2 YMR053C NA YMR315W DCG1 YIR030C ARG5,6 YER069W HAP3 YBL021C YCT1 YLL055W NA YLR352W NA YNL193W MET13 YGL125W ARG1 YOL058W TNA1 YGR260W Essential subunit of the COMPASS (Set1C) complex, which methylates histone H3 on lysine 4 and is required in transcriptional silencing near telomeres; WD40 beta propeller superfamily member and ortholog of mammalian WDR5 Sorting nexin, involved in the retrieval of late‐Golgi SNAREs from the post‐Golgi endosome to the trans‐Golgi network; interacts with Snx4p Protein that interacts with Sin3p in a two‐hybrid assay and is part of a large protein complex with Sin3p and Stb1p Protein with NADP(H) oxidoreductase activity; transcription is regulated by Stb5p in response to NADPH depletion induced by diamide; promoter contains a putative Stb5p binding site Protein of unknown function, expression is sensitive to nitrogen catabolite repression and regulated by Dal80p; contains transmembrane domain Protein that is processed in the mitochondrion to yield acetylglutamate kinase and N‐acetyl‐gamma‐glutamyl‐phosphate reductase, which catalyze the 2nd and 3rd steps in arginine biosynthesis; enzymes form a complex with Arg2p Subunit of the heme‐activated, glucose‐repressed Hap2p/3p/4p/5p CCAAT‐binding complex, a transcriptional activator and global regulator of respiratory gene expression; contains sequences contributing to both complex assembly and DNA binding High‐affinity cysteine‐specific transporter with similarity to the Dal5p family of transporters; green fluorescent protein (GFP)‐fusion protein localizes to the endoplasmic reticulum; YCT1 is not an essential gene Putative protein of unknown function with similarity to F‐box proteins; interacts with Skp1p and Cdc53p; YLR352W is not an essential gene Putative protein of unknown function; exhibits a two‐hybrid interaction with Yhr151cp in a large‐scale analysis Major isozyme of methylenetetrahydrofolate reductase, catalyzes the reduction of 5,10‐methylenetetrahydrofolate to 5‐ methyltetrahydrofolate in the methionine biosynthesis pathway Arginosuccinate synthetase, catalyzes the formation of L‐ argininosuccinate from citrulline and L‐aspartate in the arginine biosynthesis pathway; potential Cdc28p substrate High affinity nicotinic acid plasma membrane permease, responsible for uptake of low levels of nicotinic acid; expression of the gene increases in the absence of extracellular nicotinic acid or para‐ aminobenzoate (PABA) 128 adj.p.val ‐0.6238 0.0366 ‐0.6154 0.0393 ‐0.6085 0.0419 ‐0.6063 0.0424 ‐0.6057 0.0449 ‐0.5991 0.0424 ‐0.5909 0.0459 ‐0.5853 0.0400 ‐0.5850 0.0493 ‐0.5735 0.0493 ‐0.5689 0.0459 ‐0.5550 0.0493 ‐0.5346 0.0460 APPENDIX C: Transcripts with increased translation activity index in fed mtr10Δ cells The transcripts with decreased P/NP ratio (cutoff condition: log2 FC 0.6 and adjusted p-value 0.05) in mtr10Δ cells while compared to wild-type cells in fed condition are listed below and ordered according to log2 (fold-change). Transcripts with increased P/NP ratio in fed mtr10Δ cells. Symbols ORF Description log2 (mtr10[Pf/N P.f]/WT[Pf/ NP.f]) AFFX‐ 18srRnac_a t NA NA 2.9412 0.0267 AFFX‐r2‐Sc‐ 18SrRNA‐ 3_at NA NA 2.9310 0.0454 NA YHL012W Putative protein of unknown function, has some homology to Ugp1p, which encodes UDP‐glucose pyrophosphorylase 1.6485 0.0171 NA YBR238C Mitochondrial membrane protein with similarity to Rmd9p; not required for respiratory growth but causes a synthetic respiratory defect in combination with rmd9 mutations; transcriptionally up‐ regulated by TOR; deletion increases life span 1.3591 0.0403 YAP7 YOL028C Putative basic leucine zipper (bZIP) transcription factor 1.2656 0.0276 KAP123 YER110C Karyopherin beta, mediates nuclear import of ribosomal proteins prior to assembly into ribosomes and import of histones H3 and H4; localizes to the nuclear pore, nucleus, and cytoplasm; exhibits genetic interactions with RAI1 1.2417 0.0293 KAP95 YLR347C Karyopherin beta, forms a complex with Srp1p/Kap60p; interacts with nucleoporins to mediate nuclear import of NLS‐containing cargo proteins via the nuclear pore complex; regulates PC biosynthesis; GDP‐to‐GTP exchange factor for Gsp1p 1.2363 0.0346 ACC1 YNR016C Acetyl‐CoA carboxylase, biotin containing enzyme that catalyzes the carboxylation of acetyl‐CoA to form malonyl‐CoA; required for de novo biosynthesis of long‐chain fatty acids 1.2323 0.0171 129 adj.p.v al log2 (mtr10[Pf/N P.f]/WT[Pf/ NP.f]) adj.p.v al Symbols ORF Description SSA3 YBL075C ATPase involved in protein folding and the response to stress; plays a role in SRP‐dependent cotranslational protein‐membrane targeting and translocation; member of the heat shock protein 70 (HSP70) family; localized to the cytoplasm 1.1830 0.0431 CDC34 YDR054C Ubiquitin‐conjugating enzyme (E2) and catalytic subunit of SCF ubiquitin‐protein ligase complex (together with Skp1p, Rbx1p, Cdc53p, and an F‐box protein) that regulates cell cycle progression by targeting key substrates for degradation 1.1211 0.0293 RNR1 YER070W Major isoform of the large subunit of ribonucleotide‐diphosphate reductase; the RNR complex catalyzes rate‐limiting step in dNTP synthesis, regulated by DNA replication and DNA damage checkpoint pathways via localization of small subunits 1.1207 0.0205 AFFX‐r2‐Sc‐ SRB4‐5_at NA NA 1.1078 0.0302 NA Putative protein of unknown function 1.0989 0.0171 NUP192 YMR230W‐ A YJL039C Essential structural subunit of the nuclear pore complex (NPC), localizes to the nuclear periphery of nuclear pores, homologous to human p205 1.0915 0.0319 DRS2 YAL026C Aminophospholipid translocase (flippase) that maintains membrane lipid asymmetry in post‐Golgi secretory vesicles; contributes to clathrin‐coated vesicle formation and endocytosis; mutations in human homolog ATP8B1 result in liver disease 1.0772 0.0416 TIF4632 YGL049C Translation initiation factor eIF4G, subunit of the mRNA cap‐ binding protein complex (eIF4F) that also contains eIF4E (Cdc33p); associates with the poly(A)‐binding protein Pab1p, also interacts with eIF4A (Tif1p); homologous to Tif4631p 1.0711 0.0293 GSC2 YGR032W Catalytic subunit of 1,3‐beta‐glucan synthase, involved in formation of the inner layer of the spore wall; activity positively regulated by Rho1p and negatively by Smk1p; has similarity to an alternate catalytic subunit, Fks1p (Gsc1p) 1.0557 0.0496 MPT5 YGL178W Member of the Puf family of RNA‐binding proteins; binds to mRNAs encoding chromatin modifiers and spindle pole body components; involved in longevity, maintenance of cell wall integrity, and sensitivity to and recovery from pheromone arrest 1.0514 0.0301 NA YNL247W Cysteinyl‐tRNA synthetase; may interact with ribosomes, based on co‐purification experiments 1.0468 0.0293 SRP40 YKR092C Nucleolar, serine‐rich protein with a role in preribosome assembly or transport; may function as a chaperone of small nucleolar ribonucleoprotein particles (snoRNPs); immunologically and structurally to rat Nopp140 1.0348 0.0288 CDC60 YPL160W Cytosolic leucyl tRNA synthetase, ligates leucine to the appropriate tRNA 1.0205 0.0301 CHC1 YGL206C Clathrin heavy chain, subunit of the major coat protein involved in intracellular protein transport and endocytosis; two heavy chains form the clathrin triskelion structural component; the light chain (CLC1) is thought to regulate function 1.0150 0.0403 RPB2 YOR151C RNA polymerase II second largest subunit B150, part of central core; similar to bacterial beta subunit 1.0088 0.0301 130 log2 (mtr10[Pf/N P.f]/WT[Pf/ NP.f]) adj.p.v al Symbols ORF Description TCB3 YML072C Lipid‐binding protein, localized to the bud via specific mRNA transport; non‐tagged protein detected in a phosphorylated state in mitochondria; GFP‐fusion protein localizes to the cell periphery; C‐termini of Tcb1p, Tcb2p and Tcb3p interact 1.0075 0.0416 NA YGR054W 0.9970 0.0326 PMA1 YGL008C Eukaryotic initiation factor (eIF) 2A; associates specifically with both 40S subunits and 80 S ribosomes, and interacts genetically with both eIF5b and eIF4E; homologous to mammalian eIF2A Plasma membrane H+‐ATPase, pumps protons out of the cell; major regulator of cytoplasmic pH and plasma membrane potential; part of the P2 subgroup of cation‐transporting ATPases; Hsp30p plays a role in Pma1p regulation 0.9908 0.0377 SFC1 YJR095W Mitochondrial succinate‐fumarate transporter, transports succinate into and fumarate out of the mitochondrion; required for ethanol and acetate utilization 0.9668 0.0293 DED1 YOR204W 0.9665 0.0410 NA YBR056W‐A ATP‐dependent DEAD (Asp‐Glu‐Ala‐Asp)‐box RNA helicase, required for translation initiation of all yeast mRNAs; mutations in human DEAD‐box DBY are a frequent cause of male infertility Dubious open reading frame unlikely to encode a protein, based on available experimental and comparative sequence data; partially overlaps the dubious ORF YBR056C‐B 0.9631 0.0330 NA YHR210C Putative protein of unknown function; non‐essential gene; highly expressed under anaeorbic conditions; sequence similarity to aldose 1‐epimerases such as GAL10 0.9419 0.0171 COS7 YDL248W Protein of unknown function, member of the DUP380 subfamily of conserved, often subtelomerically‐encoded proteins; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐throughput studies 0.9406 0.0459 PIR3 YKL163W 0.9395 0.0248 JLP1 YLL057C 0.9375 0.0205 FAA4 YMR246W 0.9120 0.0459 NOT3 YIL038C O‐glycosylated covalently‐bound cell wall protein required for cell wall stability; expression is cell cycle regulated, peaking in M/G1 and also subject to regulation by the cell integrity pathway Fe(II)‐dependent sulfonate/alpha‐ketoglutarate dioxygenase, involved in sulfonate catabolism for use as a sulfur source; contains sequence that resembles a J domain (typified by the E. coli DnaJ protein); induced by sulphur starvation Long chain fatty acyl‐CoA synthetase, activates imported fatty acids with a preference for C12:0‐C16:0 chain lengths; functions in long chain fatty acid import; important for survival during stationary phase; localized to lipid particles Subunit of the CCR4‐NOT complex, which is a global transcriptional regulator with roles in transcription initiation and elongation and in mRNA degradation 0.9069 0.0366 CRM1 YGR218W Major karyopherin, involved in export of proteins, RNAs, and ribosomal subunits from the nucleus; exportin 0.8817 0.0464 NA YBL095W Putative protein of unknown function; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐ throughput studies 0.8798 0.0295 FLC3 YGL139W Putative FAD transporter, similar to Flc1p and Flc2p; localized to the ER 0.8766 0.0215 TRA1 YHR099W Subunit of SAGA and NuA4 histone acetyltransferase complexes; interacts with acidic activators (e.g., Gal4p) which leads to transcription activation; similar to human TRRAP, which is a cofactor for c‐Myc mediated oncogenic transformation 0.8766 0.0488 131 log2 (mtr10[Pf/N P.f]/WT[Pf/ NP.f]) adj.p.v al Symbols ORF Description NUM1 YDR150W Protein required for nuclear migration, localizes to the mother cell cortex and the bud tip; may mediate interactions of dynein and cytoplasmic microtubules with the cell cortex 0.8647 0.0454 NA YBR196C‐B Putative protein of unknown function; identified by expression profiling and mass spectrometry 0.8608 0.0293 AAD6 YFL056C Putative aryl‐alcohol dehydrogenase with similarity to P. chrysosporium aryl‐alcohol dehydrogenase, involved in the oxidative stress response; expression induced in cells treated with the mycotoxin patulin 0.8545 0.0312 DCP2 YNL118C 0.8334 0.0459 PRP16 YKR086W 0.8202 0.0308 NA YMR102C Catalytic subunit of the Dcp1p‐Dcp2p decapping enzyme complex, which removes the 5' cap structure from mRNAs prior to their degradation; member of the Nudix hydrolase family RNA helicase in the DEAH‐box family involved in the second catalytic step of splicing, exhibits ATP‐dependent RNA unwinding activity Protein of unknown function; transcription is activated by paralogous transcription factors Yrm1p and Yrr1p along with genes involved in multidrug resistance; mutant shows increased resistance to azoles; YMR102C is not an essential gene 0.8193 0.0408 PDH1 YPR002W 0.8185 0.0426 AFFX‐ YER022w5_ at NA Mitochondrial protein that participates in respiration, induced by diauxic shift; homologous to E. coli PrpD, may take part in the conversion of 2‐methylcitrate to 2‐methylisocitrate NA 0.8087 0.0288 MCM7 YBR202W Component of the hexameric MCM complex, which is important for priming origins of DNA replication in G1 and becomes an active ATP‐dependent helicase that promotes DNA melting and elongation when activated by Cdc7p‐Dbf4p in S‐phase 0.8077 0.0366 SAC1 YKL212W Phosphatidylinositol phosphate (PtdInsP) phosphatase involved in hydrolysis of PtdIns[4]P; transmembrane protein localizes to ER and Golgi; involved in protein trafficking and processing, secretion, and cell wall maintenance 0.8050 0.0291 VIP1 YLR410W Inositol hexakisphosphate (IP6) and inositol heptakisphosphate (IP7) kinase; IP7 production is important for phosphate signaling; involved in cortical actin cytoskeleton function, and invasive pseudohyphal growth analogous to S. pombe asp1 0.7931 0.0366 IRC7 YFR055W Putative cystathionine beta‐lyase; involved in copper ion homeostasis and sulfur metabolism; null mutant displays increased levels of spontaneous Rad52p foci; expression induced by nitrogen limitation in a GLN3, GAT1‐dependent manner 0.7880 0.0437 IRR1 YIL026C Subunit of the cohesin complex, which is required for sister chromatid cohesion during mitosis and meiosis and interacts with centromeres and chromosome arms, essential for viability 0.7801 0.0288 RTS1 YOR014W B‐type regulatory subunit of protein phosphatase 2A (PP2A); homolog of the mammalian B' subunit of PP2A 0.7654 0.0301 MCM1 YMR043W Transcription factor involved in cell‐type‐specific transcription and pheromone response; plays a central role in the formation of both repressor and activator complexes 0.7526 0.0488 RPN2 YIL075C Subunit of the 26S proteasome, substrate of the N‐ acetyltransferase Nat1p 0.7460 0.0349 132 log2 (mtr10[Pf/N P.f]/WT[Pf/ NP.f]) adj.p.v al Symbols ORF Description SMI1 YGR229C Protein involved in the regulation of cell wall synthesis; proposed to be involved in coordinating cell cycle progression with cell wall integrity 0.7433 0.0301 DIA1 YMR316W Protein of unknown function, involved in invasive and pseudohyphal growth; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm in a punctate pattern 0.7421 0.0454 GFA1 YKL104C Glutamine‐fructose‐6‐phosphate amidotransferase, catalyzes the formation of glucosamine‐6‐P and glutamate from fructose‐6‐P and glutamine in the first step of chitin biosynthesis 0.7396 0.0254 TCB1 YOR086C Lipid‐binding protein containing three calcium and lipid binding domains; non‐tagged protein localizes to mitochondria and GFP‐ fusion protein localizes to the cell periphery; C‐termini of Tcb1p, Tcb2p and Tcb3p interact 0.7338 0.0293 UFD2 YDL190C Ubiquitin chain assembly factor (E4) that cooperates with a ubiquitin‐activating enzyme (E1), a ubiquitin‐conjugating enzyme (E2), and a ubiquitin protein ligase (E3) to conjugate ubiquitin to substrates; also functions as an E3 0.7338 0.0293 NDD1 YOR372C Transcriptional activator essential for nuclear division; localized to the nucleus; essential component of the mechanism that activates the expression of a set of late‐S‐phase‐specific genes 0.7332 0.0252 SIC1 YLR079W Inhibitor of Cdc28‐Clb kinase complexes that controls G1/S phase transition, preventing premature S phase and ensuring genomic integrity; phosphorylation targets Sic1p for SCF(CDC4)‐dependent turnover; functional homolog of mammalian Kip1 0.7241 0.0459 SEC27 YGL137W Essential beta'‐coat protein of the COPI coatomer, involved in ER‐ to‐Golgi and Golgi‐to‐ER transport; contains WD40 domains that mediate cargo selective interactions; 45% sequence identity to mammalian beta'‐COP 0.7130 0.0437 SIS2 YKR072C Negative regulatory subunit of protein phosphatase 1 Ppz1p and also a subunit of the phosphopantothenoylcysteine decarboxylase (PPCDC; Cab3p, Sis2p, Vhs3p) complex, which catalyzes the third step of coenzyme A biosynthesis 0.7129 0.0326 RAD51 YER095W Strand exchange protein, forms a helical filament with DNA that searches for homology; involved in the recombinational repair of double‐strand breaks in DNA during vegetative growth and meiosis; homolog of Dmc1p and bacterial RecA protein 0.7101 0.0301 LHS1 YKL073W Molecular chaperone of the endoplasmic reticulum lumen, involved in polypeptide translocation and folding; nucleotide exchange factor for the ER lumenal Hsp70 chaperone Kar2p; regulated by the unfolded protein response pathway 0.7092 0.0288 MSN4 YKL062W Transcriptional activator related to Msn2p; activated in stress conditions, which results in translocation from the cytoplasm to the nucleus; binds DNA at stress response elements of responsive genes, inducing gene expression 0.7027 0.0326 ADE2 YOR128C Phosphoribosylaminoimidazole carboxylase, catalyzes a step in the 'de novo' purine nucleotide biosynthetic pathway; red pigment accumulates in mutant cells deprived of adenine 0.7018 0.0454 133 log2 (mtr10[Pf/N P.f]/WT[Pf/ NP.f]) adj.p.v al Symbols ORF Description STP1 YDR463W Transcription factor, undergoes proteolytic processing by SPS (Ssy1p‐Ptr3p‐Ssy5p)‐sensor component Ssy5p in response to extracellular amino acids; activates transcription of amino acid permease genes and may have a role in tRNA processing 0.7012 0.0454 ACS2 YLR153C Acetyl‐coA synthetase isoform which, along with Acs1p, is the nuclear source of acetyl‐coA for histone acetylation; mutants affect global transcription; required for growth on glucose; expressed under anaerobic conditions 0.6811 0.0368 NUP133 YKR082W 0.6787 0.0464 MNN5 YJL186W 0.6765 0.0366 PSD2 YGR170W Subunit of the Nup84p subcomplex of the nuclear pore complex (NPC), localizes to both sides of the NPC, required to establish a normal nucleocytoplasmic concentration gradient of the GTPase Gsp1p Alpha‐1,2‐mannosyltransferase, responsible for addition of the second alpha‐1,2‐linked mannose of the branches on the mannan backbone of oligosaccharides, localizes to an early Golgi compartment Phosphatidylserine decarboxylase of the Golgi and vacuolar membranes, converts phosphatidylserine to phosphatidylethanolamine 0.6765 0.0384 JJJ2 YJL162C Protein of unknown function, contains a J‐domain, which is a region with homology to the E. coli DnaJ protein 0.6749 0.0431 RIO2 YNL207W Essential serine kinase involved in the processing of the 20S pre‐ rRNA into mature 18S rRNA; has similarity to Rio1p 0.6679 0.0408 MUD1 YBR119W U1 snRNP A protein, homolog of human U1‐A; involved in nuclear mRNA splicing 0.6666 0.0301 CDC20 YGL116W 0.6666 0.0301 RFC4 YOL094C Cell‐cycle regulated activator of anaphase‐promoting complex/cyclosome (APC/C), which is required for metaphase/anaphase transition; directs ubiquitination of mitotic cyclins, Pds1p, and other anaphase inhibitors; potential Cdc28p substrate Subunit of heteropentameric Replication factor C (RF‐C), which is a DNA binding protein and ATPase that acts as a clamp loader of the proliferating cell nuclear antigen (PCNA) processivity factor for DNA polymerases delta and epsilon 0.6648 0.0301 TFA1 YKL028W TFIIE large subunit, involved in recruitment of RNA polymerase II to the promoter, activation of TFIIH, and promoter opening 0.6596 0.0459 YPT32 YGL210W Rab family GTPase, very similar to Ypt31p; involved in the exocytic pathway; mediates intra‐Golgi traffic or the budding of post‐Golgi vesicles from the trans‐Golgi 0.6574 0.0301 NA YGL140C Putative protein of unknown function; non‐essential gene; contains multiple predicted transmembrane domains 0.6567 0.0454 ARO9 YHR137W Aromatic aminotransferase II, catalyzes the first step of tryptophan, phenylalanine, and tyrosine catabolism 0.6498 0.0431 PFK27 YOL136C 6‐phosphofructo‐2‐kinase, catalyzes synthesis of fructose‐2,6‐ bisphosphate; inhibited by phosphoenolpyruvate and sn‐glycerol 3‐ phosphate, expression induced by glucose and sucrose, transcriptional regulation involves protein kinase A 0.6479 0.0330 TPD3 YAL016W Regulatory subunit A of the heterotrimeric protein phosphatase 2A (PP2A), which also contains regulatory subunit Cdc55p and either catalytic subunit Pph21p or Pph22p; required for cell morphogenesis and transcription by RNA polymerase III 0.6442 0.0400 134 log2 (mtr10[Pf/N P.f]/WT[Pf/ NP.f]) adj.p.v al Symbols ORF Description REB1 YBR049C RNA polymerase I enhancer binding protein; DNA binding protein which binds to genes transcribed by both RNA polymerase I and RNA polymerase II; required for termination of RNA polymerase I transcription 0.6403 0.0431 SMP1 YBR182C Putative transcription factor involved in regulating the response to osmotic stress; member of the MADS‐box family of transcription factors 0.6394 0.0485 NUP145 YGL092W Essential nucleoporin, catalyzes its own cleavage in vivo to generate a C‐terminal fragment that assembles into the Nup84p subcomplex of the nuclear pore complex, and an N‐terminal fragment of unknown function that is homologous to Nup100p 0.6392 0.0431 KES1 YPL145C Member of the oxysterol binding protein family, which includes seven yeast homologs; involved in negative regulation of Sec14p‐ dependent Golgi complex secretory functions, peripheral membrane protein that localizes to the Golgi complex 0.6191 0.0478 MNN2 YBR015C Alpha‐1,2‐mannosyltransferase, responsible for addition of the first alpha‐1,2‐linked mannose to form the branches on the mannan backbone of oligosaccharides, localizes to an early Golgi compartment 0.5941 0.0459 NA YOL098C Putative metalloprotease 0.5910 0.0459 CDC55 YGL190C Non‐essential regulatory subunit B of protein phosphatase 2A (PP2A), which has multiple roles in mitosis and protein biosynthesis; involved in regulation of mitotic exit; found in the nucleus of most cells, also at bud neck and bud tip 0.5698 0.0459 CDC3 YLR314C Component of the septin ring of the mother‐bud neck that is required for cytokinesis; septins recruit proteins to the neck and can act as a barrier to diffusion at the membrane, and they comprise the 10nm filaments seen with EM 0.5571 0.0458 ARO7 YPR060C Chorismate mutase, catalyzes the conversion of chorismate to prephenate to initiate the tyrosine/phenylalanine‐specific branch of aromatic amino acid biosynthesis 0.5515 0.0454 135 APPENDIX D: Transcripts with changed translation activity index in mtr10Δ cells in amino acid starvation condition This section includes the transcripts with decreased P/NP ratio (cutoff condition: log2 FC 0.6 and adjusted p-value 0.05) and the transcripts with increase P/NP ratio (log2 FC 0.6 and adjusted p-value 0.05) in mtr10Δ cells while compared to wild-type cells in amino acid starvation condition. The affected transcripts are ordered according to log2 (fold-change). Transcripts with decreased P/NP ratio in amino acid starved mtr10Δ cells. log2 (mtr10[Ps/ NP.s]/WT[ Ps/NP.s]) adj.p. val Symbols ORF Description AAD14 YNL331C Putative aryl‐alcohol dehydrogenase with similarity to P. chrysosporium aryl‐alcohol dehydrogenase; mutational analysis has not yet revealed a physiological role ‐1.9155 0.0302 AMS1 YGL156W Vacuolar alpha mannosidase, involved in free oligosaccharide (fOS) degradation; delivered to the vacuole in a novel pathway separate from the secretory pathway ‐1.1781 0.0479 CIR2 YOR356W Putative ortholog of human electron transfer flavoprotein dehydrogenase (ETF‐dH); found in a large supramolecular complex with other mitochondrial dehydrogenases; may have a role in oxidative stress response ‐1.0468 0.0479 NSE4 YDL105W Nuclear protein that plays a role in the function of the Smc5p‐Rhc18p complex ‐1.0143 0.0479 FLO9 YAL063C Lectin‐like protein with similarity to Flo1p, thought to be expressed and involved in flocculation ‐0.9790 0.0479 136 Transcripts with increased P/NP ratio in amino acid starved mtr10Δ cells. log2 (mtr10[Ps/ NP.s]/WT[ Ps/NP.s]) adj.P.V al symbols ORF Description NA YLR154W‐E Dubious open reading frame unlikely to encode a protein; encoded within the the 35S rRNA gene on the opposite strand 2.6224 0.0302 NA YHL012W Putative protein of unknown function, has some homology to Ugp1p, which encodes UDP‐glucose pyrophosphorylase 1.8017 0.0479 137 APPENDIX E: Transcripts with decreased translation activity index in fed msn5Δ cells The transcripts with decreased P/NP ratio (cutoff condition: log2 FC 0.6 and adjusted p-value 0.05) in msn5Δ cells while compared to wild-type cells in fed condition are listed below and ordered according to log2 (fold-change). Transcripts with decreased P/NP ratio in fed msn5Δ cells. log2 (msn5[Pf/ NP.f]/WT[ Pf/NP.f]) adj.p. val Symbols ORF Description STR3 YGL184C ‐2.6447 0.0099 MSN5 YDR335W ‐2.5962 0.0205 SUL1 YBR294W ‐2.4354 0.0239 SEO1 YAL067C Peroxisomal cystathionine beta‐lyase, converts cystathionine into homocysteine; may be redox regulated by Gto1p Karyopherin involved in nuclear import and export of proteins, including import of replication protein A and export of Swi6p, Far1p, and Pho4p; required for re‐export of mature tRNAs after their retrograde import from the cytoplasm High affinity sulfate permease; sulfate uptake is mediated by specific sulfate transporters Sul1p and Sul2p, which control the concentration of endogenous activated sulfate intermediates Putative permease, member of the allantoate transporter subfamily of the major facilitator superfamily; mutation confers resistance to ethionine sulfoxide ‐1.8658 0.0099 ARG3 YJL088W Ornithine carbamoyltransferase (carbamoylphosphate:L‐ornithine carbamoyltransferase), catalyzes the sixth step in the biosynthesis of the arginine precursor ornithine ‐1.6423 0.0246 MMP1 YLL061W High‐affinity S‐methylmethionine permease, required for utilization of S‐methylmethionine as a sulfur source; has similarity to S‐ adenosylmethionine permease Sam3p ‐1.5790 0.0099 MET3 YJR010W ATP sulfurylase, catalyzes the primary step of intracellular sulfate activation, essential for assimilatory reduction of sulfate to sulfide, involved in methionine metabolism ‐1.4117 0.0246 MHT1 YLL062C S‐methylmethionine‐homocysteine methyltransferase, functions along with Sam4p in the conversion of S‐adenosylmethionine (AdoMet) to methionine to control the methionine/AdoMet ratio ‐1.3841 0.0239 138 PRM1 YNL279W Pheromone‐regulated multispanning membrane protein involved in membrane fusion during mating; predicted to have 5 transmembrane segments and a coiled coil domain; localizes to the shmoo tip; regulated by Ste12p 139 ‐1.0819 0.0291 APPENDIX F: Summary tables of functional catalogue analysis for affected transcripts in mtr10Δ cells in fed or amino acid starvation conditions This section includes four summary results of functional catalogue (FunCat) analysis of transcriptionally differentially expressed genes (cutoff conditions: log2 FC FC 0.6 or log2 0.6 and adjusted p-value < 0.001) in mtr10Δ cells in fed or amino acid starvation conditions. The FunCat analysis was executed in the website: http://mips.helmholtzmuenchen.de/proj/funcatDB/ (Ruepp et al., 2004) Table F. 1. FunCat analysis of down-regulated transcripts in mtr10Δ cells in fed condition. FUNCTIONAL CATALOGUE METABOLISM lipid, fatty acid and isoprenoid metabolism membrane lipid metabolism phospholipid metabolism modification with fatty acids (e.g. myristylation, palmitylation, farnesylation) thiamine pyrophosphate binding transported compounds (substrates) ion transport cation transport (H+, Na+, K+, Ca2+ , NH4+, etc.) heavy metal ion transport (Cu+, Fe3+, etc.) siderophore‐iron transport lipid/fatty acid transport nucleotide/nucleoside/nucleobase transport 140 numbers of gene matches 65 22 8 7 4 1 32 11 8 6 3 6 4 p-value 0.003177 8.86E‐05 0.004182 0.005404 0.019417 0.031648 0.001356 0.007357 0.038983 0.006192 0.005559 0.002408 0.003164 continued. Table F. 1. continued. numbers of gene matches 3 8 13 5 7 FUNCTIONAL CATALOGUE vitamine/cofactor transport drug/toxin transport transport facilities transport ATPases ABC transporters p-value 0.008779 2.31E‐05 0.008971 0.027214 1.92E‐05 Table F. 2. FunCat analysis of up-regulated transcripts in mtr10Δ cells in fed condition. FUNCTIONAL CATALOGUE metabolism of lysine biosynthesis of lysine aminoadipic acid pathway degradation of lysine metabolism of phenylalanine nitrogen, sulfur and selenium metabolism nitrogen metabolism catabolism of nitrogenous compounds purin nucleotide/nucleoside/nucleobase metabolism purine nucleotide /nucleoside/nucleobase catabolism purine nucleotide/nucleoside/nucleobase anabolism electron transport and membrane‐associated energy conservation aerobic respiration fermentation lactate fermentation DNA repair mRNA processing (splicing, 5'‐, 3'‐end processing) modification by ubiquitin‐related proteins transmembrane signal transduction detoxification involving cytochrome P450 chemoperception and response pheromone response, mating‐type determination, sex‐specific proteins CELL TYPE DIFFERENTIATION fungal/microorganismic cell type differentiation fungal and other eukaryotic cell type differentiation development of asco‐ basidio‐ or zygospore UNCLASSIFIED PROTEINS 141 numbers of gene matches 4 4 4 2 4 11 3 3 10 2 5 7 10 6 2 17 16 4 7 2 20 17 35 35 35 21 81 p-value 0.009055 0.001352 0.000847 0.026075 0.011347 0.014891 0.00464 0.00464 0.00141 0.00841 0.018126 0.044798 0.00799 0.039273 0.01622 0.005148 0.0429 0.024079 0.007529 0.01622 0.034217 0.025928 0.018017 0.018017 0.018017 0.000228 0.236462 Table F. 3. FunCat analysis of down-regulated transcripts in mtr10Δ cells in amino acid starved condition. FUNCTIONAL CATALOGUE METABOLISM phosphate metabolism C‐compound and carbohydrate metabolism sugar, glucoside, polyol and carboxylate anabolism polysaccharide metabolism glycogen metabolism glycogen anabolism lipid, fatty acid and isoprenoid metabolism catabolism of vitamins, cofactors, and prosthetic groups metabolism of secondary products derived from primary amino acids metabolism of nonprotein amino acids metabolism of energy reserves (e.g. glycogen, trehalose) oxidation of fatty acids transcription activation modification with sugar residues (e.g. glycosylation, deglycosylation) CELLULAR TRANSPORT, TRANSPORT FACILITIES AND TRANSPORT ROUTES transported compounds (substrates) ion transport cation transport (H+, Na+, K+, Ca2+ , NH4+, etc.) heavy metal ion transport (Cu+, Fe3+, etc.) siderophore‐iron transport C‐compound and carbohydrate transport amino acid/amino acid derivatives transport amine / polyamine transport lipid/fatty acid transport nucleotide/nucleoside/nucleobase transport allantoin and allantoate transport vitamine/cofactor transport drug/toxin transport transport facilities channel / pore class transport carrier (electrochemical potential‐driven transport) symporter proton driven symporter ABC transporters peroxisomal transport MAPKKK cascade osmotic and salt stress response disease, virulence and defense resistance proteins chemical agent resistance detoxification INTERACTION WITH THE ENVIRONMENT 142 numbers of gene matches 167 49 58 7 12 4 4 47 2 5 4 10 3 8 65 11 121 88 29 22 15 6 12 14 4 11 8 3 4 15 43 4 5 2 2 15 5 6 10 7 7 7 21 p-value 0.0013 0.032306 0.030207 0.034247 0.0124 0.000307 0.000307 3.05E‐05 0.022994 0.02366 0.008902 0.026663 0.040793 0.031922 0.043824 0.001018 4.71E‐07 5.08E‐05 0.001191 4.41E‐05 0.000305 0.009692 2.45E‐05 0.024365 0.001436 0.0002 0.029111 0.031735 5.87E‐07 2.17E‐08 0.04027 0.018793 0.043248 0.022994 2.25E‐09 0.02366 0.030374 0.036928 0.034247 0.025432 0.012719 0.001624 0.000276 continued Table. F.3. continued. FUNCTIONAL CATALOGUE homeostasis homeostasis of cations homeostasis of metal ions (Na, K, Ca etc.) UNCLASSIFIED PROTEINS numbers of gene matches 64 29 26 20 p-value 0.001681 0.002586 0.000386 0.208561 Table F. 4. FunCat analysis of up-regulated transcripts in mtr10Δ cells in amino acid starved condition. numbers of gene matches 2 140 73 20 36 29 28 26 91 60 25 10 25 24 6 13 4 12 9 3 30 25 145 27 6 29 23 FUNCTIONAL CATALOGUE degradation of arginine CELL CYCLE AND DNA PROCESSING DNA processing DNA synthesis and replication DNA recombination and DNA repair DNA repair DNA restriction or modification DNA conformation modification (e.g. chromatin) cell cycle mitotic cell cycle and cell cycle control mitotic cell cycle M phase meiosis nuclear and chromosomal cycle chromosome condensation chromosome segregation/division nuclear division cell cycle dependent cytoskeleton reorganization spindle pole body/centrosome and microtubule cycle cell cycle dependent actin filament reorganization mRNA processing (splicing, 5'‐, 3'‐end processing) splicing PROTEIN FATE (folding, modification, destination) protein folding and stabilization modification by ubiquitin‐related proteins cytoplasmic and nuclear protein degradation proteasomal degradation (ubiquitin/proteasomal pathway) PROTEIN WITH BINDING FUNCTION OR COFACTOR REQUIREMENT (structural or catalytic) 137 protein binding 67 2 amino acid/amino acid derivatives binding 143 p-value 0.026315 1.88E‐06 0.000536 0.042889 0.010629 0.000556 0.027059 0.036794 0.000135 0.004847 0.01546 0.022457 0.011516 0.000119 0.01559 0.003567 0.016547 0.001971 0.017142 0.023532 0.003396 0.001982 0.000223 9.87E‐08 0.012322 0.007476 0.002473 5.55E‐05 1.66E‐06 0.026315 continued. Table. F. 4. continued. FUNCTIONAL CATALOGUE magnesium binding protease activator nuclear transport unfolded protein response (e.g. ER quality control) cellular sensing and response to external stimulus chemoperception and response pheromone response, mating‐type determination, sex‐specific proteins BIOGENESIS OF CELLULAR COMPONENTS cytoskeleton/structural proteins microtubule cytoskeleton UNCLASSIFIED PROTEINS 144 numbers of gene matches 2 2 14 15 37 34 0.049269 0.009375 0.032722 0.002077 0.03647 0.014459 26 102 37 11 139 0.0412 0.014298 0.006243 0.004383 0.356087 p-value APPENDIX G: Down-regulated genes in mtr10Δ cells in fed condition This section includes the list of transcriptionally differentially expressed genes (cutoff conditions: log2 FC 0.6 and adjusted p-value < 0.001) in mtr10Δ cells in fed condition. Down-regulated genes in mtr10Δ cells in fed condition log2(mtr10 .Tf/WT.Tf) Symbols ORF Description MTR10 YOR160W Nuclear import receptor, mediates the nuclear localization of proteins involved in mRNA‐nucleus export; promotes dissociation of mRNAs from the nucleus‐cytoplasm mRNA shuttling protein Npl3p; required for retrograde import of mature tRNAs ‐7.0599 0.0000 QDR2 YIL121W Multidrug transporter of the major facilitator superfamily, required for resistance to quinidine, barban, cisplatin, and bleomycin; may have a role in potassium uptake ‐4.0758 0.0000 FCY21 YER060W ‐3.1458 0.0000 BDH2 YAL061W Putative purine‐cytosine permease, very similar to Fcy2p but cannot substitute for its function Putative medium‐chain alcohol dehydrogenase with similarity to BDH1; transcription induced by constitutively active PDR1 and PDR3 ‐2.7008 0.0001 PNS1 YOR161C ‐2.4530 0.0000 NA NA FIT3 NA NA YOR383C Protein of unknown function; has similarity to Torpedo californica tCTL1p, which is postulated to be a choline transporter, neither null mutation nor overexpression affects choline transport NA NA Mannoprotein that is incorporated into the cell wall via a glycosylphosphatidylinositol (GPI) anchor, involved in the retention of siderophore‐iron in the cell wall ‐2.1966 ‐1.9974 ‐1.9076 0.0002 0.0001 0.0003 RGS2 YOR107W Negative regulator of glucose‐induced cAMP signaling; directly activates the GTPase activity of the heterotrimeric G protein alpha subunit Gpa2p ‐1.8591 0.0000 GTO3 YMR251W Omega class glutathione transferase; putative cytosolic localization ‐1.8261 0.0005 TPO4 YOR273C Polyamine transport protein, recognizes spermine, putrescine, and spermidine; localizes to the plasma membrane; member of the major facilitator superfamily ‐1.7975 0.0000 145 adj.p.val log2(mtr10 .Tf/WT.Tf) Symbols ORF Description HOR2 YER062C One of two redundant DL‐glycerol‐3‐phosphatases (RHR2/GPP1 encodes the other) involved in glycerol biosynthesis; induced in response to hyperosmotic stress and oxidative stress, and during the diauxic transition ‐1.6977 0.0002 HOP1 YIL072W Meiosis‐specific DNA binding protein that displays Red1p dependent localization to the unsynapsed axial‐lateral elements of the synaptonemal complex; required for homologous chromosome synapsis and chiasma formation ‐1.6539 0.0007 ICS2 YBR157C Protein of unknown function; null mutation does not confer any obvious defects in growth, spore germination, viability, or carbohydrate utilization ‐1.6524 0.0002 ARN2 YHL047C ‐1.6499 0.0006 FMP48 YGR052W Transporter, member of the ARN family of transporters that specifically recognize siderophore‐iron chelates; responsible for uptake of iron bound to the siderophore triacetylfusarinine C Putative protein of unknown function; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐ throughput studies; induced by treatment with 8‐methoxypsoralen and UVA irradiation ‐1.6387 0.0004 FIT2 YOR382W Mannoprotein that is incorporated into the cell wall via a glycosylphosphatidylinositol (GPI) anchor, involved in the retention of siderophore‐iron in the cell wall ‐1.6293 0.0009 RSB1 YOR049C Suppressor of sphingoid long chain base (LCB) sensitivity of an LCB‐ lyase mutation; putative integral membrane transporter or flippase that may transport LCBs from the cytoplasmic side toward the extracytoplasmic side of the membrane ‐1.5722 0.0003 ERC1 YHR032W Member of the multi‐drug and toxin extrusion (MATE) family of the multidrug/oligosaccharidyl‐lipid/polysaccharide (MOP) exporter superfamily; overproduction confers ethionine resistance and accumulation of S‐adenosylmethionine ‐1.5321 0.0002 NA YCR061W Protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm in a punctate pattern; induced by treatment with 8‐methoxypsoralen and UVA irradiation ‐1.5255 0.0003 SAP4 YGL229C ‐1.5182 0.0001 SFK1 YKL051W Protein required for function of the Sit4p protein phosphatase, member of a family of similar proteins that form complexes with Sit4p, including Sap155p, Sap185p, and Sap190p Plasma membrane protein that may act together with or upstream of Stt4p to generate normal levels of the essential phospholipid PI4P, at least partially mediates proper localization of Stt4p to the plasma membrane ‐1.5072 0.0000 NA YHR022C ‐1.4996 0.0001 THO1 YER063W Putative protein of unknown function; YHR022C is not an essential gene Conserved nuclear RNA‐binding protein; specifically binds to transcribed chromatin in a THO‐ and RNA‐dependent manner, genetically interacts with shuttling hnRNP NAB2; overproduction suppresses transcriptional defect caused by hpr1 mutation ‐1.4748 0.0001 ARN1 YHL040C Transporter, member of the ARN family of transporters that specifically recognize siderophore‐iron chelates; responsible for uptake of iron bound to ferrirubin, ferrirhodin, and related siderophores ‐1.4686 0.0002 NA TYE7 YHR140W YOR344C Putative integral membrane protein of unknown function Serine‐rich protein that contains a basic‐helix‐loop‐helix (bHLH) DNA binding motif; binds E‐boxes of glycolytic genes and contributes to their activation; may function as a transcriptional activator in Ty1‐ mediated gene expression ‐1.4314 ‐1.4233 0.0002 0.0003 146 adj.p.val log2(mtr10 .Tf/WT.Tf) Symbols ORF Description NA YOR152C TDH1 YJL052W Putative protein of unknown function; has no similarity to any known protein; YOR152C is not an essential gene Glyceraldehyde‐3‐phosphate dehydrogenase, isozyme 1, involved in glycolysis and gluconeogenesis; tetramer that catalyzes the reaction of glyceraldehyde‐3‐phosphate to 1,3 bis‐phosphoglycerate; detected in the cytoplasm and cell wall YRO2 YBR054W PRY1 PDR3 adj.p.val ‐1.4142 0.0001 ‐1.3912 0.0003 Putative protein of unknown function; the authentic, non‐tagged protein is detected in a phosphorylated state in highly purified mitochondria in high‐throughput studies; transcriptionally regulated by Haa1p ‐1.3897 0.0004 YJL079C YBL005W Protein of unknown function Transcriptional activator of the pleiotropic drug resistance network, regulates expression of ATP‐binding cassette (ABC) transporters through binding to cis‐acting sites known as PDREs (PDR responsive elements) ‐1.3703 ‐1.3401 0.0000 0.0002 NA RPS14B NA YJL191W NA Ribosomal protein 59 of the small subunit, required for ribosome assembly and 20S pre‐rRNA processing; mutations confer cryptopleurine resistance; nearly identical to Rps14Ap and similar to E. coli S11 and rat S14 ribosomal proteins ‐1.3381 ‐1.3157 0.0003 0.0002 TIR1 YER011W Cell wall mannoprotein of the Srp1p/Tip1p family of serine‐alanine‐ rich proteins; expression is downregulated at acidic pH and induced by cold shock and anaerobiosis; abundance is increased in cells cultured without shaking ‐1.3027 0.0001 PGM2 YMR105C ‐1.3001 0.0004 NA DSE1 YAR066W YER124C Phosphoglucomutase, catalyzes the conversion from glucose‐1‐ phosphate to glucose‐6‐phosphate, which is a key step in hexose metabolism; functions as the acceptor for a Glc‐phosphotransferase Putative GPI protein Daughter cell‐specific protein, may regulate cross‐talk between the mating and filamentation pathways; deletion affects cell separation after division and sensitivity to alpha‐factor and drugs affecting the cell wall ‐1.2845 ‐1.2744 0.0002 0.0006 AAR2 YBL074C ‐1.2717 0.0002 NA IRC15 NA YPL017C Component of the U5 snRNP, required for splicing of U3 precursors; originally described as a splicing factor specifically required for splicing pre‐mRNA of the MATa1 cistron NA Microtubule associated protein; regulates microtubule dynamics; required for accurate meiotic chromosome segregation; null mutant displays large budded cells due to delayed mitotic progression, increased levels of spontaneous Rad52 foci ‐1.2710 ‐1.2560 0.0003 0.0004 NA YLR126C Putative protein of unknown function with similarity to glutamine amidotransferase proteins; has Aft1p‐binding motif in the promoter; may be involved in copper and iron homeostasis; YLR126C is not an essential protein ‐1.2482 0.0004 ENO1 YGR254W Enolase I, a phosphopyruvate hydratase that catalyzes the conversion of 2‐phosphoglycerate to phosphoenolpyruvate during glycolysis and the reverse reaction during gluconeogenesis; expression is repressed in response to glucose ‐1.2413 0.0002 ENB1 YOL158C Endosomal ferric enterobactin transporter, expressed under conditions of iron deprivation; member of the major facilitator superfamily; expression is regulated by Rcs1p and affected by chloroquine treatment ‐1.2397 0.0004 FCY22 YER060W‐ A Putative purine‐cytosine permease, very similar to Fcy2p but cannot substitute for its function ‐1.2287 0.0006 147 log2(mtr10 .Tf/WT.Tf) Symbols ORF Description SPS19 YNL202W Peroxisomal 2,4‐dienoyl‐CoA reductase, auxiliary enzyme of fatty acid beta‐oxidation; homodimeric enzyme required for growth and sporulation on petroselineate medium; expression induced during late sporulation and in the presence of oleate ‐1.2265 0.0004 SHE4 YOR035C Protein containing a UCS (UNC‐45/CRO1/SHE4) domain, binds to myosin motor domains to regulate myosin function; involved in endocytosis, polarization of the actin cytoskeleton, and asymmetric mRNA localization ‐1.2201 0.0001 VMR1 YHL035C Vacuolar membrane protein involved in multiple drug resistance and metal sensitivity; ATP‐binding cassette (ABC) family member involved in drug transport; potential Cdc28p substrate; induced under respiratory conditions ‐1.2097 0.0001 TPK1 YJL164C cAMP‐dependent protein kinase catalytic subunit; promotes vegetative growth in response to nutrients via the Ras‐cAMP signaling pathway; inhibited by regulatory subunit Bcy1p in the absence of cAMP; partially redundant with Tpk2p and Tpk3p ‐1.1856 0.0004 PDR5 YOR153W Plasma membrane ATP‐binding cassette (ABC) transporter, multidrug transporter actively regulated by Pdr1p; also involved in steroid transport, cation resistance, and cellular detoxification during exponential growth ‐1.1815 0.0002 PLB2 YMR006C ‐1.1746 0.0003 KRE5 YOR336W Phospholipase B (lysophospholipase) involved in phospholipid metabolism; displays transacylase activity in vitro; overproduction confers resistance to lysophosphatidylcholine Protein required for beta‐1,6 glucan biosynthesis; mutations result in aberrant morphology and severe growth defects ‐1.1615 0.0001 RPS28B YLR264W ‐1.1388 0.0004 SMF1 YOL122C ‐1.1302 0.0003 PDR15 YDR406W Protein component of the small (40S) ribosomal subunit; nearly identical to Rps28Ap and has similarity to rat S28 ribosomal protein Divalent metal ion transporter with a broad specificity for di‐valent and tri‐valent metals; post‐translationally regulated by levels of metal ions; member of the Nramp family of metal transport proteins Plasma membrane ATP binding cassette (ABC) transporter, multidrug transporter and general stress response factor implicated in cellular detoxification; regulated by Pdr1p, Pdr3p and Pdr8p; promoter contains a PDR responsive element ‐1.1257 0.0002 NA YPR172W ‐1.1220 0.0004 SKN1 YGR143W ‐1.1034 0.0003 MDS3 YGL197W Protein of unknown function, transcriptionally activated by Yrm1p along with genes involved in multidrug resistance Protein involved in sphingolipid biosynthesis; type II membrane protein with similarity to Kre6p Putative component of the TOR regulatory pathway; negative regulator of early meiotic gene expression; required, with Pmd1p, for growth under alkaline conditions; has an N‐terminal kelch‐like domain ‐1.0982 0.0010 COA4 YLR218C Twin Cx(9)C protein involved in cytochrome c oxidase assembly or stability; localizes to the mitochondrial intermembrane space via the Mia40p‐Erv1p system; interacts genetically with CYC1 and with cytochrome c oxidase assembly factors ‐1.0981 0.0001 URA1 YKL216W Dihydroorotate dehydrogenase, catalyzes the fourth enzymatic step in the de novo biosynthesis of pyrimidines, converting dihydroorotic acid into orotic acid ‐1.0842 0.0005 ULA1 YPL003W Protein that acts together with Uba3p to activate Rub1p before its conjugation to proteins (neddylation), which may play a role in protein degradation ‐1.0828 0.0007 148 adj.p.val log2(mtr10 .Tf/WT.Tf) Symbols ORF Description NA YMR102C Protein of unknown function; transcription is activated by paralogous transcription factors Yrm1p and Yrr1p along with genes involved in multidrug resistance; mutant shows increased resistance to azoles; YMR102C is not an essential gene ‐1.0802 0.0002 NA YNL040W Putative protein of unknown function with strong similarity to alanyl‐tRNA synthases from Eubacteria; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm; YNL040W is not an essential gene ‐1.0736 0.0004 RPL18B YNL301C ‐1.0700 0.0009 YPC1 YBR183W ‐1.0673 0.0003 PXA1 YPL147W Protein component of the large (60S) ribosomal subunit, identical to Rpl18Ap and has similarity to rat L18 ribosomal protein Alkaline ceramidase that also has reverse (CoA‐independent) ceramide synthase activity, catalyzes both breakdown and synthesis of phytoceramide; overexpression confers fumonisin B1 resistance Subunit of a heterodimeric peroxisomal ATP‐binding cassette transporter complex (Pxa1p‐Pxa2p), required for import of long‐ chain fatty acids into peroxisomes; similarity to human adrenoleukodystrophy transporter and ALD‐related proteins ‐1.0534 0.0005 GPD1 YDL022W ‐1.0485 0.0008 NA YHR131C ‐1.0441 0.0005 TCA17 YEL048C ‐1.0432 0.0006 NA NA YBL029W YHL044W NAD‐dependent glycerol‐3‐phosphate dehydrogenase, key enzyme of glycerol synthesis, essential for growth under osmotic stress; expression regulated by high‐osmolarity glycerol response pathway; homolog of Gpd2p Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm; overexpression causes a cell cycle delay or arrest Protein that interacts with subunits of the TRAPP complex and may play a role its assembly or stability; mutation is synthetically lethal with gcs1 deletion; Sedlin_N family member; human Sedlin mutations cause the skeletal disorder SEDT Non‐essential protein of unknown function Putative integral membrane protein, member of DUP240 gene family; green fluorescent protein (GFP)‐fusion protein localizes to the plasma membrane in a punctate pattern ‐1.0329 ‐1.0264 0.0004 0.0007 COX11 YPL132W ‐1.0067 0.0003 NPP1 YCR026C ‐1.0040 0.0007 PPT2 YPL148C Mitochondrial inner membrane protein required for delivery of copper to the Cox1p subunit of cytochrome c oxidase; association with mitochondrial ribosomes suggests that copper delivery may occur during translation of Cox1p Nucleotide pyrophosphatase/phosphodiesterase family member; mediates extracellular nucleotide phosphate hydrolysis along with Npp2p and Pho5p; activity and expression enhanced during conditions of phosphate starvation Phosphopantetheine:protein transferase (PPTase), activates mitochondrial acyl carrier protein (Acp1p) by phosphopantetheinylation ‐0.9971 0.0002 PET111 YMR257C ‐0.9938 0.0003 FAA3 YIL009W Mitochondrial translational activator specific for the COX2 mRNA; located in the mitochondrial inner membrane Long chain fatty acyl‐CoA synthetase, activates imported fatty acids; green fluorescent protein (GFP)‐fusion protein localizes to the cell periphery ‐0.9862 0.0002 PCI8 YIL071C Possible shared subunit of Cop9 signalosome (CSN) and eIF3, binds eIF3b subunit Prt1p, has possible dual functions in transcriptional and translational control, contains a PCI (Proteasome‐COP9 signalosome (CSN)‐eIF3) domain ‐0.9855 0.0006 SCW11 YGL028C Cell wall protein with similarity to glucanases; may play a role in conjugation during mating based on its regulation by Ste12p ‐0.9825 0.0003 149 adj.p.val log2(mtr10 .Tf/WT.Tf) Symbols ORF Description RDS2 YPL133C Transcription factor involved in regulating gluconeogenesis and glyoxylate cycle genes; member of the zinc cluster family of proteins; confers resistance to ketoconazole ‐0.9706 0.0004 NA YNL234W Protein of unknown function with similarity to globins; has a functional heme‐binding domain; mutant has aneuploidy tolerance; transcription induced by stress conditions; may be involved in glucose signaling or metabolism; regulated by Rgt1 ‐0.9686 0.0010 NA YIL151C ‐0.9617 0.0005 AIM41 YOR215C Putative protein of unknown function, predicted to contain a PINc domain Putative protein of unknown function; the authentic protein is detected in highly purified mitochondria in high‐throughput studies; null mutant displays reduced frequency of mitochondrial genome loss ‐0.9591 0.0005 PCL6 YER059W Pho85p cyclin of the Pho80p subfamily; forms the major Glc8p kinase together with Pcl7p and Pho85p; involved in the control of glycogen storage by Pho85p; stabilized by Elongin C binding ‐0.9587 0.0004 YPK9 YOR291W Vacuolar protein with a possible role in sequestering heavy metals; has similarity to the type V P‐type ATPase Spf1p; homolog of human ATP13A2 (PARK9), mutations in which are associated with Parkinson disease and Kufor‐Rakeb syndrome ‐0.9567 0.0002 NA YGR122W Probable ortholog of A. nidulans PalC, which is involved in pH regulation and binds to the ESCRT‐III complex; null mutant does not properly process Rim101p and has decreased resistance to rapamycin; GFP‐fusion protein is cytoplasmic ‐0.9528 0.0004 BNA5 YLR231C ‐0.9440 0.0005 MNN1 YER001W Kynureninase, required for the de novo biosynthesis of NAD from tryptophan via kynurenine; expression regulated by Hst1p Alpha‐1,3‐mannosyltransferase, integral membrane glycoprotein of the Golgi complex, required for addition of alpha1,3‐mannose linkages to N‐linked and O‐linked oligosaccharides, one of five S. cerevisiae proteins of the MNN1 family ‐0.9379 0.0004 VPS62 YGR141W ‐0.9327 0.0005 CMC4 YMR194C‐ B ‐0.9317 0.0008 CMK1 YFR014C Calmodulin‐dependent protein kinase; may play a role in stress response, many CA++/calmodulan dependent phosphorylation substrates demonstrated in vitro, amino acid sequence similar to Cmk2p and mammalian Cam Kinase II ‐0.9263 0.0004 RRN10 YBL025W ‐0.9259 0.0003 MRF1 YGL143C Protein involved in promoting high level transcription of rDNA, subunit of UAF (upstream activation factor) for RNA polymerase I Mitochondrial translation release factor, involved in stop codon recognition and hydrolysis of the peptidyl‐tRNA bond during mitochondrial translation; lack of MRF1 causes mitochondrial genome instability ‐0.9250 0.0003 NA YOR292C Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the vacuole; YOR292C is not an essential gene ‐0.9242 0.0002 NA YPR147C Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm and is induced in response to the DNA‐damaging agent MMS ‐0.9219 0.0002 Vacuolar protein sorting (VPS) protein required for cytoplasm to vacuole targeting of proteins Protein that localizes to the mitochondrial intermembrane space via the Mia40p‐Erv1p system; contains twin cysteine‐x(9)‐cysteine motifs 150 adj.p.val log2(mtr10 .Tf/WT.Tf) Symbols ORF Description adj.p.val NA YBL095W Putative protein of unknown function; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐ throughput studies ‐0.9141 0.0002 COX10 YPL172C ‐0.9025 0.0005 LAG1 YHL003C ‐0.9025 0.0005 PSY2 YNL201C Heme A:farnesyltransferase, catalyzes the first step in the conversion of protoheme to the heme A prosthetic group required for cytochrome c oxidase activity; human ortholog is associated with mitochondrial disorders Ceramide synthase component, involved in synthesis of ceramide from C26(acyl)‐coenzyme A and dihydrosphingosine or phytosphingosine, functionally equivalent to Lac1p Putative subunit of an evolutionarily conserved protein phosphatase complex containing the catalytic subunit Pph3p and the regulatory subunit Psy4p; required for cisplatin and oxaliplatin resistance; putative homolog of mammalian R3 ‐0.9007 0.0003 CPS1 YJL172W Vacuolar carboxypeptidase yscS; expression is induced under low‐ nitrogen conditions Putative S‐adenosylmethionine‐dependent methyltransferase ‐0.8991 0.0010 ‐0.8976 0.0002 Phosphoesterase involved in downregulation of the unfolded protein response, at least in part via dephosphorylation of Ire1p; dosage‐ dependent positive regulator of the G1/S phase transition through control of the timing of START Choline kinase, catalyzing the first step in phosphatidylcholine synthesis via the CDP‐choline (Kennedy pathway); exhibits some ethanolamine kinase activity contributing to phosphatidylethanolamine synthesis via the CDP‐ethanolamine pathway Putative protein of unknown function, has some homology to Ugp1p, which encodes UDP‐glucose pyrophosphorylase Putative protein of unknown function; expression repressed in carbon limited vs carbon replete chemostat cultures; YJR030C is a non‐essential gene ‐0.8970 0.0009 ‐0.8898 0.0004 ‐0.8837 0.0004 ‐0.8808 0.0003 NA YLR137W DCR2 YLR361C CKI1 YLR133W NA YHL012W NA YJR030C RTT105 YGK3 YER104W YOL128C Protein with a role in regulation of Ty1 transposition Protein kinase related to mammalian glycogen synthase kinases of the GSK‐3 family; GSK‐3 homologs (Mck1p, Rim11p, Mrk1p, Ygk3p) are involved in control of Msn2p‐dependent transcription of stress responsive genes and in protein degradation ‐0.8801 ‐0.8783 0.0009 0.0007 NA YMR262W ‐0.8782 0.0005 AIM22 YJL046W ‐0.8759 0.0003 NA AFT2 YLR031W YPL202C ‐0.8722 ‐0.8697 0.0004 0.0002 ARI1 YGL157W Protein of unknown function; interacts weakly with Knr4p; YMR262W is not an essential gene Putative lipoate‐protein ligase, required along with Lip2 and Lip5 for lipoylation of Lat1p and Kgd2p; similar to E. coli LplA; null mutant displays reduced frequency of mitochondrial genome loss Putative protein of unknown function Iron‐regulated transcriptional activator; activates genes involved in intracellular iron use and required for iron homeostasis and resistance to oxidative stress; similar to Aft1p NADPH‐dependent aldehyde reductase, utilizes aromatic and alophatic aldehyde substrates; member of the short‐chain dehydrogenase/reductase superfamily ‐0.8694 0.0003 SPI1 YER150W ‐0.8671 0.0006 SMF3 YLR034C GPI‐anchored cell wall protein involved in weak acid resistance; basal expression requires Msn2p/Msn4p; expression is induced under conditions of stress and during the diauxic shift; similar to Sed1p Putative divalent metal ion transporter involved in iron homeostasis; transcriptionally regulated by metal ions; member of the Nramp family of metal transport proteins ‐0.8656 0.0005 NA YPR013C ‐0.8648 0.0002 Putative zinc finger protein; YPR013C is not an essential gene 151 log2(mtr10 .Tf/WT.Tf) Symbols ORF Description YOR1 YGR281W Plasma membrane ATP‐binding cassette (ABC) transporter, multidrug transporter mediates export of many different organic anions including oligomycin; similar to human cystic fibrosis transmembrane receptor (CFTR) ‐0.8613 0.0002 NA YER130C ‐0.8566 0.0004 ELO1 YJL196C Protein of unknown function; transcription is regulated by Haa1p, Sok2p and Zap1p transcriptional activators; the C. Albicans homolog (MNL1) plays a role in adaptation to stress Elongase I, medium‐chain acyl elongase, catalyzes carboxy‐terminal elongation of unsaturated C12‐C16 fatty acyl‐CoAs to C16‐C18 fatty acids ‐0.8537 0.0002 MSF1 YPR047W Mitochondrial phenylalanyl‐tRNA synthetase, active as a monomer, unlike the cytoplasmic subunit which is active as a dimer complexed to a beta subunit dimer; similar to the alpha subunit of E. coli phenylalanyl‐tRNA synthetase ‐0.8411 0.0003 TRZ1 YKR079C ‐0.8386 0.0004 NA YKR070W tRNA 3'‐end processing endonuclease tRNase Z; also localized to mitochondria and interacts genetically with Rex2 exonuclease; homolog of the human candidate prostate cancer susceptibility gene ELAC2 Putative protein of unknown function; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐ throughput studies ‐0.8352 0.0009 SPT14 YPL175W UDP‐GlcNAc‐binding and catalytic subunit of the enzyme that mediates the first step in glycosylphosphatidylinositol (GPI) biosynthesis, mutations cause defects in transcription and in biogenesis of cell wall proteins ‐0.8336 0.0005 NA YGL140C ‐0.8322 0.0002 OCT1 YKL134C Putative protein of unknown function; non‐essential gene; contains multiple predicted transmembrane domains Mitochondrial intermediate peptidase, cleaves N‐terminal residues of a subset of proteins upon import, after their cleavage by mitochondrial processing peptidase (Mas1p‐Mas2p); may contribute to mitochondrial iron homeostasis ‐0.8304 0.0005 FLX1 YIL134W ‐0.8250 0.0010 NMA1 YLR328W ‐0.8160 0.0002 RMR1 YGL250W ‐0.8142 0.0004 BUR2 YLR226W Cyclin for the Sgv1p (Bur1p) protein kinase; Sgv1p and Bur2p comprise a CDK‐cyclin complex involved in transcriptional regulation through its phosphorylation of the carboxy‐terminal domain of the largest subunit of RNA polymerase II ‐0.8083 0.0002 TOR1 YJR066W PIK‐related protein kinase and rapamycin target; subunit of TORC1, a complex that controls growth in response to nutrients by regulating translation, transcription, ribosome biogenesis, nutrient transport and autophagy; involved in meiosis ‐0.8081 0.0008 RIM15 YFL033C Glucose‐repressible protein kinase involved in signal transduction during cell proliferation in response to nutrients, specifically the establishment of stationary phase; identified as a regulator of IME2; substrate of Pho80p‐Pho85p kinase ‐0.8070 0.0007 RTC4 YNL254C Protein of unknown function; null mutation suppresses cdc13‐1 temperature sensitivity; (GFP)‐fusion protein localizes to both the cytoplasm and the nucleus ‐0.7990 0.0006 Protein required for transport of flavin adenine dinucleotide (FAD), a synthesis product of riboflavin, across the mitochondrial membrane Nicotinic acid mononucleotide adenylyltransferase, involved in pathways of NAD biosynthesis, including the de novo, NAD(+) salvage, and nicotinamide riboside salvage pathways Protein required for meiotic recombination and gene conversion; null mutant displays reduced PIS1 expression and growth defects on non‐fermentable carbon sources and minimal media; GFP‐fusion protein localizes to both cytoplasm and nucleus 152 adj.p.val log2(mtr10 .Tf/WT.Tf) Symbols ORF Description UBR2 YLR024C ‐0.7978 0.0009 POL12 YBL035C ‐0.7921 0.0004 RIB4 YOL143C ‐0.7916 0.0002 GPI12 YMR281W Cytoplasmic ubiquitin‐protein ligase (E3); required for ubiquitylation of Rpn4p; mediates formation of a Mub1p‐Ubr2p‐Rad6p complex B subunit of DNA polymerase alpha‐primase complex, required for initiation of DNA replication during mitotic and premeiotic DNA synthesis; also functions in telomere capping and length regulation Lumazine synthase (6,7‐dimethyl‐8‐ribityllumazine synthase, also known as DMRL synthase); catalyzes synthesis of immediate precursor to riboflavin ER membrane protein involved in the second step of glycosylphosphatidylinositol (GPI) anchor assembly, the de‐N‐ acetylation of the N‐acetylglucosaminylphosphatidylinositol intermediate; functional homolog of human PIG‐Lp ‐0.7909 0.0009 RIM2 YBR192W ‐0.7889 0.0004 RPL8A YHL033C ‐0.7880 0.0005 MGA1 YGR249W Mitochondrial pyrimidine nucleotide transporter; imports pyrimidine nucleoside triphosphates and exports pyrimidine nucleoside monophosphates; member of the mitochondrial carrier family Ribosomal protein L4 of the large (60S) ribosomal subunit, nearly identical to Rpl8Bp and has similarity to rat L7a ribosomal protein; mutation results in decreased amounts of free 60S subunits Protein similar to heat shock transcription factor; multicopy suppressor of pseudohyphal growth defects of ammonium permease mutants ‐0.7877 0.0008 MDM32 YOR147W ‐0.7823 0.0006 RAD16 YBR114W Mitochondrial inner membrane protein with similarity to Mdm31p, required for normal mitochondrial morphology and inheritance; interacts genetically with MMM1, MDM10, MDM12, and MDM34 Protein that recognizes and binds damaged DNA in an ATP‐ dependent manner (with Rad7p) during nucleotide excision repair; subunit of Nucleotide Excision Repair Factor 4 (NEF4) and the Elongin‐Cullin‐Socs (ECS) ligase complex ‐0.7820 0.0002 CAX4 YGR036C Dolichyl pyrophosphate (Dol‐P‐P) phosphatase with a luminally oriented active site in the ER, cleaves the anhydride linkage in Dol‐P‐ P, required for Dol‐P‐P‐linked oligosaccharide intermediate synthesis and protein N‐glycosylation ‐0.7805 0.0004 COS9 YKL219W ‐0.7794 0.0009 NA YCR015C ‐0.7785 0.0002 MNS1 YJR131W Protein of unknown function, member of the DUP380 subfamily of conserved, often subtelomerically‐encoded proteins Putative protein of unknown function; YCR015C is not an essential gene Alpha‐1,2‐mannosidase involved in ER quality control; catalyzes the removal of one mannose residue from Man9GlcNAc to produce a single isomer of Man8GlcNAc in N‐linked oligosaccharide biosynthesis; integral to ER membrane ‐0.7763 0.0007 AKR2 YOR034C Ankyrin repeat‐containing protein similar to Akr1p; member of a family of putative palmitoyltransferases containing an Asp‐His‐His‐ Cys‐cysteine rich (DHHC‐CRD) domain; possibly involved in constitutive endocytosis of Ste3p ‐0.7750 0.0005 TNA1 YGR260W High affinity nicotinic acid plasma membrane permease, responsible for uptake of low levels of nicotinic acid; expression of the gene increases in the absence of extracellular nicotinic acid or para‐ aminobenzoate (PABA) ‐0.7745 0.0004 NA YMR210W ‐0.7716 0.0007 YAP5 NA YIR018W YBR220C Putative acyltransferase with similarity to Eeb1p and Eht1p, has a minor role in medium‐chain fatty acid ethyl ester biosynthesis; may be involved in lipid metabolism and detoxification Basic leucine zipper (bZIP) transcription factor Putative protein of unknown function; YBR220C is not an essential gene ‐0.7633 ‐0.7625 0.0003 0.0004 153 adj.p.val log2(mtr10 .Tf/WT.Tf) Symbols ORF Description NA YMR010W ‐0.7545 0.0009 TUF1 YOR187W Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm; YMR010W is not an essential gene; YMR010W mRNA is transcribed with ADI1 Mitochondrial translation elongation factor Tu; comprises both GTPase and guanine nucleotide exchange factor activities, while these activities are found in separate proteins in S. pombe and humans ‐0.7500 0.0003 GDT1 YBR187W Putative protein of unknown function; expression is reduced in a gcr1 null mutant; GFP‐fusion protein localizes to the vacuole; expression pattern and physical interactions suggest a possible role in ribosome biogenesis ‐0.7455 0.0003 NA VPS75 YOR390W YNL246W Putative protein of unknown function NAP family histone chaperone; binds to histones and Rtt109p, stimulating histone acetyltransferase activity; possesses nucleosome assembly activity in vitro; proposed role in vacuolar protein sorting and in double‐strand break repair ‐0.7427 ‐0.7414 0.0004 0.0004 AMD1 YML035C ‐0.7304 0.0006 MRPS35 HEK2 YGR165W YBL032W AMP deaminase, tetrameric enzyme that catalyzes the deamination of AMP to form IMP and ammonia; may be involved in regulation of intracellular adenine nucleotide pools Mitochondrial ribosomal protein of the small subunit RNA binding protein involved in the asymmetric localization of ASH1 mRNA; represses translation of ASH1 mRNA, an effect reversed by Yck1p‐dependent phosphoryation; regulates telomere position effect and length; similarity to hnRNP‐K ‐0.7297 ‐0.7296 0.0009 0.0004 ATP2 YJR121W ‐0.7208 0.0008 MSG5 YNL053W Beta subunit of the F1 sector of mitochondrial F1F0 ATP synthase, which is a large, evolutionarily conserved enzyme complex required for ATP synthesis; phosphorylated Dual‐specificity protein phosphatase; exists in 2 isoforms; required for maintenance of a low level of signaling through the cell integrity pathway, adaptive response to pheromone; regulates and is regulated by Slt2p; dephosphorylates Fus3p ‐0.7194 0.0005 ARO4 YBR249C ‐0.7156 0.0009 STE4 YOR212W 3‐deoxy‐D‐arabino‐heptulosonate‐7‐phosphate (DAHP) synthase, catalyzes the first step in aromatic amino acid biosynthesis and is feedback‐inhibited by tyrosine or high concentrations of phenylalanine or tryptophan G protein beta subunit, forms a dimer with Ste18p to activate the mating signaling pathway, forms a heterotrimer with Gpa1p and Ste18p to dampen signaling; may recruit Rho1p to the polarized growth site during mating; contains WD40 repeats ‐0.7112 0.0009 SCS7 YMR272C ‐0.7091 0.0008 SCS3 YGL126W Sphingolipid alpha‐hydroxylase, functions in the alpha‐hydroxylation of sphingolipid‐associated very long chain fatty acids, has both cytochrome b5‐like and hydroxylase/desaturase domains, not essential for growth Protein required for inositol prototrophy, identified as an ortholog of the FIT family of proteins involved in triglyceride droplet biosynthesis; disputed role in the synthesis of inositol phospholipids from inositol ‐0.7065 0.0009 MKK1 YOR231W Mitogen‐activated kinase kinase involved in protein kinase C signaling pathway that controls cell integrity; upon activation by Bck1p phosphorylates downstream target, Slt2p; functionally redundant with Mkk2p ‐0.7036 0.0003 SSY5 YJL156C Serine protease of SPS plasma membrane amino acid sensor system (Ssy1p‐Ptr3p‐Ssy5p); contains an inhibitory domain that dissociates in response to extracellular amino acids, freeing a catalytic domain to activate transcription factor Stp1p ‐0.7007 0.0008 154 adj.p.val log2(mtr10 .Tf/WT.Tf) Symbols ORF Description VBA1 MNP1 YMR088C YGL068W ‐0.6986 ‐0.6984 0.0009 0.0003 LEU5 YHR002W ‐0.6910 0.0004 PET117 COT1 YER058W YOR316C ‐0.6893 ‐0.6873 0.0005 0.0009 BLM10 YFL007W Permease of basic amino acids in the vacuolar membrane Protein associated with the mitochondrial nucleoid; putative mitochondrial ribosomal protein with similarity to E. coli L7/L12 ribosomal protein; required for normal respiratory growth Mitochondrial carrier protein involved in the accumulation of CoA in the mitochondrial matrix; homolog of human Graves disease protein; does not encode an isozyme of Leu4p, as first hypothesized Protein required for assembly of cytochrome c oxidase Vacuolar transporter that mediates zinc transport into the vacuole; overexpression confers resistance to cobalt and rhodium Proteasome activator subunit; found in association with core particles, with and without the 19S regulatory particle; required for resistance to bleomycin, may be involved in protecting against oxidative damage; similar to mammalian PA200 ‐0.6859 0.0004 NA YMR291W Putative kinase of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm and nucleus; YMR291W is not an essential gene ‐0.6857 0.0007 NA YLR278C Zinc‐cluster protein; GFP‐fusion protein localizes to the nucleus; mutant shows moderate growth defect on caffeine; has a prion‐ domain like fragment that increases frequency of [URE3]; YLR278C is not an essential gene ‐0.6741 0.0004 YLF2 YHL014C ‐0.6738 0.0010 CIN2 YPL241C Protein of unknown function, has weak similarity to E. coli GTP‐ binding protein gtp1; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐throughput studies GTPase‐activating protein (GAP) for Cin4p; tubulin folding factor C involved in beta‐tubulin (Tub2p) folding; mutants display increased chromosome loss and benomyl sensitivity; deletion complemented by human GAP, retinitis pigmentosa 2 ‐0.6734 0.0007 NA POL2 YOL098C YNL262W Putative metalloprotease Catalytic subunit of DNA polymerase (II) epsilon, a chromosomal DNA replication polymerase that exhibits processivity and proofreading exonuclease activity; also involved in DNA synthesis during DNA repair; interacts extensively with Mrc1p ‐0.6659 ‐0.6624 0.0006 0.0009 SAT4 YCR008W Ser/Thr protein kinase involved in salt tolerance; funtions in regulation of Trk1p‐Trk2p potassium transporter; partially redundant with Hal5p; has similarity to Npr1p ‐0.6603 0.0009 HIR2 YOR038C Subunit of the HIR complex, a nucleosome assembly complex involved in regulation of histone gene transcription; recruits Swi‐Snf complexes to histone gene promoters; promotes heterochromatic gene silencing with Asf1p ‐0.6587 0.0009 CAN1 YEL063C ‐0.6553 0.0003 PSD1 YNL169C Plasma membrane arginine permease, requires phosphatidyl ethanolamine (PE) for localization, exclusively associated with lipid rafts; mutation confers canavanine resistance Phosphatidylserine decarboxylase of the mitochondrial inner membrane, converts phosphatidylserine to phosphatidylethanolamine ‐0.6495 0.0006 DIA4 YHR011W ‐0.6491 0.0005 KGD1 YIL125W Probable mitochondrial seryl‐tRNA synthetase, mutant displays increased invasive and pseudohyphal growth Component of the mitochondrial alpha‐ketoglutarate dehydrogenase complex, which catalyzes a key step in the tricarboxylic acid (TCA) cycle, the oxidative decarboxylation of alpha‐ ketoglutarate to form succinyl‐CoA ‐0.6467 0.0006 MRPL6 YHR147C Mitochondrial ribosomal protein of the large subunit ‐0.6456 0.0008 155 adj.p.val log2(mtr10 .Tf/WT.Tf) Symbols ORF Description POR2 YIL114C PET127 YOR017W RPO41 YFL036W EGT2 YNL327W Putative mitochondrial porin (voltage‐dependent anion channel), related to Por1p but not required for mitochondrial membrane permeability or mitochondrial osmotic stability Protein with a role in 5'‐end processing of mitochondrial RNAs, located in the mitochondrial membrane Mitochondrial RNA polymerase; single subunit enzyme similar to those of T3 and T7 bacteriophages; requires a specificity subunit encoded by MTF1 for promoter recognition Glycosylphosphatidylinositol (GPI)‐anchored cell wall endoglucanase required for proper cell separation after cytokinesis, expression is activated by Swi5p and tightly regulated in a cell cycle‐dependent manner CDH1 YGL003C PEX32 adj.p.val ‐0.6446 0.0004 ‐0.6425 0.0007 ‐0.6413 0.0005 ‐0.6364 0.0005 Cell‐cycle regulated activator of the anaphase‐promoting complex/cyclosome (APC/C), which directs ubiquitination of cyclins resulting in mitotic exit; targets the APC/C to specific substrates including Cdc20p, Ase1p, Cin8p and Fin1p ‐0.6317 0.0009 YBR168W Peroxisomal integral membrane protein, involved in negative regulation of peroxisome size; partially functionally redundant with Pex31p; genetic interactions suggest action at a step downstream of steps mediated by Pex28p and Pex29p ‐0.6298 0.0004 OST6 YML019W ‐0.6281 0.0008 PIR1 YKL164C Subunit of the oligosaccharyltransferase complex of the ER lumen, which catalyzes asparagine‐linked glycosylation of newly synthesized proteins; similar to and partially functionally redundant with Ost3p O‐glycosylated protein required for cell wall stability; attached to the cell wall via beta‐1,3‐glucan; mediates mitochondrial translocation of Apn1p; expression regulated by the cell integrity pathway and by Swi5p during the cell cycle ‐0.6244 0.0006 CDS1 YBR029C Phosphatidate cytidylyltransferase (CDP‐diglyceride synthetase); an enzyme that catalyzes that conversion of CTP + phosphate into diphosphate + CDP‐diaclglyerol, a critical step in the synthesis of all major yeast phospholipids ‐0.6241 0.0010 PEX2 YJL210W RING‐finger peroxin and E3 ubiquitin ligase, peroxisomal membrane protein with a C‐terminal zinc‐binding RING domain, forms translocation subcomplex with Pex10p and Pex12p which functions in peroxisomal matrix protein import ‐0.6189 0.0010 FAA1 YOR317W Long chain fatty acyl‐CoA synthetase, activates imported fatty acids with a preference for C12:0‐C16:0 chain lengths; functions in long chain fatty acid import; accounts for most acyl‐CoA synthetase activity; localized to lipid particles ‐0.6161 0.0006 IGO2 YHR132W‐ A Protein required for initiation of G0 program; prevents degradation of nutrient‐regulated mRNAs via the 5'‐3' mRNA decay pathway; phosphorylated by Rim15p; GFP protein localizes to the cytoplasm and nucleus; similar to Igo1p ‐0.6088 0.0009 NA YKL100C Putative protein of unknown function with similarity to a human minor histocompatibility antigen and signal peptide peptidases; YKL100C is not an essential gene ‐0.6087 0.0008 CFT2 YLR115W ‐0.6062 0.0008 NA YOR093C Subunit of the mRNA cleavage and polyadenlylation factor (CPF); required for pre‐mRNA cleavage, polyadenylation and poly(A) site recognition, 43% similarity with the mammalian CPSF‐100 protein. Putative protein of unknown function; deletion causes sensitivity to unfolded protein response‐inducing agents ‐0.6029 0.0007 156 APPENDIX H: Up-regulated genes in mtr10Δ cells in fed condition This section includes the list of transcriptionally differentially expressed genes (cutoff conditions: log2 FC 0.6 and adjusted p-value < 0.001) in mtr10Δ cells in fed condition. Up-regulated genes in mtr10Δ cells in fed condition log2(mtr10 .Tf/WT.Tf) adj.p.va l Adhesion subunit of a‐agglutinin of a‐cells, C‐terminal sequence acts as a ligand for alpha‐agglutinin (Sag1p) during agglutination, modified with O‐linked oligomannosyl chains, linked to anchorage subunit Aga1p via two disulfide bonds 3.6923 0.0001 YDR461W Mating pheromone a‐factor, made by a cells; interacts with alpha cells to induce cell cycle arrest and other responses leading to mating; biogenesis involves C‐terminal modification, N‐terminal proteolysis, and export; also encoded by MFA2 3.5838 0.0000 NA YDL114W 3.2686 0.0000 MFA2 YNL145W Putative protein of unknown function with similarity to acyl‐carrier‐ protein reductases; YDL114W is not an essential gene Mating pheromone a‐factor, made by a cells; interacts with alpha cells to induce cell cycle arrest and other responses leading to mating; biogenesis involves C‐terminal modification, N‐terminal proteolysis, and export; also encoded by MFA1 3.2579 0.0001 STL1 YDR536W 3.2372 0.0000 ARO10 YDR380W Glycerol proton symporter of the plasma membrane, subject to glucose‐induced inactivation, strongly but transiently induced when cells are subjected to osmotic shock Phenylpyruvate decarboxylase, catalyzes decarboxylation of phenylpyruvate to phenylacetaldehyde, which is the first specific step in the Ehrlich pathway 2.9917 0.0004 Symbols ORF Description AGA2 YGL032C MFA1 NA YNR062C Putative membrane protein of unknown function 2.9628 0.0000 SRT1 YMR101C Cis‐prenyltransferase involved in synthesis of long‐chain dolichols (19‐22 isoprene units; as opposed to Rer2p which synthesizes shorter‐chain dolichols); localizes to lipid bodies; transcription is induced during stationary phase 2.6672 0.0000 SPG4 YMR107W 2.6514 0.0002 NA YDR374C Protein required for survival at high temperature during stationary phase; not required for growth on nonfermentable carbon sources Putative protein of unknown function 2.5782 0.0000 157 log2(mtr10 .Tf/WT.Tf) adj.p.va l Putative protein of unknown function; transcription dependent upon Azf1p Aromatic aminotransferase II, catalyzes the first step of tryptophan, phenylalanine, and tyrosine catabolism Sporulation‐specific homolog of the yeast CDC3/10/11/12 family of bud neck microfilament genes; septin protein involved in sporulation; regulated by ABFI 2.4575 0.0002 2.3991 0.0002 2.3575 0.0001 YDR034W‐B Predicted tail‐anchored plasma membrane protein containing a conserved CYSTM module; related proteins in other organisms may be involved in response to stress; green fluorescent protein (GFP)‐ fusion protein localizes to the cell periphery 2.2665 0.0000 CDA1 YLR307W Chitin deacetylase, together with Cda2p involved in the biosynthesis ascospore wall component, chitosan; required for proper rigidity of the ascospore wall 2.2479 0.0001 RNP1 YLL046C Ribonucleoprotein that contains two RNA recognition motifs (RRM) 2.1298 0.0002 NA YNR064C 2.1183 0.0000 CSN9 YDR179C Epoxide hydrolase, member of the alpha/beta hydrolase fold family; may have a role in detoxification of epoxides Subunit of the Cop9 signalosome, which is required for deneddylation, or removal of the ubiquitin‐like protein Rub1p from Cdc53p (cullin); involved in adaptation to pheromone signaling 2.1149 0.0001 ECM8 YBR076W Non‐essential protein of unknown function 2.0530 0.0001 BOP2 YLR267W Protein of unknown function 2.0185 0.0001 FMP16 YDR070C Putative protein of unknown function; proposed to be involved in responding to conditions of stress; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐ throughput studies 1.9711 0.0007 DAL80 YKR034W Negative regulator of genes in multiple nitrogen degradation pathways; expression is regulated by nitrogen levels and by Gln3p; member of the GATA‐binding family, forms homodimers and heterodimers with Deh1p 1.9184 0.0002 NA YBR056W‐A Dubious open reading frame unlikely to encode a protein, based on available experimental and comparative sequence data; partially overlaps the dubious ORF YBR056C‐B 1.8641 0.0002 DIT1 YDR403W Sporulation‐specific enzyme required for spore wall maturation, involved in the production of a soluble LL‐dityrosine‐containing precursor of the spore wall; transcripts accumulate at the time of prospore enclosure 1.7701 0.0002 NA YDR042C 1.7677 0.0000 NA YFR012W‐A Putative protein of unknown function; expression is increased in ssu72‐ts69 mutant Putative protein of unknown function; identified by homology 1.7649 0.0001 PES4 YFR023W 1.7269 0.0001 PRR2 YDL214C Poly(A) binding protein, suppressor of DNA polymerase epsilon mutation, similar to Mip6p Serine/threonine protein kinase that inhibits pheromone induced signalling downstream of MAPK, possibly at the level of the Ste12p transcription factor; mutant has increased aneuploidy tolerance 1.7018 0.0000 ZIP1 YDR285W Transverse filament protein of the synaptonemal complex; required for normal levels of meiotic recombination and pairing between homologous chromosome during meiosis; potential Cdc28p substrate 1.7002 0.0001 NA YJR149W Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm 1.6709 0.0001 Symbols ORF Description ANS1 YHR126C ARO9 YHR137W SPR3 YGR059W NA 158 log2(mtr10 .Tf/WT.Tf) adj.p.va l Putative transmembrane protein involved in export of ammonia, a starvation signal that promotes cell death in aging colonies; phosphorylated in mitochondria; member of the TC 9.B.33 YaaH family; homolog of Ady2p and Y. lipolytica Gpr1p 1.6697 0.0003 YLL057C Fe(II)‐dependent sulfonate/alpha‐ketoglutarate dioxygenase, involved in sulfonate catabolism for use as a sulfur source; contains sequence that resembles a J domain (typified by the E. coli DnaJ protein); induced by sulphur starvation 1.6563 0.0002 BNA2 YJR078W Putative tryptophan 2,3‐dioxygenase or indoleamine 2,3‐ dioxygenase, required for de novo biosynthesis of NAD from tryptophan via kynurenine; interacts genetically with telomere capping gene CDC13; regulated by Hst1p and Aftp 1.6391 0.0003 LYS20 YDL182W Homocitrate synthase isozyme, catalyzes the condensation of acetyl‐ CoA and alpha‐ketoglutarate to form homocitrate, which is the first step in the lysine biosynthesis pathway; highly similar to the other isozyme, Lys21p 1.6276 0.0002 MCH2 YKL221W Protein with similarity to mammalian monocarboxylate permeases, which are involved in transport of monocarboxylic acids across the plasma membrane; mutant is not deficient in monocarboxylate transport 1.6150 0.0005 HXT9 YJL219W Putative hexose transporter that is nearly identical to Hxt11p, has similarity to major facilitator superfamily (MFS) transporters, expression of HXT9 is regulated by transcription factors Pdr1p and Pdr3p 1.6118 0.0004 NA YGL015C 1.6100 0.0001 BAR1 YIL015W Putative protein of unknown function; null mutants accumulate cargo in the Golgi Aspartyl protease secreted into the periplasmic space of mating type a cells, helps cells find mating partners, cleaves and inactivates alpha factor allowing cells to recover from alpha‐factor‐induced cell cycle arrest 1.5974 0.0005 PUT1 YLR142W Proline oxidase, nuclear‐encoded mitochondrial protein involved in utilization of proline as sole nitrogen source; PUT1 transcription is induced by Put3p in the presence of proline and the absence of a preferred nitrogen source 1.5578 0.0001 NA YML083C Putative protein of unknown function; strong increase in transcript abundance during anaerobic growth compared to aerobic growth; cells deleted for YML083C do not exhibit growth defects in anerobic or anaerobic conditions 1.5547 0.0001 DON1 YDR273W 1.5545 0.0001 TFB5 YDR079C‐A Meiosis‐specific component of the spindle pole body, part of the leading edge protein (LEP) coat, forms a ring‐like structure at the leading edge of the prospore membrane during meiosis II Component of the RNA polymerase II general transcription and DNA repair factor TFIIH; involved in transcription initiation and in nucleotide‐excision repair; homolog of Chlamydomonas reinhardtii REX1‐S protein involved in DNA repair 1.5532 0.0001 GSC2 YGR032W Catalytic subunit of 1,3‐beta‐glucan synthase, involved in formation of the inner layer of the spore wall; activity positively regulated by Rho1p and negatively by Smk1p; has similarity to an alternate catalytic subunit, Fks1p (Gsc1p) 1.5527 0.0001 LOH1 YJL038C Protein of unknown function with proposed roles in maintenance of genome integrity and also in spore wall assembly; induced during sporulation; repressed during vegetative growth by Sum1p and Hst1p; sequence similar to IRC1 1.5458 0.0002 Symbols ORF Description ATO2 YNR002C JLP1 159 log2(mtr10 .Tf/WT.Tf) adj.p.va l Putative protein of unknown function; YDL186W is not an essential gene Transcriptional regulator involved in activation of the GAL genes in response to galactose; forms a complex with Gal80p to relieve Gal80p inhibition of Gal4p; binds galactose and ATP but does not have galactokinase activity 1.5209 0.0002 1.5134 0.0000 Putative protein of unknown function; similar to the mouse IMPACT gene; YDL177C is not an essential gene Permease that serves as a gamma‐aminobutyrate (GABA) transport protein involved in the utilization of GABA as a nitrogen source; catalyzes the transport of putrescine and delta‐aminolevulinic acid (ALA); localized to the vacuolar membrane 1.4974 0.0000 1.4882 0.0001 YGL018C Specialized J‐protein that functions with Hsp70 in Fe‐S cluster biogenesis in mitochondria, involved in iron metabolism; contains a J domain typical to J‐type chaperones; localizes to the mitochondrial matrix 1.4697 0.0000 SMA1 YPL027W 1.4625 0.0006 MGT1 YDL200C 1.4582 0.0001 DIT2 YDR402C 1.4464 0.0001 KRE28 YDR532C Protein of unknown function involved in the assembly of the prospore membrane during sporulation; interacts with Spo14p DNA repair methyltransferase (6‐O‐methylguanine‐DNA methylase) involved in protection against DNA alkylation damage N‐formyltyrosine oxidase, sporulation‐specific microsomal enzyme involved in the production of N,N‐bisformyl dityrosine required for spore wall maturation, homologous to cytochrome P‐450s Subunit of a kinetochore‐microtubule binding complex with Spc105p that bridges centromeric heterochromatin and kinetochore MAPs and motors, and is also required for sister chromatid bi‐orientation and kinetochore binding of SAC components 1.3761 0.0003 DIA3 YDL024C 1.3759 0.0002 STF1 YDL130W‐A 1.3700 0.0001 CWC15 YDR163W 1.3445 0.0001 IRC18 YJL037W 1.3377 0.0004 PLM2 YDR501W Forkhead Associated domain containing protein and putative transcription factor found associated with chromatin; target of SBF transcription factor; induced in response to DNA damaging agents and deletion of telomerase; similar to TOS4 1.3218 0.0001 CYB2 YML054C Cytochrome b2 (L‐lactate cytochrome‐c oxidoreductase), component of the mitochondrial intermembrane space, required for lactate utilization; expression is repressed by glucose and anaerobic conditions 1.3192 0.0001 BUG1 YDL099W Cis‐golgi localized protein involved in ER to Golgi transport; forms a complex with the mammalian GRASP65 homolog, Grh1p; mutants are compromised for the fusion of ER‐derived vesicles with Golgi membranes 1.3176 0.0002 PAR32 YDL173W Putative protein of unknown function; hyperphosphorylated upon rapamycin treatment in a Tap42p‐dependent manner; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm; PAR32 is not an essential gene 1.3165 0.0001 Symbols ORF Description NA YDL186W GAL3 YDR009W NA YDL177C UGA4 YDL210W JAC1 Protein of unknown function, involved in invasive and pseudohyphal growth Protein involved in regulation of the mitochondrial F1F0‐ATP synthase; Stf1p and Stf2p may act as stabilizing factors that enhance inhibitory action of the Inh1p protein Non‐essential protein involved in pre‐mRNA splicing, component of a complex containing Cef1p; has similarity to S. pombe Cwf15p Putative protein of unknown function; expression induced in respiratory‐deficient cells and in carbon‐limited chemostat cultures; similar to adjacent ORF, YJL038C; null mutant displays increased levels of spontaneous Rad52p foci 160 log2(mtr10 .Tf/WT.Tf) adj.p.va l 1.2996 0.0002 1.2986 0.0004 1.2845 0.0002 1.2753 0.0001 Plasma membrane sulfite pump involved in sulfite metabolism and required for efficient sulfite efflux; major facilitator superfamily protein 1.2676 0.0004 Homocitrate synthase isozyme, catalyzes the condensation of acetyl‐ CoA and alpha‐ketoglutarate to form homocitrate, which is the first step in the lysine biosynthesis pathway; highly similar to the other isozyme, Lys20p 1.2633 0.0006 Symbols ORF Description RAD34 YDR314C MAL32 YBR299W Protein involved in nucleotide excision repair (NER); homologous to RAD4 Maltase (alpha‐D‐glucosidase), inducible protein involved in maltose catabolism; encoded in the MAL3 complex locus; functional in genomic reference strain S288C; hydrolyzes the disaccharides maltose, turanose, maltotriose, and sucrose CRR1 YLR213C PAU4 YLR461W SSU1 YPL092W LYS21 YDL131W HIM1 YDR317W Protein of unknown function involved in DNA repair 1.2462 0.0001 MPC54 YOR177C Component of the meiotic outer plaque, a membrane‐organizing center which is assembled on the cytoplasmic face of the spindle pole body during meiosis II and triggers the formation of the prospore membrane; potential Cdc28p substrate 1.2229 0.0002 MUP3 YHL036W Low affinity methionine permease, similar to Mup1p 1.2217 0.0010 OSW2 YLR054C 1.2215 0.0005 HXT14 YNL318C Protein of unknown function proposed to be involved in the assembly of the spore wall Protein with similarity to hexose transporter family members, expression is induced in low glucose and repressed in high glucose; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐throughput studies 1.2165 0.0006 YSW1 YBR148W Protein required for normal prospore membrane formation; interacts with Gip1p, which is the meiosis‐specific regulatory subunit of the Glc7p protein phosphatase; expressed specifically in spores and localizes to the prospore membrane 1.2147 0.0001 ECM11 YDR446W 1.2101 0.0003 THI13 YDL244W 1.2080 0.0002 NA YDR379C‐A 1.2051 0.0007 SSP2 YOR242C 1.2000 0.0005 NA YLL066W‐B 1.1877 0.0008 UGA3 YDL170W Non‐essential protein apparently involved in meiosis, GFP fusion protein is present in discrete clusters in the nucleus throughout mitosis; may be involved in maintaining chromatin structure Protein involved in synthesis of the thiamine precursor hydroxymethylpyrimidine (HMP); member of a subtelomeric gene family including THI5, THI11, THI12, and THI13 Protein involved in the assembly of the mitochondrial succinate dehydrogenase complex; mutations in human ortholog SDHAF1 are associated with infantile leukoencephalopathy Sporulation specific protein that localizes to the spore wall; required for sporulation at a point after meiosis II and during spore wall formation; SSP2 expression is induced midway in meiosis Putative protein of unknown function; overexpression causes a cell cycle delay or arrest Transcriptional activator necessary for gamma‐aminobutyrate (GABA)‐dependent induction of GABA genes (such as UGA1, UGA2, UGA4); zinc‐finger transcription factor of the Zn(2)‐Cys(6) binuclear cluster domain type; localized to the nucleus 1.1791 0.0003 Putative glycoside hydrolase of the spore wall envelope; required for normal spore wall assembly, possibly for cross‐linking between the glucan and chitosan layers; expressed during sporulation Member of the seripauperin multigene family encoded mainly in subtelomeric regions; active during alcoholic fermentation, regulated by anaerobiosis, negatively regulated by oxygen, repressed by heme 161 log2(mtr10 .Tf/WT.Tf) adj.p.va l Putative protein of unknown function, involved in copper metabolism; similar to C. carbonum toxD gene; member of the quinone oxidoreductase family 1.1627 0.0002 YOL058W Arginosuccinate synthetase, catalyzes the formation of L‐ argininosuccinate from citrulline and L‐aspartate in the arginine biosynthesis pathway; potential Cdc28p substrate 1.1627 0.0007 HNT2 YDR305C Dinucleoside triphosphate hydrolase; has similarity to the tumor suppressor FHIT and belongs to the histidine triad (HIT) superfamily of nucleotide‐binding proteins 1.1561 0.0001 NA YDR018C 1.1517 0.0003 DAL1 YIR027C Probable membrane protein with three predicted transmembrane domains; homologous to Ybr042cp, similar to C. elegans F55A11.5 and maize 1‐acyl‐glycerol‐3‐phosphate acyltransferase Allantoinase, converts allantoin to allantoate in the first step of allantoin degradation; expression sensitive to nitrogen catabolite repression 1.1459 0.0004 NA YLR466C‐B 1.1438 0.0001 DSF1 YEL070W Dubious open reading frame unlikely to encode a protein, based on available experimental and comparative sequence data Deletion suppressor of mpt5 mutation 1.1398 0.0004 NA YLR460C 1.1386 0.0002 MTH1 YDR277C Member of the quinone oxidoreductase family, up‐regulated in response to the fungicide mancozeb; possibly up‐regulated by iodine Negative regulator of the glucose‐sensing signal transduction pathway, required for repression of transcription by Rgt1p; interacts with Rgt1p and the Snf3p and Rgt2p glucose sensors; phosphorylated by Yck1p, triggering Mth1p degradation 1.1369 0.0006 CTS2 YDR371W 1.1306 0.0004 REC8 YPR007C Putative chitinase, functionally complements A. gossypii cts2 mutant sporulation defect Meiosis‐specific component of sister chromatid cohesion complex; maintains cohesion between sister chromatids during meiosis I; maintains cohesion between centromeres of sister chromatids until meiosis II; homolog of S. pombe Rec8p 1.1304 0.0007 CTA1 YDR256C Catalase A, breaks down hydrogen peroxide in the peroxisomal matrix formed by acyl‐CoA oxidase (Pox1p) during fatty acid beta‐ oxidation 1.1300 0.0005 SNA4 YDL123W 1.1236 0.0002 SHS1 YDL225W 1.1057 0.0008 NDT80 YHR124W Protein of unknown function, localized to the vacuolar outer membrane; predicted to be palmitoylated One of five related septins (Cdc3p, Cdc10p, Cdc11p, Cdc12p, Shs1p) that form a cortical filamentous collar at the mother‐bud neck which is necessary for normal morphogenesis and cytokinesis Meiosis‐specific transcription factor required for exit from pachytene and for full meiotic recombination; activates middle sporulation genes; competes with Sum1p for binding to promoters containing middle sporulation elements (MSE) 1.1043 0.0003 NA YDL233W Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm and nucleus; YDL233W is not an essential gene 1.1031 0.0001 NA YDR186C 1.0952 0.0001 GYP7 YDL234C Putative protein of unknown function; may interact with ribosomes, based on co‐purification experiments; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm GTPase‐activating protein for yeast Rab family members including: Ypt7p (most effective), Ypt1p, Ypt31p, and Ypt32p (in vitro); involved in vesicle mediated protein trafficking 1.0918 0.0002 NA YLL066W‐B 1.0896 0.0007 Symbols ORF Description NA YCR102C ARG1 Putative protein of unknown function; overexpression causes a cell cycle delay or arrest 162 log2(mtr10 .Tf/WT.Tf) adj.p.va l Cytoplasmic light chain dynein, microtubule motor protein; proposed to be involved in the assembly of the nuclear pore complex 1.0875 0.0002 Mitochondrial protein required for degradation of unstable forms of cytochrome c Putative protein kinase, potentially phosphorylated by Cdc28p; interacts with ribosome biogenesis factors, Cka2, Gus1 and Arc1 Component of the CCR4‐NOT complex, which has multiple roles in regulating mRNA levels including regulation of transcription and destabilizing mRNAs by deadenylation; basal transcription factor 1.0856 0.0002 1.0824 0.0002 1.0794 0.0002 Protein involved in RNA splicing by the spliceosome; component of a complex containing Cef1p; interacts genetically with ISY1 and BUD13; may bind RNA; has similarity to S. pombe Cwf21p Protein of unknown function; associates with meiosis‐specific protein Spo1p Putative aryl‐alcohol dehydrogenase with similarity to P. chrysosporium aryl‐alcohol dehydrogenase, involved in the oxidative stress response; expression induced in cells treated with the mycotoxin patulin 1.0788 0.0001 1.0787 0.0003 1.0740 0.0002 Member of the seripauperin multigene family encoded mainly in subtelomeric regions, active during alcoholic fermentation, regulated by anaerobiosis, negatively regulated by oxygen, repressed by heme DNA damage‐dependent checkpoint protein, required for cell‐cycle arrest in G1/S, intra‐S, and G2/M; transmits checkpoint signal by activating Rad53p and Chk1p; hyperphosphorylated by Mec1p and Tel1p; potential Cdc28p substrate 1.0732 0.0004 1.0708 0.0004 Putative aryl‐alcohol dehydrogenase with similarity to P. chrysosporium aryl‐alcohol dehydrogenase, involved in the oxidative stress response; expression induced in cells treated with the mycotoxin patulin 1.0706 0.0002 Putative protein of unknown function; proposed to be a palmitoylated membrane protein Protein with similarity to mammalian monocarboxylate permeases, which are involved in transport of monocarboxylic acids across the plasma membrane; mutant is not deficient in monocarboxylate transport 1.0621 0.0010 1.0609 0.0003 YDR308C Subunit of the RNA polymerase II mediator complex; associates with core polymerase subunits to form the RNA polymerase II holoenzyme; essential for transcriptional regulation; target of the global repressor Tup1p 1.0608 0.0001 YOS9 YDR057W ER quality‐control lectin; integral subunit of the HRD ligase; binds to glycans with terminal alpha‐1,6 linked mannose on misfolded N‐ glycosylated proteins and participates in targeting proteins to ERAD; member of the OS‐9 protein family 1.0606 0.0005 PAU18 YLL064C 1.0587 0.0002 NA YDR366C Protein of unknown function, member of the seripauperin multigene family encoded mainly in subtelomeric regions; identical to Pau6p Putative protein of unknown function 1.0567 0.0001 LCD1 YDR499W Essential protein required for the DNA integrity checkpoint pathways; interacts physically with Mec1p; putative homolog of S. pombe Rad26 and human ATRIP 1.0543 0.0009 Symbols ORF Description DYN2 YDR424C SUE1 YPR151C RTK1 YDL025C CDC36 YDL165W CWC21 YDR482C SPO23 YBR250W AAD4 YDL243C PAU3 YCR104W RAD9 YDR217C AAD4 YDL243C NA YKL107W MCH1 YDL054C SRB7 163 log2(mtr10 .Tf/WT.Tf) adj.p.va l Chaperone that specifically facilitates the assembly of cytochrome c oxidase, integral to the mitochondrial inner membrane; interacts with a subcomplex of subunits VII, VIIa, and VIII (Cox7p, Cox9p, and Cox8p) but not with the holoenzyme 1.0495 0.0001 YHL016C Plasma membrane transporter for both urea and polyamines, expression is highly sensitive to nitrogen catabolite repression and induced by allophanate, the last intermediate of the allantoin degradative pathway 1.0471 0.0001 COA2 YPL189C‐A Cytochrome oxidase assembly factor; null mutation results in respiratory deficiency with specific loss of cytochrome oxidase activity; functions downstream of assembly factors Mss51p and Coa1p and interacts with assembly factor Shy1p 1.0452 0.0006 MTD1 YKR080W NAD‐dependent 5,10‐methylenetetrahydrafolate dehydrogenase, plays a catalytic role in oxidation of cytoplasmic one‐carbon units; expression is regulated by Bas1p and Bas2p, repressed by adenine, and may be induced by inositol and choline 1.0384 0.0001 NA YLR466C‐B 1.0374 0.0002 DFM1 YDR411C Dubious open reading frame unlikely to encode a protein, based on available experimental and comparative sequence data Endoplasmic reticulum (ER) localized protein involved in ER‐ associated protein degradation (ERAD), ER stress and homeostasis; interacts with components of ERAD‐L and ERAD‐C and Cdc48p; derlin‐like family member similar to Der1p 1.0350 0.0002 NA YDL073W 1.0313 0.0002 NA YDR248C 1.0260 0.0002 BUR6 YER159C Subunit of a heterodimeric NC2 transcription regulator complex with Ncb2p; complex binds to TBP and can repress transcription by preventing preinitiation complex assembly or stimulate activated transcription; homologous to human NC2alpha 1.0236 0.0003 RTC2 YBR147W Protein of unknown function; identified in a screen for mutants with decreased levels of rDNA transcription; detected in highly purified mitochondria; null mutant suppresses cdc13‐1; similar to a G‐protein coupled receptor from S. pombe 1.0235 0.0003 SLM3 YDL033C 1.0226 0.0003 PAU8 YAL068C 1.0213 0.0002 NA YBR085C‐A 1.0140 0.0003 LYS1 YIR034C 1.0115 0.0009 LRS4 YDR439W tRNA‐specific 2‐thiouridylase, responsible for 2‐thiolation of the wobble base of mitochondrial tRNAs; human ortholog is implicated in myoclonus epilepsy associated with ragged red fibers (MERRF) Protein of unknown function, member of the seripauperin multigene family encoded mainly in subtelomeric regions Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm and to the nucleus Saccharopine dehydrogenase (NAD+, L‐lysine‐forming), catalyzes the conversion of saccharopine to L‐lysine, which is the final step in the lysine biosynthesis pathway; also has mRNA binding activity Nucleolar protein that forms a complex with Csm1p, and then Mam1p at kinetochores during meiosis I to mediate accurate homolog segregation; required for condensin recruitment to the replication fork barrier site and rDNA repeat segregation 1.0093 0.0003 POL3 YDL102W 1.0085 0.0001 Symbols ORF Description PET100 YDR079W DUR3 Putative protein of unknown function; YDL073W is not an essential gene Putative protein of unknown function; sequence similarity to bacterial and human gluconokinase; green fluorescent protein (GFP)‐ fusion protein localizes to the cytoplasm; upregulated by deletion of the RNAP‐II associated factor, PAF1 Catalytic subunit of DNA polymerase delta; required for chromosomal DNA replication during mitosis and meiosis, intragenic recombination, repair of double strand DNA breaks, and DNA replication during nucleotide excision repair (NER) 164 log2(mtr10 .Tf/WT.Tf) adj.p.va l Highly conserved mitochondrial protein, essential for t6A modification of mitochondrial tRNAs that decode ANN codons; similar to Kae1p and E. coli YgjD, both of which are also required for tRNA t6A modification 1.0077 0.0006 Putative protein of unknown function; likely member of BLOC complex involved in endosomal cargo sorting; green fluorescent protein (GFP)‐fusion protein localizes to endosomes Ethanolamine kinase, primarily responsible for phosphatidylethanolamine synthesis via the CDP‐ethanolamine pathway; exhibits some choline kinase activity, thus contributing to phosphatidylcholine synthesis via the CDP‐choline pathway 1.0006 0.0002 0.9881 0.0001 Non‐essential kinetochore protein, subunit of the Ctf19 central kinetochore complex (Ctf19p‐Mcm21p‐Okp1p‐Mcm22p‐Mcm16p‐ Ctf3p‐Chl4p‐Mcm19p‐Nkp1p‐Nkp2p‐Ame1p‐Mtw1p) Putative protein of unknown function 0.9880 0.0001 0.9870 0.0002 YDR263C Mitochondrial nuclease functioning in DNA repair and replication, modulates the stability of the mitochondrial genome, induced by exposure to mutagens, also induced during meiosis at a time nearly coincident with commitment to recombination 0.9836 0.0003 GCV1 YDR019C 0.9817 0.0003 SHU2 YDR078C T subunit of the mitochondrial glycine decarboxylase complex, required for the catabolism of glycine to 5,10‐methylene‐THF; expression is regulated by levels of levels of 5,10‐methylene‐THF in the cytoplasm Protein involved in a Rad51p‐, Rad54p‐dependent pathway for homologous recombination repair, important for error‐free repair of spontaneous and induced DNA lesions to protect the genome from mutation; associates with Shu1p, Psy3p, and Csm2p 0.9733 0.0002 ARO80 YDR421W Zinc finger transcriptional activator of the Zn2Cys6 family; activates transcription of aromatic amino acid catabolic genes in the presence of aromatic amino acids 0.9711 0.0001 RNH202 YDR279W 0.9698 0.0001 UBX3 YDL091C 0.9661 0.0004 CBS1 YDL069C Ribonuclease H2 subunit, required for RNase H2 activity; related to human AGS2 that causes Aicardi‐Goutieres syndrome UBX (ubiquitin regulatory X) domain‐containing protein that interacts with Cdc48p, green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm in a punctate pattern Mitochondrial translational activator of the COB mRNA; membrane protein that interacts with translating ribosomes, acts on the COB mRNA 5'‐untranslated leader 0.9651 0.0007 MBR1 YKL093W Protein involved in mitochondrial functions and stress response; overexpression suppresses growth defects of hap2, hap3, and hap4 mutants 0.9647 0.0003 SCM3 YDL139C Nonhistone component of centromeric chromatin that binds stoichiometrically to CenH3‐H4 histones, required for kinetochore assembly; required for G2/M progression and localization of Cse4p; may protect Cse4p from ubiquitylation 0.9641 0.0001 PFA5 YDR459C Palmitoyltransferase with autoacylation activity; likely functions in pathway(s) outside Ras; member of a family of putative palmitoyltransferases containing an Asp‐His‐His‐Cys‐cysteine rich (DHHC‐CRD) domain 0.9595 0.0003 QCR7 YDR529C Subunit 7 of the ubiquinol cytochrome‐c reductase complex, which is a component of the mitochondrial inner membrane electron transport chain; oriented facing the mitochondrial matrix; N‐ terminus appears to play a role in complex assembly 0.9559 0.0001 POX1 YGL205W Fatty‐acyl coenzyme A oxidase, involved in the fatty acid beta‐ oxidation pathway; localized to the peroxisomal matrix 0.9550 0.0006 Symbols ORF Description QRI7 YDL104C SNN1 YNL086W EKI1 YDR147W NKP1 YDR383C NA YDR132C DIN7 165 log2(mtr10 .Tf/WT.Tf) adj.p.va l Putative GPI‐anchored aspartic protease, member of the yapsin family of proteases involved in cell wall growth and maintenance; located in the cytoplasm and endoplasmic reticulum Putative kinase 0.9548 0.0001 0.9515 0.0001 Putative protein of unknown function; identified by fungal homology and RT‐PCR Membrane anchor subunit of succinate dehydrogenase (Sdh1p, Sdh2p, Sdh3p, Sdh4p), which couples the oxidation of succinate to the transfer of electrons to ubiquinone as part of the TCA cycle and the mitochondrial respiratory chain 0.9512 0.0009 0.9420 0.0004 Anti‐silencing protein that causes derepression of silent loci when overexpressed Protein that interacts with CCT (chaperonin containing TCP‐1) complex and has a role in actin and tubulin folding; has weak similarity to phosducins, which are G‐protein regulators 0.9388 0.0002 0.9385 0.0002 Symbols ORF Description YPS7 YDR349C NA YDR109C NA YER078W‐A SDH4 YDR178W ASF2 YDL197C PLP1 YDR183W PEX3 YDR329C Peroxisomal membrane protein (PMP) required for proper localization and stability of PMPs; anchors peroxisome retention factor Inp1p at the peroxisomal membrane; interacts with Pex19p 0.9377 0.0001 NA YDR239C 0.9375 0.0002 SEC1 YDR164C 0.9349 0.0001 REF2 YDR195W Protein of unknown function that may interact with ribosomes, based on co‐purification experiments Sm‐like protein involved in docking and fusion of exocytic vesicles through binding to assembled SNARE complexes at the membrane; localization to sites of secretion (bud neck and bud tip) is dependent on SNARE function RNA‐binding protein involved in the cleavage step of mRNA 3'‐end formation prior to polyadenylation, and in snoRNA maturation; part of holo‐CPF subcomplex APT, which associates with 3'‐ends of snoRNA‐ and mRNA‐encoding genes 0.9342 0.0002 VPS60 YDR486C 0.9334 0.0002 NA YGR146C‐A Putative protein of unknown function 0.9322 0.0004 SPC19 YDR201W Essential subunit of the Dam1 complex (aka DASH complex), couples kinetochores to the force produced by MT depolymerization thereby aiding in chromosome segregation; also localized to nuclear side of spindle pole body 0.9291 0.0003 NA YCR108C 0.9270 0.0002 PRM9 YAR031W Putative protein of unknown function; identified by fungal homology and RT‐PCR Pheromone‐regulated protein with 3 predicted transmembrane segments and an FF sequence, a motif involved in COPII binding; member of DUP240 gene family 0.9258 0.0002 AAD14 YNL331C Putative aryl‐alcohol dehydrogenase with similarity to P. chrysosporium aryl‐alcohol dehydrogenase; mutational analysis has not yet revealed a physiological role 0.9206 0.0002 CRG1 YHR209W 0.9196 0.0003 QRI1 YDL103C mRNA binding protein and putative S‐adenosylmethionine‐ dependent methyltransferase; mediates cantharidin resistance UDP‐N‐acetylglucosamine pyrophosphorylase, catalyzes the formation of UDP‐N‐acetylglucosamine (UDP‐GlcNAc), which is important in cell wall biosynthesis, protein N‐glycosylation, and GPI anchor biosynthesis 0.9185 0.0002 Cytoplasmic and vacuolar membrane protein involved in late endosome to vacuole transport; required for normal filament maturation during pseudohyphal growth; may function in targeting cargo proteins for degradation; interacts with Vta1p 166 log2(mtr10 .Tf/WT.Tf) adj.p.va l Subunit of TFIIH and nucleotide excision repair factor 3 complexes, involved in transcription initiation, required for nucleotide excision repair; ring finger protein similar to mammalian CAK and TFIIH subunit 0.9177 0.0002 YDR058C Triacylglycerol lipase that is localized to the mitochondria; has lipolytic activity towards triacylglycerols and diacylglycerols when expressed in E. coli 0.9150 0.0004 TAF10 YDR167W 0.9088 0.0009 ADE17 YMR120C Subunit (145 kDa) of TFIID and SAGA complexes, involved in RNA polymerase II transcription initiation and in chromatin modification Enzyme of 'de novo' purine biosynthesis containing both 5‐ aminoimidazole‐4‐carboxamide ribonucleotide transformylase and inosine monophosphate cyclohydrolase activities, isozyme of Ade16p; ade16 ade17 mutants require adenine and histidine 0.9074 0.0002 MCT1 YOR221C Predicted malonyl‐CoA:ACP transferase, putative component of a type‐II mitochondrial fatty acid synthase that produces intermediates for phospholipid remodeling 0.9028 0.0002 NA YDR514C Putative protein of unknown function 0.8995 0.0009 NA YNL195C 0.8991 0.0004 NA YDR056C Putative protein of unknown function; shares a promoter with YNL194C; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐throughput studies Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the endoplasmic reticulum; YDR056C is not an essential protein 0.8946 0.0002 NA YAL064W 0.8929 0.0003 NA YPL071C Protein of unknown function; may interact with ribosomes, based on co‐purification experiments Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to both the cytoplasm and the nucleus 0.8918 0.0004 PTP1 YDL230W Phosphotyrosine‐specific protein phosphatase that dephosphorylates a broad range of substrates in vivo, including Fpr3p; localized to the cytoplasm and the mitochondria 0.8882 0.0002 APA2 YDR530C Diadenosine 5',5''‐P1,P4‐tetraphosphate phosphorylase II (AP4A phosphorylase), involved in catabolism of bis(5'‐nucleosidyl) tetraphosphates; has similarity to Apa1p 0.8841 0.0002 PEP7 YDR323C Multivalent adaptor protein that facilitates vesicle‐mediated vacuolar protein sorting by ensuring high‐fidelity vesicle docking and fusion, which are essential for targeting of vesicles to the endosome; required for vacuole inheritance 0.8804 0.0007 ADE4 YMR300C Phosphoribosylpyrophosphate amidotransferase (PRPPAT; amidophosphoribosyltransferase), catalyzes first step of the 'de novo' purine nucleotide biosynthetic pathway 0.8738 0.0007 RAM1 YDL090C Beta subunit of the CAAX farnesyltransferase (FTase) that prenylates the a‐factor mating pheromone and Ras proteins; required for the membrane localization of Ras proteins and a‐factor; homolog of the mammalian FTase beta subunit 0.8734 0.0002 DOS2 YDR068W 0.8733 0.0002 KIN1 YDR122W Protein of unknown function, green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm Serine/threonine protein kinase involved in regulation of exocytosis; localizes to the cytoplasmic face of the plasma membrane; closely related to Kin2p 0.8690 0.0006 VMS1 YDR049W 0.8680 0.0002 Symbols ORF Description TFB3 YDR460W TGL2 Zinc finger protein, forms a mitochondrially‐associated complex with Cdc48p and Npl4p under oxidative stress that is required for ubiquitin‐mediated mitochondria‐associated protein degradation (MAD); conserved in C. elegans and human 167 log2(mtr10 .Tf/WT.Tf) adj.p.va l Putative protein of unknown function; identified by fungal homology and RT‐PCR Protein of unknown function that may interact with ribosomes, based on co‐purification experiments; Myb‐like DNA‐binding protein that may bind to the Ter region of rDNA; interacts physically with Fob1p 0.8669 0.0009 0.8669 0.0003 YDL194W Plasma membrane low glucose sensor that regulates glucose transport; contains 12 predicted transmembrane segments and a long C‐terminal tail required for induction of hexose transporters; also senses fructose and mannose; similar to Rgt2p 0.8669 0.0003 NRG1 YDR043C Transcriptional repressor that recruits the Cyc8p‐Tup1p complex to promoters; mediates glucose repression and negatively regulates a variety of processes including filamentous growth and alkaline pH response 0.8664 0.0009 QCR2 YPR191W Subunit 2 of the ubiquinol cytochrome‐c reductase complex, which is a component of the mitochondrial inner membrane electron transport chain; phosphorylated; transcription is regulated by Hap1p, Hap2p/Hap3p, and heme 0.8652 0.0003 NBP2 YDR162C Protein involved in the HOG (high osmolarity glycerol) pathway, negatively regulates Hog1p by recruitment of phosphatase Ptc1p the Pbs2p‐Hog1p complex, found in the nucleus and cytoplasm, contains an SH3 domain that binds Pbs2p 0.8637 0.0003 DAL4 YIR028W Allantoin permease; expression sensitive to nitrogen catabolite repression and induced by allophanate, an intermediate in allantoin degradation 0.8630 0.0007 NA YDR179W‐A Putative protein of unknown function 0.8618 0.0004 DAD1 YDR016C Essential subunit of the Dam1 complex (aka DASH complex), couples kinetochores to the force produced by MT depolymerization thereby aiding in chromosome segregation; is transferred to the kinetochore prior to mitosis 0.8603 0.0006 NA YDR262W Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the vacuole and is induced in response to the DNA‐damaging agent MMS; gene expression increases in response to Zymoliase treatment 0.8577 0.0003 ATG10 YLL042C 0.8574 0.0008 YPD1 YDL235C Conserved E2‐like conjugating enzyme that mediates formation of the Atg12p‐Atg5p conjugate, which is a critical step in autophagy Phosphorelay intermediate protein, phosphorylated by the plasma membrane sensor Sln1p in response to osmotic stress and then in turn phosphorylates the response regulators Ssk1p in the cytosol and Skn7p in the nucleus 0.8542 0.0004 NA YJL144W Cytoplasmic hydrophilin of unknown function, possibly involved in the dessication response; expression induced by osmotic stress, starvation and during stationary phase; GFP‐fusion protein is induced by the DNA‐damaging agent MMS 0.8533 0.0004 DPB4 YDR121W 0.8505 0.0009 NA YHL048C‐A 0.8493 0.0006 NA YDR131C 0.8471 0.0002 RAV2 YDR202C Shared subunit of DNA polymerase (II) epsilon and of ISW2/yCHRAC chromatin accessibility complex; involved in both chromosomal DNA replication and in inheritance of telomeric silencing Putative protein of unknown function; identified by expression profiling and mass spectrometry F‐box protein, substrate‐specific adaptor subunit that recruits substrates to a core ubiquitination complex Subunit of RAVE (Rav1p, Rav2p, Skp1p), a complex that associates with the V1 domain of the vacuolar membrane (H+)‐ATPase (V‐ ATPase) and promotes assembly and reassembly of the holoenzyme 0.8461 0.0003 Symbols ORF Description NA YDR246W‐A NA YDR026C SNF3 168 log2(mtr10 .Tf/WT.Tf) adj.p.va l ATPase involved in protein folding and the response to stress; plays a role in SRP‐dependent cotranslational protein‐membrane targeting and translocation; member of the heat shock protein 70 (HSP70) family; localized to the cytoplasm 0.8460 0.0005 YJR022W Lsm (Like Sm) protein; forms heteroheptameric complex (with Lsm2p, Lsm3p, Lsm4p, Lsm5p, Lsm6p, and Lsm7p) that is part of spliceosomal U6 snRNP and is also implicated in processing of pre‐ tRNA, pre‐snoRNA, and pre‐rRNA 0.8442 0.0009 CKS1 YBR135W Cyclin‐dependent protein kinase regulatory subunit and adaptor; modulates proteolysis of M‐phase targets through interactions with the proteasome; role in transcriptional regulation, recruiting proteasomal subunits to target gene promoters 0.8431 0.0003 NA YIL082W‐A Retrotransposon TYA Gag and TYB Pol genes; transcribed/translated as one unit; polyprotein is processed to make a nucleocapsid‐like protein (Gag), reverse transcriptase (RT), protease (PR), and integrase (IN); similar to retroviral genes 0.8429 0.0004 PEX19 YDL065C Chaperone and import receptor for newly‐synthesized class I peroxisomal membrane proteins (PMPs), binds PMPs in the cytoplasm and delivers them to the peroxisome for subsequent insertion into the peroxisomal membrane 0.8410 0.0008 DAL3 YIR032C Ureidoglycolate hydrolase, converts ureidoglycolate to glyoxylate and urea in the third step of allantoin degradation; expression sensitive to nitrogen catabolite repression 0.8359 0.0005 NA YDL086W 0.8359 0.0003 NA YOL155W‐A 0.8341 0.0003 OCA6 YDR067C Putative protein of unknown function; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐ throughput studies; YDL086W is not an essential gene Putative protein of unknown function; identified by expression profiling and mass spectrometry Cytoplasmic protein required for replication of Brome mosaic virus in S. cerevisiae, which is a model system for studying positive‐strand RNA virus replication; null mutation confers sensitivity to tunicamycin and DTT 0.8333 0.0002 SNU23 YDL098C 0.8318 0.0004 SDC1 YDR469W Component of U4/U6.U5 snRNP involved in mRNA splicing via spliceosome Subunit of the COMPASS (Set1C) complex, which methylates lysine 4 of histone H3 and is required in chromatin silencing at telomeres; contains a Dpy‐30 domain that mediates interaction with Bre2p; similar to C. elegans and human DPY‐30 0.8307 0.0008 CDC7 YDL017W DDK (Dbf4‐dependent kinase) catalytic subunit required for firing origins and replication fork progression in S phase through phosphorylation of Mcm2‐7p complexes and Cdc45p; kinase activity correlates with cyclical DBF4 expression 0.8304 0.0003 PDS1 YDR113C Securin, inhibits anaphase by binding separin Esp1p; blocks cyclin destruction and mitotic exit, essential for meiotic progression and mitotic cell cycle arrest; localization is cell‐cycle dependent and regulated by Cdc28p phosphorylation 0.8265 0.0004 IPK1 YDR315C Inositol 1,3,4,5,6‐pentakisphosphate 2‐kinase, nuclear protein required for synthesis of 1,2,3,4,5,6‐hexakisphosphate (phytate), which is integral to cell function; has 2 motifs conserved in other fungi; ipk1 gle1 double mutant is inviable 0.8220 0.0009 REG1 YDR028C Regulatory subunit of type 1 protein phosphatase Glc7p, involved in negative regulation of glucose‐repressible genes 0.8218 0.0004 Symbols ORF Description SSA3 YBL075C LSM8 169 log2(mtr10 .Tf/WT.Tf) adj.p.va l Ubiquitin‐conjugating enzyme that mediates selective degradation of short‐lived and abnormal proteins; plays a role in vesicle biogenesis and ER‐associated protein degradation (ERAD); component of the cellular stress response 0.8193 0.0005 YDR125C Protein of unknown function, similar to Rlp24p 0.8192 0.0002 LYS9 YNR050C Saccharopine dehydrogenase (NADP+, L‐glutamate‐forming); catalyzes the formation of saccharopine from alpha‐aminoadipate 6‐ semialdehyde, the seventh step in lysine biosynthesis pathway; exhibits genetic and physical interactions with TRM112 0.8153 0.0007 EAF7 YNL136W 0.8142 0.0005 DIG2 YDR480W Subunit of the NuA4 histone acetyltransferase complex, which acetylates the N‐terminal tails of histones H4 and H2A MAP kinase‐responsive inhibitor of the Ste12p transcription factor, involved in the regulation of mating‐specific genes and the invasive growth pathway; related regulators Dig1p and Dig2p bind to Ste12p 0.8129 0.0004 ACN9 YDR511W Protein of the mitochondrial intermembrane space, required for acetate utilization and gluconeogenesis; has orthologs in higher eukaryotes 0.8123 0.0003 HIS4 YCL030C Multifunctional enzyme containing phosphoribosyl‐ATP pyrophosphatase, phosphoribosyl‐AMP cyclohydrolase, and histidinol dehydrogenase activities; catalyzes the second, third, ninth and tenth steps in histidine biosynthesis 0.8070 0.0003 RBS1 YDL189W 0.8008 0.0003 SSF2 YDR312W 0.8005 0.0006 DLD2 YDL178W Protein of unknown function, identified as a high copy suppressor of psk1 psk2 mutations that confer temperature‐sensitivity for galactose utilization; proposed to bind single‐stranded nucleic acids via its R3H domain Protein required for ribosomal large subunit maturation, functionally redundant with Ssf1p; member of the Brix family D‐lactate dehydrogenase, located in the mitochondrial matrix 0.7929 0.0008 SMD3 YLR147C Core Sm protein Sm D3; part of heteroheptameric complex (with Smb1p, Smd1p, Smd2p, Sme1p, Smx3p, and Smx2p) that is part of the spliceosomal U1, U2, U4, and U5 snRNPs; homolog of human Sm D3 0.7861 0.0008 SNM1 YDR478W Subunit of RNase MRP, which cleaves pre‐rRNA and has a role in cell cycle‐regulated degradation of daughter cell‐specific mRNAs; binds to the NME1 RNA subunit of RNase MRP 0.7850 0.0004 PRM5 YIL117C 0.7824 0.0005 UFD2 YDL190C Pheromone‐regulated protein, predicted to have 1 transmembrane segment; induced during cell integrity signaling Ubiquitin chain assembly factor (E4) that cooperates with a ubiquitin‐activating enzyme (E1), a ubiquitin‐conjugating enzyme (E2), and a ubiquitin protein ligase (E3) to conjugate ubiquitin to substrates; also functions as an E3 0.7821 0.0007 SSP1 YHR184W Protein involved in the control of meiotic nuclear division and coordination of meiosis with spore formation; transcription is induced midway through meiosis 0.7797 0.0004 MRP10 YDL045W‐A 0.7797 0.0004 PTC1 YDL006W Mitochondrial ribosomal protein of the small subunit; contains twin cysteine‐x9‐cysteine motifs Type 2C protein phosphatase (PP2C); dephosphorylates Hog1p, inactivating osmosensing MAPK cascade; involved in Fus3p activation during pheromone response; deletion affects precursor tRNA splicing, mitochondrial inheritance, and sporulation 0.7767 0.0003 Symbols ORF Description UBC1 YDR177W ECM18 170 log2(mtr10 .Tf/WT.Tf) adj.p.va l GTPase‐activating protein for the polarity‐establishment protein Cdc42p; implicated in control of septin organization, pheromone response, and haploid invasive growth; regulated by Pho85p and Cdc28p 0.7750 0.0009 YDR027C Component of the GARP (Golgi‐associated retrograde protein) complex, Vps51p‐Vps52p‐Vps53p‐Vps54p, which is required for the recycling of proteins from endosomes to the late Golgi; potentially phosphorylated by Cdc28p 0.7746 0.0003 GCD6 YDR211W Catalytic epsilon subunit of the translation initiation factor eIF2B, the guanine‐nucleotide exchange factor for eIF2; activity subsequently regulated by phosphorylated eIF2; first identified as a negative regulator of GCN4 expression 0.7711 0.0006 MCM21 YDR318W Protein involved in minichromosome maintenance; component of the COMA complex (Ctf19p, Okp1p, Mcm21p, Ame1p) that bridges kinetochore subunits that are in contact with centromeric DNA and the subunits bound to microtubules 0.7708 0.0009 KIN28 YDL108W Serine/threonine protein kinase, subunit of the transcription factor TFIIH; involved in transcription initiation at RNA polymerase II promoters 0.7704 0.0002 JNM1 YMR294W Component of the yeast dynactin complex, consisting of Nip100p, Jnm1p, and Arp1p; required for proper nuclear migration and spindle partitioning during mitotic anaphase B 0.7699 0.0008 COX20 YDR231C 0.7690 0.0004 HEM13 YDR044W Mitochondrial inner membrane protein, required for proteolytic processing of Cox2p and its assembly into cytochrome c oxidase Coproporphyrinogen III oxidase, an oxygen requiring enzyme that catalyzes the sixth step in the heme biosynthetic pathway; transcription is repressed by oxygen and heme (via Rox1p and Hap1p) 0.7686 0.0006 STP1 YDR463W Transcription factor, undergoes proteolytic processing by SPS (Ssy1p‐ Ptr3p‐Ssy5p)‐sensor component Ssy5p in response to extracellular amino acids; activates transcription of amino acid permease genes and may have a role in tRNA processing 0.7677 0.0004 SWA2 YDR320C 0.7654 0.0009 AIR2 YDL175C Auxilin‐like protein involved in vesicular transport; clathrin‐binding protein required for uncoating of clathrin‐coated vesicles Zinc knuckle protein, involved in nuclear RNA processing and degredation as a component of the TRAMP complex; stimulates the poly(A) polymerase activity of Pap2p in vitro; functionally redundant with Air1p 0.7639 0.0004 DET1 YDR051C Acid phosphatase involved in the non‐vesicular transport of sterols in both directions between the endoplasmic reticulum and plasma membrane; deletion confers sensitivity to nickel 0.7615 0.0004 COX9 YDL067C 0.7525 0.0003 DUN1 YDL101C Subunit VIIa of cytochrome c oxidase, which is the terminal member of the mitochondrial inner membrane electron transport chain Cell‐cycle checkpoint serine‐threonine kinase required for DNA damage‐induced transcription of certain target genes, phosphorylation of Rad55p and Sml1p, and transient G2/M arrest after DNA damage; also regulates postreplicative DNA repair 0.7516 0.0006 NA YDL012C Tail‐anchored plasma membrane protein containing a conserved CYSTM module, possibly involved in response to stress; may contribute to non‐homologous end‐joining (NHEJ) based on ydl012c htz1 double null phenotype 0.7508 0.0003 PAU14 YIL176C Protein of unknown function, member of the seripauperin multigene family encoded mainly in subtelomeric regions; identical to Pau1p 0.7465 0.0005 Symbols ORF Description RGA2 YDR379W VPS54 171 log2(mtr10 .Tf/WT.Tf) adj.p.va l Subunit of a heterodimeric nuclear SUMO activating enzyme (E1) with Aos1p; activates Smt3p (SUMO) before its conjugation to proteins (sumoylation), which may play a role in protein targeting; essential for viability 0.7462 0.0003 YDR059C Ubiquitin‐conjugating enzyme that mediates selective degradation of short‐lived, abnormal, or excess proteins, including histone H3; central component of the cellular stress response; expression is heat inducible 0.7460 0.0008 SPO13 YHR014W Meiosis‐specific protein, involved in maintaining sister chromatid cohesion during meiosis I as well as promoting proper attachment of kinetochores to the spindle during meiosis I and meiosis II 0.7424 0.0009 FRQ1 YDR373W N‐myristoylated calcium‐binding protein that may have a role in intracellular signaling through its regulation of the phosphatidylinositol 4‐kinase Pik1p; member of the recoverin/frequenin branch of the EF‐hand superfamily 0.7416 0.0003 NPL3 YDR432W RNA‐binding protein that promotes elongation, regulates termination, and carries poly(A) mRNA from nucleus to cytoplasm; required for pre‐mRNA splicing; dissociation from mRNAs promoted by Mtr10p; phosphorylated by Sky1p in the cytoplasm 0.7413 0.0003 Symbols ORF Description UBA2 YDR390C UBC5 MRPL28 YDR462W Mitochondrial ribosomal protein of the large subunit 0.7407 0.0004 GIC2 YDR309C Redundant rho‐like GTPase Cdc42p effector; homolog of Gic1p; involved in initiation of budding and cellular polarization; interacts with Cdc42p via the Cdc42/Rac‐interactive binding (CRIB) domain and with PI(4,5)P2 via a polybasic region 0.7396 0.0004 GIR2 YDR152W Highly‐acidic cytoplasmic RWD domain‐containing protein of unknown function; forms a complex with Rbg2p; interacts with Rbg1p and Gcn1p; associates with translating ribosomes; putative intrinsically unstructured protein 0.7382 0.0007 STE7 YDL159W Signal transducing MAP kinase kinase involved in pheromone response, where it phosphorylates Fus3p, and in the pseudohyphal/invasive growth pathway, through phosphorylation of Kss1p; phosphorylated by Ste11p, degraded by ubiquitin pathway 0.7365 0.0004 PDC2 YDR081C Transcription factor required for the synthesis of the glycolytic enzyme pyruvate decarboxylase, required for high level expression of both the THI and the PDC genes 0.7351 0.0003 APC4 YDR118W Subunit of the Anaphase‐Promoting Complex/Cyclosome (APC/C), which is a ubiquitin‐protein ligase required for degradation of anaphase inhibitors, including mitotic cyclins, during the metaphase/anaphase transition 0.7349 0.0004 GCS1 YDL226C ADP‐ribosylation factor GTPase activating protein (ARF GAP), involved in ER‐Golgi transport; shares functional similarity with Glo3p 0.7328 0.0010 NA YDL176W Protein of unknown function, predicted by computational methods to be involved in fructose‐1,6‐bisphosphatase (Fbp1p) degradation; interacts with components of the GID complex; YDL176W is not an essential gene 0.7307 0.0008 CWC2 YDL209C Member of the NineTeen Complex (NTC) that contains Prp19p and stabilizes U6 snRNA in catalytic forms of the spliceosome containing U2, U5, and U6 snRNAs; binds directly to U6 snRNA; similar to S. pombe Cwf2 0.7295 0.0006 OPT2 YPR194C Oligopeptide transporter; member of the OPT family, with potential orthologs in S. pombe and C. albicans; also plays a role in formation of mature vacuoles 0.7236 0.0008 172 log2(mtr10 .Tf/WT.Tf) adj.p.va l Dethiobiotin synthetase, catalyzes the third step in the biotin biosynthesis pathway; BIO4 is in a cluster of 3 genes (BIO3, BIO4, and BIO5) that mediate biotin synthesis; expression appears to be repressed at low iron levels 0.7226 0.0006 YDR251W Essential protein of unknown function; exhibits variable expression during colony morphogenesis; overexpression permits survival without protein phosphatase 2A, inhibits growth, and induces a filamentous phenotype 0.7219 0.0004 TRS23 YDR246W One of 10 subunits of the transport protein particle (TRAPP) complex of the cis‐Golgi which mediates vesicle docking and fusion; involved in endoplasmic reticulum (ER) to Golgi membrane traffic; human homolog is TRAPPC4 0.7207 0.0006 TMN2 YDR107C Protein with a role in cellular adhesion and filamentous growth; similar to Emp70p and Tmn3p; member of the evolutionarily conserved Transmembrane Nine family of proteins with nine membrane‐spanning segments 0.7190 0.0006 IVY1 YDR229W 0.7179 0.0004 NA YDR348C Phospholipid‐binding protein that interacts with both Ypt7p and Vps33p, may partially counteract the action of Vps33p and vice versa, localizes to the rim of the vacuole as cells approach stationary phase Protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cell periphery and bud neck; potential Cdc28p substrate 0.7140 0.0003 STN1 YDR082W Telomere end‐binding and capping protein, plays a key role with Pol12p in linking telomerase action with completion of lagging strand synthesis, and in a regulatory step required for telomere capping 0.7138 0.0005 PCF11 YDR228C mRNA 3' end processing factor, essential component of cleavage and polyadenylation factor IA (CF IA), involved in pre‐mRNA 3' end processing and in transcription termination; binds C‐terminal domain of largest subunit of RNA pol II (Rpo21p) 0.7107 0.0004 HDA2 YDR295C Subunit of a possibly tetrameric trichostatin A‐sensitive class II histone deacetylase complex containing an Hda1p homodimer and an Hda2p‐Hda3p heterodimer; involved in telomere maintenance 0.7102 0.0006 SPG1 YGR236C Protein required for survival at high temperature during stationary phase; not required for growth on nonfermentable carbon sources; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐throughput studies 0.7072 0.0008 NA YGR067C 0.7071 0.0005 PBP4 YDL053C Putative protein of unknown function; contains a zinc finger motif similar to that of Adr1p Pbp1p binding protein, interacts strongly with Pab1p‐binding protein 1 (Pbp1p) in the yeast two‐hybrid system; also interacts with Lsm12p in a copurification assay 0.7068 0.0008 NA YGR226C Dubious open reading frame, unlikely to encode a protein; not conserved in closely related Saccharomyces species; overlaps significantly with a verified ORF, AMA1/YGR225W 0.7066 0.0009 RPC11 YDR045C RNA polymerase III subunit C11; mediates pol III RNA cleavage activity and is important for termination of transcription; homologous to TFIIS 0.7057 0.0005 SIZ1 YDR409W SUMO/Smt3 ligase that promotes the attachment of sumo (Smt3p; small ubiquitin‐related modifier) to proteins; binds Ubc9p and may bind septins; specifically required for sumoylation of septins in vivo; localized to the septin ring 0.7030 0.0003 Symbols ORF Description BIO4 YNR057C PAM1 173 log2(mtr10 .Tf/WT.Tf) adj.p.va l 0.7028 0.0004 0.7024 0.0010 0.7013 0.0004 0.6979 0.0003 Bifunctional enzyme containing both alcohol dehydrogenase and glutathione‐dependent formaldehyde dehydrogenase activities, functions in formaldehyde detoxification and formation of long chain and complex alcohols, regulated by Hog1p‐Sko1p 0.6977 0.0004 YMR263W Subunit of a histone deacetylase complex, along with Rpd3p and Sin3p, that is involved in silencing at telomeres, rDNA, and silent mating‐type loci; involved in telomere maintenance 0.6971 0.0009 GRX3 YDR098C Hydroperoxide and superoxide‐radical responsive glutathione‐ dependent oxidoreductase; monothiol glutaredoxin subfamily member along with Grx4p and Grx5p; protects cells from oxidative damage 0.6967 0.0003 THI7 YLR237W Plasma membrane transporter responsible for the uptake of thiamine, member of the major facilitator superfamily of transporters; mutation of human ortholog causes thiamine‐ responsive megaloblastic anemia 0.6912 0.0006 NA YDR115W 0.6897 0.0009 GDH2 YDL215C Putative mitochondrial ribosomal protein of the large subunit, has similarity to E. coli L34 ribosomal protein; required for respiratory growth, as are most mitochondrial ribosomal proteins NAD(+)‐dependent glutamate dehydrogenase, degrades glutamate to ammonia and alpha‐ketoglutarate; expression sensitive to nitrogen catabolite repression and intracellular ammonia levels 0.6887 0.0004 SPT3 YDR392W Subunit of the SAGA and SAGA‐like transcriptional regulatory complexes, interacts with Spt15p to activate transcription of some RNA polymerase II‐dependent genes, also functions to inhibit transcription at some promoters 0.6878 0.0008 PCK1 YKR097W Phosphoenolpyruvate carboxykinase, key enzyme in gluconeogenesis, catalyzes early reaction in carbohydrate biosynthesis, glucose represses transcription and accelerates mRNA degradation, regulated by Mcm1p and Cat8p, located in the cytosol 0.6870 0.0003 PEX5 YDR244W Peroxisomal membrane signal receptor for the C‐terminal tripeptide signal sequence (PTS1) of peroxisomal matrix proteins, required for peroxisomal matrix protein import; also proposed to have PTS1‐ receptor independent functions 0.6823 0.0008 NTG2 YOL043C 0.6777 0.0004 GLE1 YDL207W DNA N‐glycosylase and apurinic/apyrimidinic (AP) lyase involved in base excision repair, localizes to the nucleus; sumoylated Cytoplasmic nucleoporin required for polyadenylated RNA export but not for protein import; component of Nup82p nuclear pore subcomplex; contains a nuclear export signal 0.6772 0.0004 GPR1 YDL035C Plasma membrane G protein coupled receptor (GPCR) that interacts with the heterotrimeric G protein alpha subunit, Gpa2p, and with Plc1p; sensor that integrates nutritional signals with the modulation of cell fate via PKA and cAMP synthesis 0.6758 0.0005 Symbols ORF Description TMS1 YDR105C YSP2 YDR326C GIP2 YER054C ENT1 YDL161W Vacuolar membrane protein of unknown function that is conserved in mammals; predicted to contain eleven transmembrane helices; interacts with Pdr5p, a protein involved in multidrug resistance Protein involved in programmed cell death; mutant shows resistance to cell death induced by amiodarone or intracellular acidification Putative regulatory subunit of the protein phosphatase Glc7p, involved in glycogen metabolism; contains a conserved motif (GVNK motif) that is also found in Gac1p, Pig1p, and Pig2p Epsin‐like protein involved in endocytosis and actin patch assembly and functionally redundant with Ent2p; binds clathrin via a clathrin‐ binding domain motif at C‐terminus SFA1 YDL168W SAP30 174 log2(mtr10 .Tf/WT.Tf) adj.p.va l Exosome non‐catalytic core component; involved in 3'‐5' RNA processing and degradation in both the nucleus and the cytoplasm; has similarity to E. coli RNase PH and to human hRrp42p (EXOSC7) 0.6737 0.0005 YNL146W Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the endoplasmic reticulum; YNL146W is not an essential gene 0.6725 0.0006 YSY6 YBR162W‐A Protein whose expression suppresses a secretory pathway mutation in E. coli; has similarity to the mammalian RAMP4 protein involved in secretion 0.6723 0.0010 NA YDR391C Putative protein of unknown function, possibly involved in zinc homeostasis; Bdf1p‐dependent transcription induced by salt stress; green fluorescent protein (GFP)‐fusion protein localizes to both the cytoplasm and the nucleus 0.6706 0.0004 SYF1 YDR416W Member of the NineTeen Complex (NTC) that contains Prp19p and stabilizes U6 snRNA in catalytic forms of the spliceosome containing U2, U5, and U6 snRNAs; null mutant has splicing defect and arrests in G2/M; homologs in human and C. elegans 0.6684 0.0007 MSL1 YIR009W U2B component of U2 snRNP, involved in splicing, binds the U2 snRNA stem‐loop IV in vitro but requires association of Lea1p for in vivo binding; does not contain the conserved C‐terminal RNA binding domain found in other family members 0.6682 0.0008 PSF1 YDR013W Subunit of the GINS complex (Sld5p, Psf1p, Psf2p, Psf3p), which is localized to DNA replication origins and implicated in assembly of the DNA replication machinery 0.6675 0.0005 SPO12 YHR152W 0.6662 0.0003 NA YDL157C Nucleolar protein of unknown function, positive regulator of mitotic exit; involved in regulating release of Cdc14p from the nucleolus in early anaphase, may play similar role in meiosis Putative protein of unknown function; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐ throughput studies 0.6653 0.0007 ADE1 YAR015W 0.6634 0.0006 PKH1 YDR490C N‐succinyl‐5‐aminoimidazole‐4‐carboxamide ribotide (SAICAR) synthetase, required for 'de novo' purine nucleotide biosynthesis; red pigment accumulates in mutant cells deprived of adenine Serine/threonine protein kinase involved in sphingolipid‐mediated signaling pathway that controls endocytosis; activates Ypk1p and Ykr2p, components of signaling cascade required for maintenance of cell wall integrity; redundant with Pkh2p 0.6596 0.0009 NA YCR076C 0.6595 0.0010 SLY1 YDR189W Putative protein of unknown function; YCR076C is not an essential gene Hydrophilic protein involved in vesicle trafficking between the ER and Golgi; SM (Sec1/Munc‐18) family protein that binds the tSNARE Sed5p and stimulates its assembly into a trans‐SNARE membrane‐ protein complex 0.6574 0.0007 PHO8 YDR481C Repressible alkaline phosphatase, a glycoprotein localized to the vacuole; regulated by levels of inorganic phosphate and by a system consisting of Pho4p, Pho9p, Pho80p, Pho81p and Pho85p; dephosphorylates phosphotyrosyl peptides 0.6553 0.0005 DAL2 YIR029W Allantoicase, converts allantoate to urea and ureidoglycolate in the second step of allantoin degradation; expression sensitive to nitrogen catabolite repression and induced by allophanate, an intermediate in allantoin degradation 0.6542 0.0008 PRM3 YPL192C Pheromone‐regulated protein required for nuclear envelope fusion during karyogamy; localizes to the outer face of the nuclear membrane; interacts with Kar5p at the spindle pole body 0.6539 0.0005 Symbols ORF Description RRP42 YDL111C NA 175 log2(mtr10 .Tf/WT.Tf) adj.p.va l Mitochondrial outer membrane protein, required for assembly of the translocase of the outer membrane (TOM) complex and thereby for mitochondrial protein import; N terminus is exposed to the cytosol: transmembrane segment is highly conserved 0.6496 0.0004 YDR108W Subunit of TRAPPIII (transport protein particle), a multimeric guanine nucleotide‐exchange factor for Ypt1p, required for membrane expansion during autophagy and the CVT pathway; directs Ypt1p to the PAS; late post‐replication meiotic role 0.6490 0.0007 MSS2 YDL107W Peripherally bound inner membrane protein of the mitochondrial matrix involved in membrane insertion of C‐terminus of Cox2p, interacts genetically and physically with Cox18p 0.6489 0.0009 DIF1 YLR437C Protein that regulates the nuclear localization of ribonucleotide reductase Rnr2p and Rnr4p subunits; phosphorylated by Dun1p in response to DNA damage and degraded; N‐terminal half has similarity to S. pombe Spd1 protein 0.6489 0.0008 NTG1 YAL015C DNA N‐glycosylase and apurinic/apyrimidinic (AP) lyase involved in base excision repair; acts in both nucleus and mitochondrion; creates a double‐strand break at mtDNA origins that stimulates replication in response to oxidative stress 0.6471 0.0005 PRP9 YDL030W Subunit of the SF3a splicing factor complex, required for spliceosome assembly; acts after the formation of the U1 snRNP‐ pre‐mRNA complex 0.6412 0.0005 SME1 YOR159C 0.6371 0.0005 SLX5 YDL013W Core Sm protein Sm E; part of heteroheptameric complex (with Smb1p, Smd1p, Smd2p, Smd3p, Smx3p, and Smx2p) that is part of the spliceosomal U1, U2, U4, and U5 snRNPs; homolog of human Sm E Subunit of the Slx5‐Slx8 SUMO‐targeted ubiquitin ligase (STUbL) complex, stimulated by SUMO‐modified substrates; contains a RING domain and two SIMs (SUMO‐interacting motifs); forms SUMO‐ dependent nuclear foci, including DNA repair centers 0.6345 0.0009 CRP1 YHR146W Protein that binds to cruciform DNA structures 0.6345 0.0004 FMP32 YFL046W Putative protein of unknown function; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐ throughput studies 0.6308 0.0006 SEC20 YDR498C Membrane glycoprotein v‐SNARE involved in retrograde transport from the Golgi to the ER; required for N‐ and O‐glycosylation in the Golgi but not in the ER; interacts with the Dsl1p complex through Tip20p 0.6284 0.0010 NA YDR061W Protein with similarity to ATP‐binding cassette (ABC) transporter family members; lacks predicted membrane‐spanning regions; transcriptionally activated by Yrm1p along with genes involved in multidrug resistance 0.6282 0.0009 CTH1 YDR151C 0.6258 0.0007 SLI15 YBR156C Member of the CCCH zinc finger family; has similarity to mammalian Tis11 protein, which activates transcription and also has a role in mRNA degradation; may function with Tis11p in iron homeostasis Subunit of the conserved chromosomal passenger complex (CPC; Ipl1p‐Sli15p‐Bir1p‐Nbl1p), which regulates kinetochore‐microtubule attachments, activation of the spindle tension checkpoint, and mitotic spindle disassembly 0.6227 0.0004 SOK1 YDR006C 0.6209 0.0007 Symbols ORF Description MIM1 YOL026C TRS85 Protein whose overexpression suppresses the growth defect of mutants lacking protein kinase A activity; involved in cAMP‐ mediated signaling; localized to the nucleus; similar to the mouse testis‐specific protein PBS13 176 log2(mtr10 .Tf/WT.Tf) adj.p.va l Flavin adenine dinucleotide (FAD) synthetase, performs the second step in synthesis of FAD from riboflavin Transcription factor that stimulates expression of proteasome genes; Rpn4p levels are in turn regulated by the 26S proteasome in a negative feedback control mechanism; RPN4 is transcriptionally regulated by various stress responses 0.6176 0.0006 0.6161 0.0009 Protein of unknown function that associates with ribosomes; has a putative RNA binding domain; in mammals the corresponding protein, eIF2D, has been shown to possess translation initiation factor activity Mitochondrial ribosomal protein of the small subunit, has similarity to E. coli S10 ribosomal protein; essential for viability, unlike most other mitoribosomal proteins 0.6159 0.0007 0.6157 0.0009 YLR408C Putative protein of unknown function; likely member of BLOC complex involved in endosomal cargo sorting; green fluorescent protein (GFP)‐fusion protein localizes to the endosome; YLR408C is not an essential gene 0.6137 0.0005 NA YDR319C Putative protein of unknown function, identified as an ortholog of the highly conserved FIT family of proteins involved in triglyceride droplet biosynthesis; interacts with Sst2p and Hsp82p in high‐ throughput two‐hybrid screens 0.6133 0.0008 MTC5 YDR128W Protein of unknown function; mtc5 is synthetically sick with cdc13‐1 0.6131 0.0006 RAD30 YDR419W DNA polymerase eta, involved in translesion synthesis during post‐ replication repair; catalyzes the synthesis of DNA opposite cyclobutane pyrimidine dimers and other lesions; mutations in human pol eta are responsible for XPV 0.6125 0.0008 TFB1 YDR311W Subunit of TFIIH and nucleotide excision repair factor 3 complexes, required for nucleotide excision repair, target for transcriptional activators 0.6007 0.0005 Symbols ORF Description FAD1 YDL045C RPN4 YDL020C TMA64 YDR117C RSM10 YDR041W BLS1 177 APPENDIX I: Down-regulated genes in mtr10Δ cells in amino acid starvation condition This section includes the list of transcriptionally differentially expressed genes (cutoff conditions: log2 FC 0.6 and adjusted p-value < 0.001) in mtr10Δ cells in amino acid starved condition. Down-regulated genes in mtr10Δ cells in amino acid starved condition log2(mtr10. Ts/WT.TS) adj.p.v al Nuclear import receptor, mediates the nuclear localization of proteins involved in mRNA‐nucleus export; promotes dissociation of mRNAs from the nucleus‐cytoplasm mRNA shuttling protein Npl3p; required for retrograde import of mature tRNAs ‐6.5259 0.0000 YMR251W Omega class glutathione transferase; putative cytosolic localization ‐3.5739 0.0000 FCY21 YER060W ‐2.9267 0.0000 PNS1 YOR161C Putative purine‐cytosine permease, very similar to Fcy2p but cannot substitute for its function Protein of unknown function; has similarity to Torpedo californica tCTL1p, which is postulated to be a choline transporter, neither null mutation nor overexpression affects choline transport ‐2.9240 0.0000 NA YHR022C ‐2.8403 0.0000 QDR2 YIL121W Putative protein of unknown function; YHR022C is not an essential gene Multidrug transporter of the major facilitator superfamily, required for resistance to quinidine, barban, cisplatin, and bleomycin; may have a role in potassium uptake ‐2.7694 0.0000 NA FRE5 YHR140W YOR384W Putative integral membrane protein of unknown function Putative ferric reductase with similarity to Fre2p; expression induced by low iron levels; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐throughput studies ‐2.6771 ‐2.6487 0.0000 0.0005 ASG7 YJL170C Protein that regulates signaling from a G protein beta subunit Ste4p and its relocalization within the cell; specific to a‐cells and induced by alpha‐factor ‐2.6013 0.0001 NA YHL012W Putative protein of unknown function, has some homology to Ugp1p, which encodes UDP‐glucose pyrophosphorylase ‐2.5857 0.0000 Symbols ORF Description MTR10 YOR160W GTO3 178 log2(mtr10. Ts/WT.TS) adj.p.v al Meiosis‐specific DNA binding protein that displays Red1p dependent localization to the unsynapsed axial‐lateral elements of the synaptonemal complex; required for homologous chromosome synapsis and chiasma formation ‐2.5684 0.0001 YLR136C mRNA‐binding protein expressed during iron starvation; binds to a sequence element in the 3'‐untranslated regions of specific mRNAs to mediate their degradation; involved in iron homeostasis ‐2.5208 0.0003 ECM13 YBL043W ‐2.4759 0.0002 IRC15 YPL017C Non‐essential protein of unknown function; induced by treatment with 8‐methoxypsoralen and UVA irradiation Microtubule associated protein; regulates microtubule dynamics; required for accurate meiotic chromosome segregation; null mutant displays large budded cells due to delayed mitotic progression, increased levels of spontaneous Rad52 foci ‐2.4546 0.0000 FIT2 YOR382W Mannoprotein that is incorporated into the cell wall via a glycosylphosphatidylinositol (GPI) anchor, involved in the retention of siderophore‐iron in the cell wall ‐2.4289 0.0001 NA NA RGS2 NA NA YOR107W NA NA Negative regulator of glucose‐induced cAMP signaling; directly activates the GTPase activity of the heterotrimeric G protein alpha subunit Gpa2p ‐2.3974 ‐2.3489 ‐2.2947 0.0000 0.0001 0.0000 NA YHL044W Putative integral membrane protein, member of DUP240 gene family; green fluorescent protein (GFP)‐fusion protein localizes to the plasma membrane in a punctate pattern ‐2.2924 0.0000 BSC5 YNR069C Protein of unknown function, ORF exhibits genomic organization compatible with a translational readthrough‐dependent mode of expression ‐2.2588 0.0000 SFK1 YKL051W ‐2.2187 0.0000 NA YOL163W ‐2.2123 0.0002 BDH2 YAL061W ‐2.2117 0.0001 DCI1 YOR180C ‐2.0800 0.0000 FTR1 YER145C Plasma membrane protein that may act together with or upstream of Stt4p to generate normal levels of the essential phospholipid PI4P, at least partially mediates proper localization of Stt4p to the plasma membrane Putative protein of unknown function; member of the Dal5p subfamily of the major facilitator family Putative medium‐chain alcohol dehydrogenase with similarity to BDH1; transcription induced by constitutively active PDR1 and PDR3 Peroxisomal protein; identification as a delta(3,5)‐delta(2,4)‐dienoyl‐ CoA isomerase involved in fatty acid metabolism is disputed High affinity iron permease involved in the transport of iron across the plasma membrane; forms complex with Fet3p; expression is regulated by iron ‐2.0392 0.0001 ARN1 YHL040C ‐1.9903 0.0000 PTH1 YHR189W ‐1.9587 0.0004 RSB1 YOR049C Transporter, member of the ARN family of transporters that specifically recognize siderophore‐iron chelates; responsible for uptake of iron bound to ferrirubin, ferrirhodin, and related siderophores One of two (see also PTH2) mitochondrially‐localized peptidyl‐tRNA hydrolases; dispensable for respiratory growth on rich medium, but required for respiratory growth on minimal medium Suppressor of sphingoid long chain base (LCB) sensitivity of an LCB‐ lyase mutation; putative integral membrane transporter or flippase that may transport LCBs from the cytoplasmic side toward the extracytoplasmic side of the membrane ‐1.9518 0.0001 ENT4 YLL038C ‐1.9459 0.0002 Symbols ORF Description HOP1 YIL072W TIS11 Protein of unknown function, contains an N‐terminal epsin‐like domain; proposed to be involved in the trafficking of Arn1p in the absence of ferrichrome 179 log2(mtr10. Ts/WT.TS) adj.p.v al Cell wall mannoprotein that localizes specifically to birth scars of daughter cells, linked to a beta‐1,3‐ and beta‐1,6‐glucan heteropolymer through a phosphodiester bond; required for propionic acid resistance ‐1.9204 0.0001 Transporter, member of the ARN family of transporters that specifically recognize siderophore‐iron chelates; responsible for uptake of iron bound to the siderophore triacetylfusarinine C Phosphoglucomutase, catalyzes the conversion from glucose‐1‐ phosphate to glucose‐6‐phosphate, which is a key step in hexose metabolism; functions as the acceptor for a Glc‐phosphotransferase Putative GPI protein Peroxisomal 2,4‐dienoyl‐CoA reductase, auxiliary enzyme of fatty acid beta‐oxidation; homodimeric enzyme required for growth and sporulation on petroselineate medium; expression induced during late sporulation and in the presence of oleate ‐1.9007 0.0002 ‐1.8989 0.0000 ‐1.8971 ‐1.8932 0.0000 0.0000 YHL018W Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to mitochondria and is induced in response to the DNA‐damaging agent MMS ‐1.8807 0.0001 VPS73 YGL104C ‐1.8740 0.0001 VMR1 YHL035C Mitochondrial protein; mutation affects vacuolar protein sorting; putative transporter; member of the sugar porter family Vacuolar membrane protein involved in multiple drug resistance and metal sensitivity; ATP‐binding cassette (ABC) family member involved in drug transport; potential Cdc28p substrate; induced under respiratory conditions ‐1.8671 0.0000 ICS2 YBR157C Protein of unknown function; null mutation does not confer any obvious defects in growth, spore germination, viability, or carbohydrate utilization ‐1.8568 0.0001 SAP4 YGL229C ‐1.8559 0.0000 BAR1 YIL015W ‐1.8466 0.0002 NRT1 YOR071C ‐1.8354 0.0001 NA YPL113C ‐1.8255 0.0001 MAL13 YGR288W ‐1.8198 0.0001 FIT3 YOR383C Protein required for function of the Sit4p protein phosphatase, member of a family of similar proteins that form complexes with Sit4p, including Sap155p, Sap185p, and Sap190p Aspartyl protease secreted into the periplasmic space of mating type a cells, helps cells find mating partners, cleaves and inactivates alpha factor allowing cells to recover from alpha‐factor‐induced cell cycle arrest High‐affinity nicotinamide riboside transporter; also transports thiamine with low affinity; shares sequence similarity with Thi7p and Thi72p; proposed to be involved in 5‐fluorocytosine sensitivity Glyoxylate reductase; acts on glyoxylate and hydroxypyruvate substrates; YPL113C is not an essential gene MAL‐activator protein, part of complex locus MAL1; nonfunctional in genomic reference strain S288C Mannoprotein that is incorporated into the cell wall via a glycosylphosphatidylinositol (GPI) anchor, involved in the retention of siderophore‐iron in the cell wall ‐1.8045 0.0002 CMC4 YMR194C‐ B Protein that localizes to the mitochondrial intermembrane space via the Mia40p‐Erv1p system; contains twin cysteine‐x(9)‐cysteine motifs ‐1.8038 0.0000 AAR2 YBL074C ‐1.7559 0.0000 FRE8 YLR047C Component of the U5 snRNP, required for splicing of U3 precursors; originally described as a splicing factor specifically required for splicing pre‐mRNA of the MATa1 cistron Protein with sequence similarity to iron/copper reductases, involved in iron homeostasis; deletion mutant has iron deficiency/accumulation growth defects; expression increased in the absence of copper‐responsive transcription factor Mac1p ‐1.7489 0.0006 Symbols ORF Description CWP1 YKL096W ARN2 YHL047C PGM2 YMR105C NA SPS19 YAR066W YNL202W NA 180 log2(mtr10. Ts/WT.TS) adj.p.v al Receptor for alpha‐factor pheromone; seven transmembrane‐ domain GPCR that interacts with both pheromone and a heterotrimeric G protein to initiate the signaling response that leads to mating between haploid a and alpha cells ‐1.7461 0.0003 Protein of unknown function, member of the DUP380 subfamily of conserved, often subtelomerically‐encoded proteins Putative protein of unknown function; non‐essential gene identified in a screen for mutants with increased levels of rDNA transcription; contains two putative transmembrane spans, but no significant homology to other known proteins ‐1.7450 0.0000 ‐1.7288 0.0003 YCR100C YKL217W Putative protein of unknown function Lactate transporter, required for uptake of lactate and pyruvate; phosphorylated; expression is derepressed by transcriptional activator Cat8p during respiratory growth, and repressed in the presence of glucose, fructose, and mannose ‐1.7260 ‐1.7220 0.0001 0.0003 PPT2 YPL148C Phosphopantetheine:protein transferase (PPTase), activates mitochondrial acyl carrier protein (Acp1p) by phosphopantetheinylation ‐1.6967 0.0000 PMA2 YPL036W Plasma membrane H+‐ATPase, isoform of Pma1p, involved in pumping protons out of the cell; regulator of cytoplasmic pH and plasma membrane potential ‐1.6932 0.0000 RDS1 YCR106W ‐1.6585 0.0001 NA YJR154W ‐1.6521 0.0000 FRE2 YKL220C ‐1.6436 0.0000 TPO4 YOR273C Zinc cluster transcription factor involved in conferring resistance to cycloheximide Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm Ferric reductase and cupric reductase, reduces siderophore‐bound iron and oxidized copper prior to uptake by transporters; expression induced by low iron levels but not by low copper levels Polyamine transport protein, recognizes spermine, putrescine, and spermidine; localizes to the plasma membrane; member of the major facilitator superfamily ‐1.6415 0.0000 STE3 YKL178C Receptor for a factor pheromone, couples to MAP kinase cascade to mediate pheromone response; transcribed in alpha cells and required for mating by alpha cells, ligand bound receptors endocytosed and recycled to the plasma membrane; GPCR ‐1.6289 0.0000 ENB1 YOL158C Endosomal ferric enterobactin transporter, expressed under conditions of iron deprivation; member of the major facilitator superfamily; expression is regulated by Rcs1p and affected by chloroquine treatment ‐1.6237 0.0001 NA YLR152C ‐1.6226 0.0000 PRM1 YNL279W Putative protein of unknown function; YLR152C is not an essential gene Pheromone‐regulated multispanning membrane protein involved in membrane fusion during mating; predicted to have 5 transmembrane segments and a coiled coil domain; localizes to the shmoo tip; regulated by Ste12p ‐1.6171 0.0002 GEM1 YAL048C Evolutionarily‐conserved tail‐anchored outer mitochondrial membrane GTPase which regulates mitochondrial morphology; cells lacking Gem1p contain collapsed, globular, or grape‐like mitochondria; not required for pheromone‐induced cell death ‐1.6141 0.0001 PEX18 YHR160C Peroxin required for targeting of peroxisomal matrix proteins containing PTS2; interacts with Pex7p; partially redundant with Pex21p ‐1.5940 0.0002 Symbols ORF Description STE2 YFL026W COS9 YKL219W RRT6 YGL146C NA JEN1 181 log2(mtr10. Ts/WT.TS) adj.p.v al GTPase‐activating protein for Gpa1p, regulates desensitization to alpha factor pheromone; also required to prevent receptor‐ independent signaling of the mating pathway; member of the RGS (regulator of G‐protein signaling) family Glycogen synthase with similarity to Gsy2p, the more highly expressed yeast homolog; expression induced by glucose limitation, nitrogen starvation, environmental stress, and entry into stationary phase ‐1.5914 0.0002 ‐1.5879 0.0001 Ceramide synthase component, involved in synthesis of ceramide from C26(acyl)‐coenzyme A and dihydrosphingosine or phytosphingosine, functionally equivalent to Lac1p Serine‐rich protein that contains a basic‐helix‐loop‐helix (bHLH) DNA binding motif; binds E‐boxes of glycolytic genes and contributes to their activation; may function as a transcriptional activator in Ty1‐ mediated gene expression Ferrioxamine B transporter, member of the ARN family of transporters that specifically recognize siderophore‐iron chelates; transcription is induced during iron deprivation and diauxic shift; potentially phosphorylated by Cdc28p ‐1.5733 0.0000 ‐1.5672 0.0001 ‐1.5529 0.0004 YEL048C Protein that interacts with subunits of the TRAPP complex and may play a role its assembly or stability; mutation is synthetically lethal with gcs1 deletion; Sedlin_N family member; human Sedlin mutations cause the skeletal disorder SEDT ‐1.5262 0.0001 0.0006 ‐1.5230 0.0005 SCS7 YMR272C ‐1.5021 0.0000 FCY22 ‐1.5015 0.0001 NFT1 YER060W‐ A YKR103W ‐1.5011 0.0006 PPM2 YOL141W Putative protein of unknown function; identified by fungal homology and RT‐PCR NADP(+)‐dependent glutamate dehydrogenase, synthesizes glutamate from ammonia and alpha‐ketoglutarate; rate of alpha‐ ketoglutarate utilization differs from Gdh1p; expression regulated by nitrogen and carbon sources Sphingolipid alpha‐hydroxylase, functions in the alpha‐hydroxylation of sphingolipid‐associated very long chain fatty acids, has both cytochrome b5‐like and hydroxylase/desaturase domains, not essential for growth Putative purine‐cytosine permease, very similar to Fcy2p but cannot substitute for its function Putative transporter of the multidrug resistance‐associated protein (MRP) subfamily; adjacent ORFs YKR103W and YKR104W are merged in different strain backgrounds. AdoMet‐dependent tRNA methyltransferase also involved in methoxycarbonylation; required for the synthesis of wybutosine (yW), a modified guanosine found at the 3'‐position adjacent to the anticodon of phe‐tRNA; similarity to Ppm1p ‐1.5260 GDH3 YOR316C‐ A YAL062W ‐1.4985 0.0001 SDP1 YIL113W Stress‐inducible dual‐specificity MAP kinase phosphatase, negatively regulates Slt2p MAP kinase by direct dephosphorylation, diffuse localization under normal conditions shifts to punctate localization after heat shock ‐1.4830 0.0006 NA YCR061W ‐1.4802 0.0001 NA YJL160C Protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm in a punctate pattern; induced by treatment with 8‐methoxypsoralen and UVA irradiation Putative protein of unknown function; member of the PIR (proteins with internal repeats) family of cell wall proteins; non‐essential gene that is required for sporulation; mRNA is weakly cell cycle regulated, peaking in mitosis ‐1.4756 0.0002 CRC1 YOR100C ‐1.4702 0.0001 Symbols ORF Description SST2 YLR452C GSY1 YFR015C LAG1 YHL003C TYE7 YOR344C SIT1 YEL065W TCA17 NA Mitochondrial inner membrane carnitine transporter, required for carnitine‐dependent transport of acetyl‐CoA from peroxisomes to mitochondria during fatty acid beta‐oxidation 182 log2(mtr10. Ts/WT.TS) adj.p.v al Member of the multi‐drug and toxin extrusion (MATE) family of the multidrug/oligosaccharidyl‐lipid/polysaccharide (MOP) exporter superfamily; overproduction confers ethionine resistance and accumulation of S‐adenosylmethionine ‐1.4639 0.0001 Transporter of thiamine or related compound; shares sequence similarity with Thi7p Repressor of G1 transcription that binds to SCB binding factor (SBF) at SCB target promoters in early G1; phosphorylation of Whi5p by the CDK, Cln3p/Cdc28p relieves repression and promoter binding by Whi5; periodically expressed in G1 ‐1.4613 0.0002 ‐1.4612 0.0007 Essential protein required for pre‐mRNA splicing; associates transiently with the spliceosomal NTC ("nineteen complex") and acts after Prp2p to promote the first catalytic reaction of splicing ‐1.4600 0.0001 Ferro‐O2‐oxidoreductase required for high‐affinity iron uptake and involved in mediating resistance to copper ion toxicity, belongs to class of integral membrane multicopper oxidases Protein of unknown function; interacts weakly with Knr4p; YMR262W is not an essential gene Galactose‐1‐phosphate uridyl transferase, synthesizes glucose‐1‐ phosphate and UDP‐galactose from UDP‐D‐glucose and alpha‐D‐ galactose‐1‐phosphate in the second step of galactose catabolism ‐1.4573 0.0003 ‐1.4455 0.0000 ‐1.4409 0.0001 Putative protein of unknown function; non‐essential gene; interacts genetically with CHS5, a gene involved in chitin biosynthesis Putative divalent metal ion transporter involved in iron homeostasis; transcriptionally regulated by metal ions; member of the Nramp family of metal transport proteins ‐1.4402 0.0001 ‐1.4194 0.0000 Putative paralog of ATR1, but not required for boron tolerance; identified as a heat‐induced gene in a high‐throughout screen; YMR279C is not an essential gene ‐1.4192 0.0000 YPL147W Subunit of a heterodimeric peroxisomal ATP‐binding cassette transporter complex (Pxa1p‐Pxa2p), required for import of long‐ chain fatty acids into peroxisomes; similarity to human adrenoleukodystrophy transporter and ALD‐related proteins ‐1.4104 0.0001 DCR2 YLR361C Phosphoesterase involved in downregulation of the unfolded protein response, at least in part via dephosphorylation of Ire1p; dosage‐ dependent positive regulator of the G1/S phase transition through control of the timing of START ‐1.3977 0.0001 NA YJL213W Protein of unknown function that may interact with ribosomes; periodically expressed during the yeast metabolic cycle; phosphorylated in vitro by the mitotic exit network (MEN) kinase complex, Dbf2p/Mob1p ‐1.3949 0.0000 ATG29 YPL166W Autophagy‐specific protein that is required for recruitment of other ATG proteins to the pre‐autophagosomal structure (PAS); interacts with Atg17p and localizas to the PAS in a manner interdependent with Atg17p and Cis1p; not conserved ‐1.3914 0.0002 SKN1 YGR143W ‐1.3789 0.0000 GSY2 YLR258W Protein involved in sphingolipid biosynthesis; type II membrane protein with similarity to Kre6p Glycogen synthase, similar to Gsy1p; expression induced by glucose limitation, nitrogen starvation, heat shock, and stationary phase; activity regulated by cAMP‐dependent, Snf1p and Pho85p kinases as well as by the Gac1p‐Glc7p phosphatase ‐1.3707 0.0003 Symbols ORF Description ERC1 YHR032W THI72 YOR192C WHI5 YOR083W YJU2 YKL095W FET3 YMR058W NA YMR262W GAL7 YBR018C NA YGL081W SMF3 YLR034C NA YMR279C PXA1 183 log2(mtr10. Ts/WT.TS) adj.p.v al Putative protein of unknown function with similarity to hexokinases; transcript is upregulated during sporulation and the unfolded protein response; YLR446W is not an essential gene ‐1.3653 0.0000 YHR105W Endosomal protein of unknown function that contains a phox (PX) homology domain and binds to both phosphatidylinositol‐3‐ phosphate (PtdIns(3)P) and proteins involved in ER‐Golgi or vesicular transport ‐1.3632 0.0009 AUS1 YOR011W ‐1.3577 0.0003 MCH5 YOR306C Transporter of the ATP‐binding cassette family, involved in uptake of sterols and anaerobic growth Plasma membrane riboflavin transporter; facilitates the uptake of vitamin B2; required for FAD‐dependent processes; sequence similarity to mammalian monocarboxylate permeases, however mutants are not deficient in monocarboxylate transport ‐1.3486 0.0002 VBA2 ATP10 YBR293W YLR393W Permease of basic amino acids in the vacuolar membrane Mitochondrial inner membrane protein required for assembly of the F0 sector of mitochondrial F1F0 ATP synthase, interacts genetically with ATP6 ‐1.3380 ‐1.3348 0.0002 0.0002 SPS100 YHR139C Protein required for spore wall maturation; expressed during sporulation; may be a component of the spore wall; expression also induced in cells treated with the mycotoxin patulin ‐1.3317 0.0001 RPS28B YLR264W ‐1.3233 0.0001 MRS4 YKR052C Protein component of the small (40S) ribosomal subunit; nearly identical to Rps28Ap and has similarity to rat S28 ribosomal protein Iron transporter that mediates Fe2+ transport across the inner mitochondrial membrane; mitochondrial carrier family member, similar to and functionally redundant with Mrs3p; active under low‐ iron conditions; may transport other cations ‐1.3206 0.0003 NA YGR117C ‐1.3190 0.0001 MMT2 YPL224C ‐1.3181 0.0000 ROD1 YOR018W ‐1.3108 0.0006 KRE5 YOR336W ‐1.3035 0.0000 NA YLR126C Protein required for beta‐1,6 glucan biosynthesis; mutations result in aberrant morphology and severe growth defects Putative protein of unknown function with similarity to glutamine amidotransferase proteins; has Aft1p‐binding motif in the promoter; may be involved in copper and iron homeostasis; YLR126C is not an essential protein ‐1.3034 0.0002 DTR1 YBR180W Putative dityrosine transporter, required for spore wall synthesis; expressed during sporulation; member of the major facilitator superfamily (DHA1 family) of multidrug resistance transporters ‐1.3009 0.0002 NA YKR104W ‐1.2994 0.0000 RTC4 YNL254C Putative transporter of the multidrug resistance‐associated protein (MRP) subfamily; contains a stop codon in S288C; adjacent ORFs YKR103W and YKR104W are merged in different strain backgrounds Protein of unknown function; null mutation suppresses cdc13‐1 temperature sensitivity; (GFP)‐fusion protein localizes to both the cytoplasm and the nucleus ‐1.2966 0.0000 NA NA YER085C YKR041W ‐1.2916 ‐1.2874 0.0004 0.0008 Symbols ORF Description NA YLR446W YPT35 Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm Putative metal transporter involved in mitochondrial iron accumulation; closely related to Mmt1p Membrane protein that binds the ubiquitin ligase Rsp5p via its 2 PY motifs; overexpression confers resistance to the GST substrate o‐ dinitrobenzene,zinc, and calcium; proposed to regulate the endocytosis of plasma membrane proteins Putative protein of unknown function Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm and nucleus 184 log2(mtr10. Ts/WT.TS) adj.p.v al Protein required for oxidation of specific cysteine residues of the transcription factor Yap1p, resulting in the nuclear localization of Yap1p in response to stress ‐1.2870 0.0002 Minor isoform of pyruvate decarboxylase, decarboxylates pyruvate to acetaldehyde, involved in amino acid catabolism; transcription is glucose‐ and ethanol‐dependent, and is strongly induced during sulfur limitation Lipase required for intravacuolar lysis of autophagic bodies and Cvt bodies; targeted to intravacuolar vesicles during autophagy via the multivesicular body (MVB) pathway ‐1.2821 0.0001 ‐1.2792 0.0002 YMR013C Dolichol kinase, catalyzes the terminal step in dolichyl monophosphate (Dol‐P) biosynthesis; required for viability and for normal rates of lipid intermediate synthesis and protein N‐ glycosylation ‐1.2789 0.0000 PRY1 SFP1 YJL079C YLR403W Protein of unknown function Transcription factor that controls expression of ribosome biogenesis genes in response to nutrients and stress, regulates G2/M transitions during mitotic cell cycle and DNA‐damage response, modulates cell size; regulated by TORC1 and Mrs6p ‐1.2753 ‐1.2672 0.0000 0.0002 RPL8A YHL033C ‐1.2662 0.0000 RRN10 YBL025W ‐1.2646 0.0000 MPD2 YOL088C Ribosomal protein L4 of the large (60S) ribosomal subunit, nearly identical to Rpl8Bp and has similarity to rat L7a ribosomal protein; mutation results in decreased amounts of free 60S subunits Protein involved in promoting high level transcription of rDNA, subunit of UAF (upstream activation factor) for RNA polymerase I Member of the protein disulfide isomerase (PDI) family, exhibits chaperone activity; overexpression suppresses the lethality of a pdi1 deletion but does not complement all Pdi1p functions; undergoes oxidation by Ero1p ‐1.2607 0.0000 NBL1 YHR199C‐ A Subunit of the conserved chromosomal passenger complex (CPC; Ipl1p‐Sli15p‐Bir1p‐Nbl1p), which regulates mitotic chromosome segregation; not required for the kinase activity of the complex; mediates the interaction of Sli15p and Bir1p ‐1.2588 0.0004 NA YPL264C ‐1.2550 0.0000 FUS1 YCL027W Putative membrane protein of unknown function; physically interacts with Hsp82p; YPL264C is not an essential gene Membrane protein localized to the shmoo tip, required for cell fusion; expression regulated by mating pheromone; proposed to coordinate signaling, fusion, and polarization events required for fusion; potential Cdc28p substrate ‐1.2361 0.0004 FMP43 YGR243W ‐1.2345 0.0006 BDS1 YOL164W ‐1.2324 0.0003 AVT6 YER119C ‐1.2318 0.0002 NA YGR205W ‐1.2307 0.0005 SIP4 YJL089W ‐1.2269 0.0004 Symbols ORF Description YBP1 YBR216C PDC6 YGR087C ATG15 YCR068W SEC59 Putative protein of unknown function; expression regulated by osmotic and alkaline stresses; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐throughput studies Bacterially‐derived sulfatase required for use of alkyl‐ and aryl‐ sulfates as sulfur sources Vacuolar aspartate and glutamate exporter; member of a family of seven genes (AVT1‐7) related to vesicular GABA‐glycine transporters; involved in compartmentalizing acidic amino acids in response to nitrogen starvation ATP‐binding protein of unknown function; crystal structure resembles that of E.coli pantothenate kinase and other small kinases C6 zinc cluster transcriptional activator that binds to the carbon source‐responsive element (CSRE) of gluconeogenic genes; involved in the positive regulation of gluconeogenesis; regulated by Snf1p protein kinase; localized to the nucleus 185 log2(mtr10. Ts/WT.TS) adj.p.v al Transcriptional activator related to Msn2p; activated in stress conditions, which results in translocation from the cytoplasm to the nucleus; binds DNA at stress response elements of responsive genes, inducing gene expression ‐1.2260 0.0001 YHR210C Putative protein of unknown function; non‐essential gene; highly expressed under anaeorbic conditions; sequence similarity to aldose 1‐epimerases such as GAL10 ‐1.2259 0.0000 SMF1 YOL122C Divalent metal ion transporter with a broad specificity for di‐valent and tri‐valent metals; post‐translationally regulated by levels of metal ions; member of the Nramp family of metal transport proteins ‐1.2249 0.0001 NA SDS3 NA YIL084C NA Component of the Rpd3p/Sin3p deacetylase complex required for its structural integrity and catalytic activity, involved in transcriptional silencing and required for sporulation; cells defective in SDS3 display pleiotropic phenotypes ‐1.2236 ‐1.2199 0.0002 0.0002 VAC8 YEL013W ‐1.2193 0.0002 ELO1 YJL196C ‐1.2160 0.0000 ULA1 YPL003W Phosphorylated and palmitoylated vacuolar membrane protein that interacts with Atg13p, required for the cytoplasm‐to‐vacuole targeting (Cvt) pathway; interacts with Nvj1p to form nucleus‐ vacuole junctions Elongase I, medium‐chain acyl elongase, catalyzes carboxy‐terminal elongation of unsaturated C12‐C16 fatty acyl‐CoAs to C16‐C18 fatty acids Protein that acts together with Uba3p to activate Rub1p before its conjugation to proteins (neddylation), which may play a role in protein degradation ‐1.2092 0.0002 NA YOR214C ‐1.2084 0.0002 BNA4 YBL098W Putative protein of unknown function; YOR214C is not an essential gene Kynurenine 3‐mono oxygenase, required for the de novo biosynthesis of NAD from tryptophan via kynurenine; expression regulated by Hst1p; putative therapeutic target for Huntington disease ‐1.2011 0.0000 MUC1 YIR019C GPI‐anchored cell surface glycoprotein (flocculin) required for pseudohyphal formation, invasive growth, flocculation, and biofilms; transcriptionally regulated by the MAPK pathway (via Ste12p and Tec1p) and the cAMP pathway (via Flo8p) ‐1.1963 0.0002 NA ICL1 YOL014W YER065C Putative protein of unknown function Isocitrate lyase, catalyzes the formation of succinate and glyoxylate from isocitrate, a key reaction of the glyoxylate cycle; expression of ICL1 is induced by growth on ethanol and repressed by growth on glucose ‐1.1961 ‐1.1951 0.0004 0.0006 RTG1 YOL067C ‐1.1927 0.0002 GYP6 YJL044C ‐1.1918 0.0000 PDR5 YOR153W Transcription factor (bHLH) involved in interorganelle communication between mitochondria, peroxisomes, and nucleus GTPase‐activating protein (GAP) for the yeast Rab family member, Ypt6p; involved in vesicle mediated protein transport Plasma membrane ATP‐binding cassette (ABC) transporter, multidrug transporter actively regulated by Pdr1p; also involved in steroid transport, cation resistance, and cellular detoxification during exponential growth ‐1.1879 0.0001 NA SFG1 YNR068C YOR315W Putative protein of unknown function Nuclear protein, putative transcription factor required for growth of superficial pseudohyphae (which do not invade the agar substrate) but not for invasive pseudohyphal growth; may act together with Phd1p; potential Cdc28p substrate ‐1.1876 ‐1.1804 0.0002 0.0006 YAP5 YIR018W Basic leucine zipper (bZIP) transcription factor ‐1.1781 0.0000 Symbols ORF Description MSN4 YKL062W NA 186 log2(mtr10. Ts/WT.TS) adj.p.v al ‐1.1708 0.0001 ‐1.1673 0.0000 Protein containing a UCS (UNC‐45/CRO1/SHE4) domain, binds to myosin motor domains to regulate myosin function; involved in endocytosis, polarization of the actin cytoskeleton, and asymmetric mRNA localization ‐1.1664 0.0000 ATP‐NADH kinase; phosphorylates both NAD and NADH; homooctameric structure consisting of 60‐kDa subunits; sequence similarity to Utr1p and Pos5p; overexpression complements certain pos5 phenotypes Mitochondrial 3‐oxoacyl‐[acyl‐carrier‐protein] reductase, may comprise a type II mitochondrial fatty acid synthase along with Mct1p ‐1.1660 0.0002 ‐1.1610 0.0001 YLR031W YOR080W Putative protein of unknown function Origin‐binding F‐box protein that forms an SCF ubiquitin ligase complex with Skp1p and Cdc53p; plays a role in DNA replication, involved in invasive and pseudohyphal growth ‐1.1590 ‐1.1574 0.0001 0.0002 STE12 YHR084W Transcription factor that is activated by a MAP kinase signaling cascade, activates genes involved in mating or pseudohyphal/invasive growth pathways; cooperates with Tec1p transcription factor to regulate genes specific for invasive growth ‐1.1567 0.0001 NA YER039C‐A ‐1.1560 0.0003 GLN3 YER040W Putative protein of unknown function; YER039C‐A is not an essential gene Transcriptional activator of genes regulated by nitrogen catabolite repression (NCR), localization and activity regulated by quality of nitrogen source ‐1.1541 0.0003 AGX1 YFL030W Alanine:glyoxylate aminotransferase (AGT), catalyzes the synthesis of glycine from glyoxylate, which is one of three pathways for glycine biosynthesis in yeast; has similarity to mammalian and plant alanine:glyoxylate aminotransferases ‐1.1526 0.0001 YPC1 YBR183W ‐1.1476 0.0001 ROG3 YFR022W Alkaline ceramidase that also has reverse (CoA‐independent) ceramide synthase activity, catalyzes both breakdown and synthesis of phytoceramide; overexpression confers fumonisin B1 resistance Protein that binds the ubiquitin ligase Rsp5p via its 2 PY motifs; has similarity to Rod1p; mutation suppresses the temperature sensitivity of an mck1 rim11 double mutant; proposed to regulate the endocytosis of plasma membrane proteins ‐1.1472 0.0001 TDH1 YJL052W Glyceraldehyde‐3‐phosphate dehydrogenase, isozyme 1, involved in glycolysis and gluconeogenesis; tetramer that catalyzes the reaction of glyceraldehyde‐3‐phosphate to 1,3 bis‐phosphoglycerate; detected in the cytoplasm and cell wall ‐1.1443 0.0003 NA YMR253C Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm in a punctate pattern; YMR253C is not an essential gene ‐1.1336 0.0003 DFG16 YOR030W Probable multiple transmembrane protein, involved in diploid invasive and pseudohyphal growth upon nitrogen starvation; required for accumulation of processed Rim101p ‐1.1248 0.0003 Symbols ORF Description PIC2 YER053C PEX32 YBR168W Mitochondrial phosphate carrier, imports inorganic phosphate into mitochondria; functionally redundant with Mir1p but less abundant than Mir1p under normal conditions; expression is induced at high temperature Peroxisomal integral membrane protein, involved in negative regulation of peroxisome size; partially functionally redundant with Pex31p; genetic interactions suggest action at a step downstream of steps mediated by Pex28p and Pex29p SHE4 YOR035C YEF1 YEL041W OAR1 YKL055C NA DIA2 187 log2(mtr10. Ts/WT.TS) adj.p.v al ‐1.1127 0.0004 ‐1.1111 ‐1.1105 0.0002 0.0000 One of two redundant DL‐glycerol‐3‐phosphatases (RHR2/GPP1 encodes the other) involved in glycerol biosynthesis; induced in response to hyperosmotic stress and oxidative stress, and during the diauxic transition ‐1.1097 0.0005 YBR093C Repressible acid phosphatase (1 of 3) that also mediates extracellular nucleotide‐derived phosphate hydrolysis; secretory pathway derived cell surface glycoprotein; induced by phosphate starvation and coordinately regulated by PHO4 and PHO2 ‐1.1082 0.0001 RRT13 YER066W ‐1.1061 0.0001 PHO80 YOL001W Putative protein of unknown function; non‐essential gene identified in a screen for mutants with decreased levels of rDNA transcription Cyclin, interacts with cyclin‐dependent kinase Pho85p; regulates the response to nutrient levels and environmental conditions, including the response to phosphate limitation and stress‐dependent calcium signaling ‐1.1057 0.0001 NA YEL025C ‐1.1055 0.0002 FEN2 YCR028C ‐1.1051 0.0001 ICP55 YER078C Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to both the cytoplasm and the nucleus Plasma membrane H+‐pantothenate symporter; confers sensitivity to the antifungal agent fenpropimorph Mitochondrial aminopeptidase; cleaves the N termini of at least 38 imported proteins after cleavage by the mitochondrial processing peptidase (MPP), thereby increasing their stability; member of the aminopeptidase P family ‐1.1051 0.0000 PDR18 YNR070W Putative transporter of the ATP‐binding cassette (ABC) family, implicated in pleiotropic drug resistance; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐ throughput studies ‐1.1007 0.0001 FIT1 YDR534C Mannoprotein that is incorporated into the cell wall via a glycosylphosphatidylinositol (GPI) anchor, involved in the retention of siderophore‐iron in the cell wall ‐1.0976 0.0006 SGF29 YCL010C Probable subunit of SAGA histone acetyltransferase complex ‐1.0947 0.0002 YPK9 YOR291W Vacuolar protein with a possible role in sequestering heavy metals; has similarity to the type V P‐type ATPase Spf1p; homolog of human ATP13A2 (PARK9), mutations in which are associated with Parkinson disease and Kufor‐Rakeb syndrome ‐1.0924 0.0001 SRB8 YCR081W Subunit of the RNA polymerase II mediator complex; associates with core polymerase subunits to form the RNA polymerase II holoenzyme; essential for transcriptional regulation; involved in glucose repression ‐1.0917 0.0001 PDR15 YDR406W Plasma membrane ATP binding cassette (ABC) transporter, multidrug transporter and general stress response factor implicated in cellular detoxification; regulated by Pdr1p, Pdr3p and Pdr8p; promoter contains a PDR responsive element ‐1.0877 0.0001 YPK2 YMR104C Protein kinase with similarity to serine/threonine protein kinase Ypk1p; functionally redundant with YPK1 at the genetic level; participates in a signaling pathway required for optimal cell wall integrity; homolog of mammalian kinase SGK ‐1.0858 0.0009 Symbols ORF Description YSP1 YHR155W NA PPE1 YLR173W YHR075C Mitochondrial protein with a potential role in promoting mitochondrial fragmentation during programmed cell death in response to high levels of alpha‐factor mating pheromone or the drug amiodarone Putative protein of unknown function Protein with carboxyl methyl esterase activity that may have a role in demethylation of the phosphoprotein phosphatase catalytic subunit; also identified as a small subunit mitochondrial ribosomal protein HOR2 YER062C PHO5 188 log2(mtr10. Ts/WT.TS) adj.p.v al Putative protein of unknown function with similarity to a human minor histocompatibility antigen and signal peptide peptidases; YKL100C is not an essential gene ‐1.0832 0.0000 YPL095C Acyl‐coenzymeA:ethanol O‐acyltransferase responsible for the major part of medium‐chain fatty acid ethyl ester biosynthesis during fermentation; possesses short‐chain esterase activity; may be involved in lipid metabolism and detoxification ‐1.0828 0.0002 NA YPR071W Putative membrane protein; YPR071W is not an essential gene ‐1.0822 0.0000 GUT2 YIL155C Mitochondrial glycerol‐3‐phosphate dehydrogenase; expression is repressed by both glucose and cAMP and derepressed by non‐ fermentable carbon sources in a Snf1p, Rsf1p, Hap2/3/4/5 complex dependent manner ‐1.0817 0.0004 TPO1 YLL028W Polyamine transporter that recognizes spermine, putrescine, and spermidine; catalyzes uptake of polyamines at alkaline pH and excretion at acidic pH; phosphorylation enhances activity and sorting to the plasma membrane ‐1.0812 0.0001 FRE6 YLL051C ‐1.0798 0.0001 NA YNL040W Putative ferric reductase with similarity to Fre2p; expression induced by low iron levels Putative protein of unknown function with strong similarity to alanyl‐ tRNA synthases from Eubacteria; green fluorescent protein (GFP)‐ fusion protein localizes to the cytoplasm; YNL040W is not an essential gene ‐1.0787 0.0002 NA YPR157W ‐1.0739 0.0006 AQY2 YLL052C ‐1.0738 0.0001 NA YMR102C Protein of unknown function; transcription is activated by paralogous transcription factors Yrm1p and Yrr1p along with genes involved in multidrug resistance; mutant shows increased resistance to azoles; YMR102C is not an essential gene ‐1.0731 0.0001 TAL1 YLR354C ‐1.0717 0.0000 NA YLR164W Transaldolase, enzyme in the non‐oxidative pentose phosphate pathway; converts sedoheptulose 7‐phosphate and glyceraldehyde 3‐phosphate to erythrose 4‐phosphate and fructose 6‐phosphate Mitochondrial inner membrane of unknown function; similar to Tim18p and Sdh4p; expression induced by nitrogen limitation in a GLN3, GAT1‐dependent manner ‐1.0683 0.0000 ECL1 YGR146C Protein of unknown function, affects chronological lifespan; induced by iron homeostasis transcription factor Aft2p; multicopy suppressor of temperature sensitive hsf1 mutant; induced by treatment with 8‐ methoxypsoralen and UVA irradiation ‐1.0671 0.0003 PAC1 YOR269W Protein involved in nuclear migration, part of the dynein/dynactin pathway; targets dynein to microtubule tips, which is necessary for sliding of microtubules along bud cortex; synthetic lethal with bni1; homolog of human LIS1 ‐1.0662 0.0001 MAL33 YBR297W ‐1.0646 0.0003 PIP2 YOR363C MAL‐activator protein, part of complex locus MAL3; nonfunctional in genomic reference strain S288C Autoregulatory oleate‐specific transcriptional activator of peroxisome proliferation, contains Zn(2)‐Cys(6) cluster domain, forms heterodimer with Oaf1p, binds oleate response elements (OREs), activates beta‐oxidation genes ‐1.0645 0.0001 Symbols ORF Description NA YKL100C EEB1 Putative protein of unknown function; induced by treatment with 8‐ methoxypsoralen and UVA irradiation Water channel that mediates the transport of water across cell membranes, only expressed in proliferating cells, controlled by osmotic signals, may be involved in freeze tolerance; disrupted by a stop codon in many S. cerevisiae strains 189 log2(mtr10. Ts/WT.TS) adj.p.v al Putative protein of unknown function; expression is induced by activation of the HOG1 mitogen‐activated signaling pathway and this induction is Hog1p/Pbs2p dependent; YJL107C and adjacent ORF, YJL108C are merged in related fungi ‐1.0630 0.0000 Self‐glucosylating initiator of glycogen synthesis, also glucosylates n‐ dodecyl‐beta‐D‐maltoside; similar to mammalian glycogenin NA Zinc transporter; localizes to the ER; null mutant is sensitive to calcofluor white, leads to zinc accumulation in cytosol; ortholog of the mouse KE4 and member of the ZIP (ZRT, IRT‐like Protein) family Putative histone acetylase with a role in transcriptional silencing, sequence‐specific activator of histone genes, binds specifically and cooperatively to pairs of UAS elements in core histone promoters, functions at or near the TATA box ‐1.0627 0.0004 ‐1.0611 ‐1.0600 0.0002 0.0001 ‐1.0592 0.0002 Putative protein of unknown function Essential membrane protein localized at the nuclear envelope and spindle pole body (SPB), required for insertion of the newly duplicated SPB into the nuclear envelope; potentially phosphorylated by Cdc28p ‐1.0588 ‐1.0585 0.0002 0.0006 Mitochondrial inner membrane protein with a role in maintaining mitochondrial morphology and normal cardiolipin levels; proposed to be involved in N‐acylethanolamine metabolism; related to mammalian N‐acylPE‐specific phospholipase D Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to both the cytoplasm and the nucleus and is induced in response to the DNA‐damaging agent MMS Protein of unknown function involved in maintenance of proper telomere length NA Putative protein of unknown function; non‐essential gene; contains multiple predicted transmembrane domains Putative zinc finger protein; YPR013C is not an essential gene ‐1.0585 0.0006 ‐1.0449 0.0003 ‐1.0407 0.0001 ‐1.0368 ‐1.0345 0.0004 0.0000 ‐1.0341 0.0000 ‐1.0308 0.0001 ‐1.0293 0.0001 ‐1.0274 0.0001 ‐1.0235 0.0001 Symbols ORF Description NA YJL107C GLG1 YKR058W NA YKE4 NA YIL023C SPT10 YJL127C NA MPS2 YIL089W YGL075C FMP30 YPL103C NA YGR126W NA YPL041C NA NA NA YGL140C NA YPR013C NA YOR152C OST6 YML019W SAD1 YFR005C NIT1 YIL164C HCM1 YCR065W Forkhead transcription factor that drives S‐phase specific expression of genes involved in chromosome segregation, spindle dynamics, and budding; suppressor of calmodulin mutants with specific SPB assembly defects; telomere maintenance role ‐1.0222 0.0004 ZRT2 YLR130C Low‐affinity zinc transporter of the plasma membrane; transcription is induced under low‐zinc conditions by the Zap1p transcription factor ‐1.0193 0.0003 NA PDR3 YAL037W YBL005W Putative protein of unknown function Transcriptional activator of the pleiotropic drug resistance network, regulates expression of ATP‐binding cassette (ABC) transporters through binding to cis‐acting sites known as PDREs (PDR responsive elements) ‐1.0187 ‐1.0176 0.0001 0.0003 Putative protein of unknown function; has no similarity to any known protein; YOR152C is not an essential gene Subunit of the oligosaccharyltransferase complex of the ER lumen, which catalyzes asparagine‐linked glycosylation of newly synthesized proteins; similar to and partially functionally redundant with Ost3p Conserved zinc‐finger domain protein involved in pre‐mRNA splicing, required for assembly of U4 snRNA into the U4/U6 particle Nitrilase, member of the nitrilase branch of the nitrilase superfamily; in closely related species and other S. cerevisiae strain backgrounds YIL164C and adjacent ORF, YIL165C, likely constitute a single ORF encoding a nitrilase gene 190 log2(mtr10. Ts/WT.TS) adj.p.v al Long chain fatty acyl‐CoA synthetase, activates imported fatty acids; green fluorescent protein (GFP)‐fusion protein localizes to the cell periphery ‐1.0170 0.0001 YAL018C YGR031W Putative protein of unknown function Putative protein of unknown function; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐ throughput studies ‐1.0150 ‐1.0138 0.0009 0.0002 ERS1 YCR075C ‐1.0047 0.0001 ARG80 YMR042W ‐1.0047 0.0006 NA ARG8 YBL029W YOL140W ‐0.9999 ‐0.9993 0.0002 0.0001 HST2 YPL015C Protein with similarity to human cystinosin, which is a H(+)‐driven transporter involved in L‐cystine export from lysosomes and implicated in the disease cystinosis; contains seven transmembrane domains Transcription factor involved in regulation of arginine‐responsive genes; acts with Arg81p and Arg82p Non‐essential protein of unknown function Acetylornithine aminotransferase, catalyzes the fourth step in the biosynthesis of the arginine precursor ornithine Cytoplasmic member of the silencing information regulator 2 (Sir2) family of NAD(+)‐dependent protein deacetylases; modulates nucleolar (rDNA) and telomeric silencing; possesses NAD(+)‐ dependent histone deacetylase activity in vitro ‐0.9981 0.0008 EDS1 YBR033W Putative zinc cluster protein; YBR033W is not an essential gene ‐0.9975 0.0003 EAR1 YMR171C ‐0.9972 0.0001 TTI1 YKL033W Specificity factor required for Rsp5p‐dependent ubiquitination and sorting of specific cargo proteins at the multivesicular body; mRNA is targeted to the bud via the mRNA transport system involving She2p Putative protein of unknown function; subunit of the ASTRA complex which is part of the chromatin remodeling machinery; similar to S. pombe Tti1p; detected in highly purified mitochondria in high‐ throughput studies ‐0.9968 0.0002 NA YBL095W Putative protein of unknown function; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐ throughput studies ‐0.9932 0.0001 MAL11 YGR289C ‐0.9924 0.0004 NA YPR003C Inducible high‐affinity maltose transporter (alpha‐glucoside transporter); encoded in the MAL1 complex locus; broad substrate specificity that includes maltotriose; required for isomaltose utilization Putative sulfate permease; physically interacts with Hsp82p; green fluorescent protein (GFP)‐fusion protein localizes to the ER; YPR003C is not an essential gene ‐0.9902 0.0001 MCM3 YEL032W Protein involved in DNA replication; component of the Mcm2‐7 hexameric complex that binds chromatin as a part of the pre‐ replicative complex ‐0.9890 0.0002 NA YPL191C Putative protein of unknown function; diploid deletion strain exhibits high budding index; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm ‐0.9887 0.0002 ALG6 YOR002W Alpha 1,3 glucosyltransferase, involved in transfer of oligosaccharides from dolichyl pyrophosphate to asparagine residues of proteins during N‐linked protein glycosylation; mutations in human ortholog are associated with disease ‐0.9852 0.0002 SER3 YER081W ‐0.9837 0.0001 DSD1 YGL196W 3‐phosphoglycerate dehydrogenase, catalyzes the first step in serine and glycine biosynthesis; isozyme of Ser33p D‐serine dehydratase (aka D‐serine ammonia‐lyase); converts D‐ serine to pyruvate and ammonia by a reaction dependent on pyridoxal 5'‐phosphate and zinc; may play a role in D‐serine detoxification; L‐serine is not a substrate ‐0.9830 0.0002 Symbols ORF Description FAA3 YIL009W NA NA 191 log2(mtr10. Ts/WT.TS) adj.p.v al Probable ortholog of A. nidulans PalC, which is involved in pH regulation and binds to the ESCRT‐III complex; null mutant does not properly process Rim101p and has decreased resistance to rapamycin; GFP‐fusion protein is cytoplasmic ‐0.9814 0.0002 YKL133C Putative protein of unknown function; has similarity to Mgr3p, but unlike MGR3, is not required for growth of cells lacking the mitochondrial genome (null mutation does not confer a petite‐ negative phenotype) ‐0.9811 0.0006 PIR3 YKL163W O‐glycosylated covalently‐bound cell wall protein required for cell wall stability; expression is cell cycle regulated, peaking in M/G1 and also subject to regulation by the cell integrity pathway ‐0.9810 0.0001 THO1 YER063W Conserved nuclear RNA‐binding protein; specifically binds to transcribed chromatin in a THO‐ and RNA‐dependent manner, genetically interacts with shuttling hnRNP NAB2; overproduction suppresses transcriptional defect caused by hpr1 mutation ‐0.9795 0.0002 GUT1 YHL032C Glycerol kinase, converts glycerol to glycerol‐3‐phosphate; glucose repression of expression is mediated by Adr1p and Ino2p‐Ino4p; derepression of expression on non‐fermentable carbon sources is mediated by Opi1p and Rsf1p ‐0.9757 0.0001 HTD2 YHR067W Mitochondrial 3‐hydroxyacyl‐thioester dehydratase involved in fatty acid biosynthesis, required for respiratory growth and for normal mitochondrial morphology ‐0.9748 0.0001 MUM3 YOR298W Protein of unknown function involved in the organization of the outer spore wall layers; has similarity to the tafazzins superfamily of acyltransferases ‐0.9732 0.0010 NA NA YPL236C YOR292C NA Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the vacuole; YOR292C is not an essential gene ‐0.9695 ‐0.9685 0.0001 0.0001 PLB2 YMR006C ‐0.9672 0.0003 NA MRK1 YPR196W YDL079C Phospholipase B (lysophospholipase) involved in phospholipid metabolism; displays transacylase activity in vitro; overproduction confers resistance to lysophosphatidylcholine Putative maltose activator Glycogen synthase kinase 3 (GSK‐3) homolog; one of four GSK‐3 homologs in S. cerevisiae that function to activate Msn2p‐dependent transcription of stress responsive genes and that function in protein degradation ‐0.9672 ‐0.9658 0.0002 0.0002 LYS5 YGL154C Phosphopantetheinyl transferase involved in lysine biosynthesis; converts inactive apo‐form of Lys2p (alpha‐aminoadipate reductase) into catalytically active holo‐form by posttranslational addition of phosphopantetheine ‐0.9650 0.0002 BUR2 YLR226W Cyclin for the Sgv1p (Bur1p) protein kinase; Sgv1p and Bur2p comprise a CDK‐cyclin complex involved in transcriptional regulation through its phosphorylation of the carboxy‐terminal domain of the largest subunit of RNA polymerase II ‐0.9645 0.0000 MNS1 YJR131W ‐0.9639 0.0001 NA YJR030C Alpha‐1,2‐mannosidase involved in ER quality control; catalyzes the removal of one mannose residue from Man9GlcNAc to produce a single isomer of Man8GlcNAc in N‐linked oligosaccharide biosynthesis; integral to ER membrane Putative protein of unknown function; expression repressed in carbon limited vs carbon replete chemostat cultures; YJR030C is a non‐essential gene ‐0.9639 0.0001 Symbols ORF Description NA YGR122W NA 192 log2(mtr10. Ts/WT.TS) adj.p.v al Putative protein of unknown function with similarity to Tfc7p and prokaryotic phosphotransfer enzymes; null mutant shows alterations in glucose metabolism; GFP‐fusion protein localizes to the cytoplasm and nucleus Putative protein of unknown function; expression induced under calcium shortage Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm; overexpression causes a cell cycle delay or arrest ‐0.9619 0.0003 ‐0.9593 0.0004 ‐0.9593 0.0004 YJL164C cAMP‐dependent protein kinase catalytic subunit; promotes vegetative growth in response to nutrients via the Ras‐cAMP signaling pathway; inhibited by regulatory subunit Bcy1p in the absence of cAMP; partially redundant with Tpk2p and Tpk3p ‐0.9571 0.0006 NIS1 YNL078W ‐0.9533 0.0002 STE4 YOR212W ‐0.9501 0.0001 RAD16 YBR114W Protein localized in the bud neck at G2/M phase; physically interacts with septins; possibly involved in a mitotic signaling network G protein beta subunit, forms a dimer with Ste18p to activate the mating signaling pathway, forms a heterotrimer with Gpa1p and Ste18p to dampen signaling; may recruit Rho1p to the polarized growth site during mating; contains WD40 repeats Protein that recognizes and binds damaged DNA in an ATP‐ dependent manner (with Rad7p) during nucleotide excision repair; subunit of Nucleotide Excision Repair Factor 4 (NEF4) and the Elongin‐Cullin‐Socs (ECS) ligase complex ‐0.9497 0.0000 NA YGL036W Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm; YGL036W is not an essential gene ‐0.9474 0.0006 HER1 YOR227W Protein of unknown function required for proliferation or remodeling of the ER that is caused by overexpression of Hmg2p; may interact with ribosomes, based on co‐purification experiments ‐0.9443 0.0002 YAT2 YER024W Carnitine acetyltransferase; has similarity to Yat1p, which is a carnitine acetyltransferase associated with the mitochondrial outer membrane ‐0.9440 0.0008 NA YPL272C ‐0.9436 0.0003 RPL9B YNL067W Putative protein of unknown function; gene expression induced in response to ketoconazole; YPL272C is not an essential gene Protein component of the large (60S) ribosomal subunit, nearly identical to Rpl9Ap and has similarity to E. coli L6 and rat L9 ribosomal proteins ‐0.9432 0.0001 FAR1 YJL157C Cyclin‐dependent kinase inhibitor that mediates cell cycle arrest in response to pheromone; also forms a complex with Cdc24p, Ste4p, and Ste18p that may specify the direction of polarized growth during mating; potential Cdc28p substrate ‐0.9400 0.0003 RDS2 YPL133C Transcription factor involved in regulating gluconeogenesis and glyoxylate cycle genes; member of the zinc cluster family of proteins; confers resistance to ketoconazole ‐0.9391 0.0002 KAR9 YPL269W Karyogamy protein required for correct positioning of the mitotic spindle and for orienting cytoplasmic microtubules, localizes at the shmoo tip in mating cells and at the tip of the growing bud in small‐ budded cells through anaphase ‐0.9366 0.0002 FHL1 YPR104C Regulator of ribosomal protein transcription; has forkhead associated domain that binds phosphorylated proteins; also has forkhead DNA‐binding domain but does not bind DNA in vitro; suppresses RNA pol III and splicing factor prp4 mutants ‐0.9336 0.0004 Symbols ORF Description NA YNL108C NA YJR124C NA YHR131C TPK1 193 log2(mtr10. Ts/WT.TS) adj.p.v al Mitochondrial NADH kinase, phosphorylates NADH; also phosphorylates NAD(+) with lower specificity; required for the response to oxidative stress ‐0.9297 0.0002 YFL025C GPI inositol deacylase of the ER that negatively regulates COPII vesicle formation, prevents production of vesicles with defective subunits, required for proper discrimination between resident ER proteins and Golgi‐bound cargo molecules ‐0.9257 0.0001 ATG32 YIL146C Mitochondrial‐anchored transmembrane receptor that interacts with the autophagy adaptor protein, Atg11p, and is essential for mitophagy, the selective vacuolar degradation of mitochondria in response to starvation ‐0.9237 0.0002 PRY3 NA YJL078C YER130C Protein of unknown function Protein of unknown function; transcription is regulated by Haa1p, Sok2p and Zap1p transcriptional activators; the C. Albicans homolog (MNL1) plays a role in adaptation to stress ‐0.9213 ‐0.9211 0.0003 0.0002 NA YEL057C ‐0.9197 0.0005 ART5 YGR068C Protein of unknown function involved in telomere maintenance; target of UME6 regulation Protein proposed to regulate the endocytosis of plasma membrane proteins by recruiting the ubiquitin ligase Rsp5p to its target in the plasma membrane ‐0.9171 0.0002 PSF2 YJL072C Subunit of the GINS complex (Sld5p, Psf1p, Psf2p, Psf3p), which is localized to DNA replication origins and implicated in assembly of the DNA replication machinery ‐0.9164 0.0002 NA YPR147C Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm and is induced in response to the DNA‐damaging agent MMS ‐0.9160 0.0001 HMLALPHA 1 YCL066W Silenced copy of ALPHA1 at HML, encoding a transcriptional coactivator involved in the regulation of mating‐type alpha‐specific gene expression ‐0.9150 0.0003 TOS3 YGL179C Protein kinase, related to and functionally redundant with Elm1p and Sak1p for the phosphorylation and activation of Snf1p; functionally orthologous to LKB1, a mammalian kinase associated with Peutz‐ Jeghers cancer‐susceptibility syndrome ‐0.9136 0.0002 NA YOL024W ‐0.9128 0.0001 AGP2 YBR132C Putative protein of unknown function; predicted to have thiol‐ disulfide oxidoreductase active site High affinity polyamine permease, preferentially uses spermidine over putrescine; expression is down‐regulated by osmotic stress; plasma membrane carnitine transporter, also functions as a low‐ affinity amino acid permease ‐0.9121 0.0004 IRC23 YOR044W Putative protein of unknown function; green fluorescent protein (GFP)‐fusion localizes to the ER; null mutant displays increased levels of spontaneous Rad52p foci ‐0.9119 0.0001 NA YMR010W ‐0.9113 0.0002 YGK3 YOL128C Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm; YMR010W is not an essential gene; YMR010W mRNA is transcribed with ADI1 Protein kinase related to mammalian glycogen synthase kinases of the GSK‐3 family; GSK‐3 homologs (Mck1p, Rim11p, Mrk1p, Ygk3p) are involved in control of Msn2p‐dependent transcription of stress responsive genes and in protein degradation ‐0.9102 0.0003 PET127 YOR017W Protein with a role in 5'‐end processing of mitochondrial RNAs, located in the mitochondrial membrane ‐0.9086 0.0001 Symbols ORF Description POS5 YPL188W BST1 194 log2(mtr10. Ts/WT.TS) adj.p.v al Probable Hsp40p co‐chaperone, has a DnaJ‐like domain and appears to be involved in ER‐associated degradation of misfolded proteins containing a tightly folded cytoplasmic domain; inhibits replication of Brome mosaic virus in S. cerevisiae ‐0.9078 0.0001 YPR026W Acid trehalase required for utilization of extracellular trehalose ‐0.9075 0.0002 IST2 YBR086C Plasma membrane protein that may be involved in osmotolerance, localizes to the mother cell in small‐budded cells and to the bud in medium‐ and large‐budded cells; mRNA is transported to the bud tip by an actomyosin‐driven process ‐0.9068 0.0004 SCW11 YGL028C ‐0.9062 0.0002 NA YBR220C ‐0.9047 0.0001 NA YLR137W Cell wall protein with similarity to glucanases; may play a role in conjugation during mating based on its regulation by Ste12p Putative protein of unknown function; YBR220C is not an essential gene Putative S‐adenosylmethionine‐dependent methyltransferase ‐0.9044 0.0001 BNA1 YJR025C 3‐hydroxyanthranilic acid dioxygenase, required for the de novo biosynthesis of NAD from tryptophan via kynurenine; expression regulated by Hst1p ‐0.9031 0.0002 ORC5 YNL261W Subunit of the origin recognition complex, which directs DNA replication by binding to replication origins and is also involved in transcriptional silencing ‐0.9023 0.0002 GAL10 YBR019C UDP‐glucose‐4‐epimerase, catalyzes the interconversion of UDP‐ galactose and UDP‐D‐glucose in galactose metabolism; also catalyzes the conversion of alpha‐D‐glucose or alpha‐D‐galactose to their beta‐ anomers ‐0.9008 0.0006 NA YKR070W Putative protein of unknown function; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐ throughput studies ‐0.9008 0.0003 ECM3 YOR092W Non‐essential protein of unknown function; involved in signal transduction and the genotoxic response; induced rapidly in response to treatment with 8‐methoxypsoralen and UVA irradiation ‐0.8967 0.0001 USA1 YML029W Scaffold subunit of the Hrd1p ubiquitin ligase that also promotes ligase oligomerization; involved in ER‐associated protein degradation (ERAD); interacts with the U1 snRNP‐specific protein, Snp1p ‐0.8952 0.0002 NA YOR302W ‐0.8937 0.0006 NA YER140W CPA1 uORF, Arginine attenuator peptide, regulates translation of the CPA1 mRNA Putative protein of unknown function; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐ throughput studies; genetic interactions suggest a role in folding of ER membrane proteins ‐0.8931 0.0003 RPS7B YNL096C Protein component of the small (40S) ribosomal subunit, nearly identical to Rps7Ap; interacts with Kti11p; deletion causes hypersensitivity to zymocin; has similarity to rat S7 and Xenopus S8 ribosomal proteins ‐0.8891 0.0005 TNA1 YGR260W High affinity nicotinic acid plasma membrane permease, responsible for uptake of low levels of nicotinic acid; expression of the gene increases in the absence of extracellular nicotinic acid or para‐ aminobenzoate (PABA) ‐0.8880 0.0001 APL3 YBL037W Alpha‐adaptin, large subunit of the clathrin associated protein complex (AP‐2); involved in vesicle mediated transport ‐0.8878 0.0003 Symbols ORF Description JID1 YPR061C ATH1 195 log2(mtr10. Ts/WT.TS) adj.p.v al Plasma membrane ATP‐binding cassette (ABC) transporter, weak‐ acid‐inducible multidrug transporter required for weak organic acid resistance; induced by sorbate and benzoate and regulated by War1p; mutants exhibit sorbate hypersensitivity ‐0.8864 0.0001 YPR047W Mitochondrial phenylalanyl‐tRNA synthetase, active as a monomer, unlike the cytoplasmic subunit which is active as a dimer complexed to a beta subunit dimer; similar to the alpha subunit of E. coli phenylalanyl‐tRNA synthetase ‐0.8855 0.0001 NA YHR202W Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the vacuole, while HA‐tagged protein is found in the soluble fraction, suggesting cytoplasmic localization ‐0.8824 0.0002 BNA6 YFR047C Quinolinate phosphoribosyl transferase, required for the de novo biosynthesis of NAD from tryptophan via kynurenine; expression regulated by Hst1p ‐0.8819 0.0002 AMD1 YML035C AMP deaminase, tetrameric enzyme that catalyzes the deamination of AMP to form IMP and ammonia; may be involved in regulation of intracellular adenine nucleotide pools ‐0.8779 0.0001 TOR1 YJR066W PIK‐related protein kinase and rapamycin target; subunit of TORC1, a complex that controls growth in response to nutrients by regulating translation, transcription, ribosome biogenesis, nutrient transport and autophagy; involved in meiosis ‐0.8779 0.0003 TEL1 YBL088C Protein kinase primarily involved in telomere length regulation; contributes to cell cycle checkpoint control in response to DNA damage; functionally redundant with Mec1p; homolog of human ataxia telangiectasia (ATM) gene ‐0.8760 0.0001 NA MIG3 YBR074W YER028C Putative metalloprotease Probable transcriptional repressor involved in response to toxic agents such as hydroxyurea that inhibit ribonucleotide reductase; phosphorylation by Snf1p or the Mec1p pathway inactivates Mig3p, allowing induction of damage response genes ‐0.8758 ‐0.8750 0.0001 0.0006 PRP8 YHR165C Component of the U4/U6‐U5 snRNP complex, involved in the second catalytic step of splicing; mutations of human Prp8 cause retinitis pigmentosa ‐0.8737 0.0001 NA ATR1 YMR147W YML116W ‐0.8728 ‐0.8719 0.0003 0.0004 SPO21 YOL091W Putative protein of unknown function Multidrug efflux pump of the major facilitator superfamily, required for resistance to aminotriazole and 4‐nitroquinoline‐N‐oxide Component of the meiotic outer plaque of the spindle pole body, involved in modifying the meiotic outer plaque that is required prior to prospore membrane formation ‐0.8718 0.0009 MRF1 YGL143C Mitochondrial translation release factor, involved in stop codon recognition and hydrolysis of the peptidyl‐tRNA bond during mitochondrial translation; lack of MRF1 causes mitochondrial genome instability ‐0.8716 0.0002 NA YOR186W ‐0.8711 0.0005 RTC2 YBR147W Putative protein of unknown function; proper regulation of expression during heat stress is sphingolipid‐dependent Protein of unknown function; identified in a screen for mutants with decreased levels of rDNA transcription; detected in highly purified mitochondria; null mutant suppresses cdc13‐1; similar to a G‐protein coupled receptor from S. pombe ‐0.8693 0.0002 NSE5 YML023C Essential subunit of the Mms21‐Smc5‐Smc6 complex; required for cell viability and DNA repair ‐0.8640 0.0001 Symbols ORF Description PDR12 YPL058C MSF1 196 log2(mtr10. Ts/WT.TS) adj.p.v al Ribosomal protein 59 of the small subunit, required for ribosome assembly and 20S pre‐rRNA processing; mutations confer cryptopleurine resistance; nearly identical to Rps14Ap and similar to E. coli S11 and rat S14 ribosomal proteins ‐0.8635 0.0007 Middle sporulation‐specific mitogen‐activated protein kinase (MAPK) required for production of the outer spore wall layers; negatively regulates activity of the glucan synthase subunit Gsc2p Ubiquitin‐specific protease that cleaves ubiquitin from ubiquitinated proteins Protein with similarity to hect domain E3 ubiquitin‐protein ligases, not essential for viability Putative protein of unknown function; deletion mutant has increased glycogen accumulation and displays elongated buds; green fluorescent protein (GFP)‐fusion protein localizes to the nucleus Putative protein of unknown function with similarity to Pip2p, an oleate‐specific transcriptional activator of peroxisome proliferation; YLL054C is not an essential gene ‐0.8620 0.0001 ‐0.8615 0.0006 ‐0.8606 0.0006 ‐0.8586 0.0002 ‐0.8578 0.0006 Dipeptidyl aminopeptidase, Golgi integral membrane protein that cleaves on the carboxyl side of repeating ‐X‐Ala‐ sequences, required for maturation of alpha factor, transcription is induced by a‐factor Protein of unknown function, required for growth on glycerol as a carbon source; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐throughput studies Putative membrane glycoprotein with strong similarity to Vth2p and Pep1p/Vps10p, may be involved in vacuolar protein sorting Putative transcription factor containing a zinc finger; overexpression increases salt tolerance through increased expression of the ENA1 (Na+/Li+ extrusion pump) gene while gene disruption decreases both salt tolerance and ENA1 expression ‐0.8570 0.0001 ‐0.8554 0.0006 ‐0.8536 0.0001 ‐0.8528 0.0002 Putative basic leucine zipper (bZIP) transcription factor Vacuolar protein sorting protein that forms part of the multimeric membrane‐associated retromer complex along with Vps35p, Vps29p, Vps17p, and Vps5p; essential for endosome‐to‐Golgi retrograde protein transport ‐0.8501 ‐0.8499 0.0007 0.0006 YNR061C YPL134C Putative protein of unknown function Mitochondrial inner membrane transporter, exports 2‐oxoadipate and 2‐oxoglutarate from the mitochondrial matrix to the cytosol for lysine and glutamate biosynthesis and lysine catabolism; suppresses, in multicopy, an fmc1 null mutation ‐0.8476 ‐0.8435 0.0003 0.0002 NA YBL112C ‐0.8435 0.0006 PCI8 YIL071C Putative protein of unknown function; YBL112C is contained within TEL02L Possible shared subunit of Cop9 signalosome (CSN) and eIF3, binds eIF3b subunit Prt1p, has possible dual functions in transcriptional and translational control, contains a PCI (Proteasome‐COP9 signalosome (CSN)‐eIF3) domain ‐0.8421 0.0006 MSB2 YGR014W Mucin family member involved in the Cdc42p‐ and MAP kinase‐ dependent filamentous growth signaling pathway; also functions as an osmosensor in parallel to the Sho1p‐mediated pathway; potential Cdc28p substrate ‐0.8396 0.0002 DSE2 YHR143W Daughter cell‐specific secreted protein with similarity to glucanases, degrades cell wall from the daughter side causing daughter to separate from mother; expression is repressed by cAMP ‐0.8379 0.0003 Symbols ORF Description RPS14B YJL191W SMK1 YPR054W UBP11 YKR098C HUL4 YJR036C NA YGR071C NA YLL054C STE13 YOR219C GDS1 YOR355W VTH1 YIL173W HAL9 YOL089C YAP7 PEP8 YOL028C YJL053W NA ODC1 197 log2(mtr10. Ts/WT.TS) adj.p.v al Minor CTP synthase isozyme (see also URA7), catalyzes the ATP‐ dependent transfer of the amide nitrogen from glutamine to UTP, forming CTP, the final step in de novo biosynthesis of pyrimidines; involved in phospholipid biosynthesis ‐0.8373 0.0002 YLR251W Protein required for ethanol metabolism; induced by heat shock and localized to the inner mitochondrial membrane; homologous to mammalian peroxisomal membrane protein Mpv17 ‐0.8367 0.0008 GLG2 YJL137C ‐0.8360 0.0001 SPO73 YER046W Self‐glucosylating initiator of glycogen synthesis, also glucosylates n‐ dodecyl‐beta‐D‐maltoside; similar to mammalian glycogenin Meiosis‐specific protein of unknown function, required for spore wall formation during sporulation; dispensible for both nuclear divisions during meiosis ‐0.8351 0.0002 CLG1 YGL215W ‐0.8347 0.0006 NA YHR080C ‐0.8346 0.0009 GZF3 YJL110C GATA zinc finger protein and Dal80p homolog that negatively regulates nitrogen catabolic gene expression by competing with Gat1p for GATA site binding; function requires a repressive carbon source; dimerizes with Dal80p and binds to Tor1p ‐0.8323 0.0001 INP52 YNL106C Polyphosphatidylinositol phosphatase, dephosphorylates a number of phosphatidylinositols (PIs) to PI; involved in endocytosis; hyperosmotic stress causes translocation to actin patches; synaptojanin‐like protein with a Sac1 domain ‐0.8315 0.0007 RAD1 YPL022W Single‐stranded DNA endonuclease (with Rad10p), cleaves single‐ stranded DNA during nucleotide excision repair and double‐strand break repair; subunit of Nucleotide Excision Repair Factor 1 (NEF1); homolog of human XPF protein ‐0.8298 0.0003 YSC84 YHR016C ‐0.8286 0.0004 CTL1 YMR180C Actin‐binding protein involved in bundling of actin filaments and endocytosis of actin cortical patches; activity stimulated by Las17p; contains SH3 domain similar to Rvs167p RNA 5'‐triphosphatase, localizes to both the nucleus and cytoplasm ‐0.8259 0.0001 ATP2 YJR121W Beta subunit of the F1 sector of mitochondrial F1F0 ATP synthase, which is a large, evolutionarily conserved enzyme complex required for ATP synthesis; phosphorylated ‐0.8250 0.0002 PET111 YMR257C ‐0.8245 0.0002 DBF20 YPR111W Mitochondrial translational activator specific for the COX2 mRNA; located in the mitochondrial inner membrane Ser/Thr kinase involved in late nuclear division, one of the mitotic exit network (MEN) proteins; necessary for the execution of cytokinesis ‐0.8236 0.0008 ANT1 YPR128C Peroxisomal adenine nucleotide transporter; involved in beta‐ oxidation of medium‐chain fatty acid; required for peroxisome proliferation ‐0.8231 0.0001 AVT1 YJR001W ‐0.8216 0.0003 NA YCR023C ‐0.8214 0.0001 TIR4 YOR009W Vacuolar transporter, imports large neutral amino acids into the vacuole; member of a family of seven S. cerevisiae genes (AVT1‐7) related to vesicular GABA‐glycine transporters Vacuolar membrane protein of unknown function; member of the multidrug resistance family; YCR023C is not an essential gene Cell wall mannoprotein of the Srp1p/Tip1p family of serine‐alanine‐ rich proteins; expressed under anaerobic conditions and required for anaerobic growth; transcription is also induced by cold shock ‐0.8214 0.0003 Symbols ORF Description URA8 YJR103W SYM1 Cyclin‐like protein that interacts with Pho85p; has sequence similarity to G1 cyclins PCL1 and PCL2 Protein of unknown function that may interact with ribosomes, based on co‐purification experiments; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐ throughput studies 198 log2(mtr10. Ts/WT.TS) adj.p.v al ‐0.8212 0.0005 ‐0.8207 0.0003 ‐0.8206 0.0002 Putative protein of unknown function; identified in a screen for mutants with increased levels of rDNA transcription; green fluorescent protein (GFP)‐fusion protein localizes to lipid particles Kynureninase, required for the de novo biosynthesis of NAD from tryptophan via kynurenine; expression regulated by Hst1p Dual‐specificity protein phosphatase; exists in 2 isoforms; required for maintenance of a low level of signaling through the cell integrity pathway, adaptive response to pheromone; regulates and is regulated by Slt2p; dephosphorylates Fus3p ‐0.8196 0.0005 ‐0.8181 0.0005 ‐0.8162 0.0002 YBR077C Component of the EGO complex, which is involved in the regulation of microautophagy, and of the GSE complex, which is required for proper sorting of amino acid permease Gap1p; gene exhibits synthetic genetic interaction with MSS4 ‐0.8146 0.0002 NA HOL1 YMR265C YNR055C ‐0.8143 ‐0.8129 0.0001 0.0001 RMR1 YGL250W Putative protein of unknown function Putative transporter in the major facilitator superfamily (DHA1 family) of multidrug resistance transporters; mutations in membrane‐spanning domains permit cation and histidinol uptake Protein required for meiotic recombination and gene conversion; null mutant displays reduced PIS1 expression and growth defects on non‐fermentable carbon sources and minimal media; GFP‐fusion protein localizes to both cytoplasm and nucleus ‐0.8118 0.0002 RPS0B YLR048W Protein component of the small (40S) ribosomal subunit, nearly identical to Rps0Ap; required for maturation of 18S rRNA along with Rps0Ap; deletion of either RPS0 gene reduces growth rate, deletion of both genes is lethal ‐0.8110 0.0002 MNN2 YBR015C Alpha‐1,2‐mannosyltransferase, responsible for addition of the first alpha‐1,2‐linked mannose to form the branches on the mannan backbone of oligosaccharides, localizes to an early Golgi compartment ‐0.8106 0.0006 AKR2 YOR034C Ankyrin repeat‐containing protein similar to Akr1p; member of a family of putative palmitoyltransferases containing an Asp‐His‐His‐ Cys‐cysteine rich (DHHC‐CRD) domain; possibly involved in constitutive endocytosis of Ste3p ‐0.8073 0.0002 RIM15 YFL033C Glucose‐repressible protein kinase involved in signal transduction during cell proliferation in response to nutrients, specifically the establishment of stationary phase; identified as a regulator of IME2; substrate of Pho80p‐Pho85p kinase ‐0.8063 0.0004 FLO10 YKR102W ‐0.8044 0.0006 NA YPL067C Lectin‐like protein with similarity to Flo1p, thought to be involved in flocculation Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm; YPL067C is not an essential gene ‐0.8044 0.0003 YBP2 YGL060W ‐0.8033 0.0001 Symbols ORF Description ECM25 YJL201W NA YMR210W NA YGR266W Non‐essential protein of unknown function; promoter contains a consensus binding sequence for factor Abf1p Putative acyltransferase with similarity to Eeb1p and Eht1p, has a minor role in medium‐chain fatty acid ethyl ester biosynthesis; may be involved in lipid metabolism and detoxification Protein of unknown function, predicted to contain a single transmembrane domain; mutant has increased aneuploidy tolerance; localized to both the mitochondrial outer membrane and the plasma membrane RRT8 YOL048C BNA5 YLR231C MSG5 YNL053W SLM4 Central kinetochore associated protein that mediates mitotic progression; interacts with several central kinetochore proteins and the centromeric histone Cse4p; role in resistance to oxidative stress; similarity to Ybp1p and Slk19p 199 log2(mtr10. Ts/WT.TS) adj.p.v al Zinc‐finger transcription factor that regulates genes involved in branched chain amino acid biosynthesis and ammonia assimilation; positively regulated by alpha‐isopropylmalate, an intermediate in leucine biosynthesis ‐0.8030 0.0003 YPR005C Cytoplasmic protein involved in halotolerance; decreases intracellular Na+ (via Ena1p) and increases intracellular K+ by decreasing efflux; expression repressed by Ssn6p‐Tup1p and Sko1p and induced by NaCl, KCl, and sorbitol through Gcn4p ‐0.7989 0.0002 GPI10 YGL142C Integral membrane protein involved in glycosylphosphatidylinositol (GPI) anchor synthesis; putative alpha 1,2 mannosyltransferase required for addition of the third mannose onto the GPI core structure; human PIG‐Bp is a functional homolog ‐0.7988 0.0001 ECM38 YLR299W ‐0.7968 0.0006 TKL2 YBR117C Gamma‐glutamyltranspeptidase, major glutathione‐degrading enzyme; involved in detoxification of electrophilic xenobiotics; expression induced mainly by nitrogen starvation Transketolase, similar to Tkl1p; catalyzes conversion of xylulose‐5‐ phosphate and ribose‐5‐phosphate to sedoheptulose‐7‐phosphate and glyceraldehyde‐3‐phosphate in the pentose phosphate pathway; needed for synthesis of aromatic amino acids ‐0.7956 0.0009 GTO1 YGR154C ‐0.7953 0.0004 NA YFL054C ‐0.7943 0.0008 CLN3 YAL040C ‐0.7939 0.0001 EXO5 YBR163W Mitochondrial 5'‐3' exonuclease and sliding exonuclease, required for mitochondrial genome maintenance; distantly related to the RecB nuclease domain of bacterial RecBCD recombinases; may be regulated by the transcription factor Ace2 ‐0.7923 0.0002 DSS1 YMR287C 3'‐5' exoribonuclease, component of the mitochondrial degradosome along with the ATP‐dependent RNA helicase Suv3p; the degradosome associates with the ribosome and mediates turnover of aberrant or unprocessed RNAs ‐0.7906 0.0004 NAM7 YMR080C ATP‐dependent RNA helicase of the SFI superfamily involved in nonsense mediated mRNA decay; required for efficient translation termination at nonsense codons and targeting of NMD substrates to P‐bodies; involved in telomere maintenance ‐0.7897 0.0002 LAA1 YJL207C AP‐1 accessory protein; colocalizes with clathrin to the late‐Golgi apparatus; involved in TGN‐endosome transport; physically interacts with AP‐1; similar to the mammalian p200; may interact with ribosomes; YJL207C is a non‐essential gene ‐0.7891 0.0006 USE1 YGL098W Essential SNARE protein localized to the ER, involved in retrograde traffic from the Golgi to the ER; forms a complex with the SNAREs Sec22p, Sec20p and Ufe1p ‐0.7867 0.0003 NA CYC1 YGR016W YJR048W Putative protein of unknown function Cytochrome c, isoform 1; electron carrier of the mitochondrial intermembrane space that transfers electrons from ubiquinone‐ cytochrome c oxidoreductase to cytochrome c oxidase during cellular respiration ‐0.7861 ‐0.7861 0.0006 0.0002 Symbols ORF Description LEU3 YLR451W HAL1 Omega‐class glutathione transferase; induced under oxidative stress; putative peroxisomal localization Putative channel‐like protein; similar to Fps1p; mediates passive diffusion of glycerol in the presence of ethanol G1 cyclin involved in cell cycle progression; activates Cdc28p kinase to promote the G1 to S phase transition; plays a role in regulating transcription of the other G1 cyclins, CLN1 and CLN2; regulated by phosphorylation and proteolysis 200 log2(mtr10. Ts/WT.TS) adj.p.v al Mitochondrial intermediate peptidase, cleaves N‐terminal residues of a subset of proteins upon import, after their cleavage by mitochondrial processing peptidase (Mas1p‐Mas2p); may contribute to mitochondrial iron homeostasis ‐0.7851 0.0003 Protein component of the large (60S) ribosomal subunit, has similarity to Rpl6Ap and to rat L6 ribosomal protein; binds to 5.8S rRNA Transmembrane protein subunit of the glycosylphosphatidylinositol transamidase complex that adds GPIs to newly synthesized proteins; human PIG‐Tp homolog Catalytic subunit of the mitochondrial inner membrane i‐AAA protease complex, which is responsible for degradation of unfolded or misfolded mitochondrial gene products; mutation causes an elevated rate of mitochondrial turnover ‐0.7845 0.0003 ‐0.7839 0.0003 ‐0.7835 0.0006 YOR038C Subunit of the HIR complex, a nucleosome assembly complex involved in regulation of histone gene transcription; recruits Swi‐Snf complexes to histone gene promoters; promotes heterochromatic gene silencing with Asf1p ‐0.7819 0.0002 PAU2 YEL049W Member of the seripauperin multigene family encoded mainly in subtelomeric regions, active during alcoholic fermentation, regulated by anaerobiosis, negatively regulated by oxygen, repressed by heme ‐0.7817 0.0003 ERD1 YDR414C Predicted membrane protein required for the retention of lumenal endoplasmic reticulum proteins; mutants secrete the endogenous ER protein, BiP (Kar2p) ‐0.7807 0.0006 CAT2 YML042W ‐0.7803 0.0001 STE6 YKL209C Carnitine acetyl‐CoA transferase present in both mitochondria and peroxisomes, transfers activated acetyl groups to carnitine to form acetylcarnitine which can be shuttled across membranes Plasma membrane ATP‐binding cassette (ABC) transporter required for the export of a‐factor, catalyzes ATP hydrolysis coupled to a‐ factor transport; contains 12 transmembrane domains and two ATP binding domains; expressed only in MATa cells ‐0.7801 0.0006 GCY1 YOR120W ‐0.7769 0.0006 COG4 YPR105C ‐0.7747 0.0006 MEC1 YBR136W Putative NADP(+) coupled glycerol dehydrogenase, proposed to be involved in an alternative pathway for glycerol catabolism; also has mRNA binding activity; member of the aldo‐keto reductase (AKR) family Essential component of the conserved oligomeric Golgi complex (Cog1p through Cog8p), a cytosolic tethering complex that functions in protein trafficking to mediate fusion of transport vesicles to Golgi compartments Genome integrity checkpoint protein and PI kinase superfamily member; signal transducer required for cell cycle arrest and transcriptional responses prompted by damaged or unreplicated DNA; monitors and participates in meiotic recombination ‐0.7746 0.0002 RPL18A YOL120C ‐0.7744 0.0003 SHR5 YOL110W Protein component of the large (60S) ribosomal subunit, identical to Rpl18Bp and has similarity to rat L18 ribosomal protein; intron of RPL18A pre‐mRNA forms stem‐loop structures that are a target for Rnt1p cleavage leading to degradation Subunit of a palmitoyltransferase, composed of Shr5p and Erf2p, that adds a palmitoyl lipid moiety to heterolipidated substrates such as Ras1p and Ras2p through a thioester linkage; palmitoylation is required for Ras2p membrane localization ‐0.7740 0.0008 AGC1 YPR021C ‐0.7738 0.0002 Symbols ORF Description 1‐Oct YKL134C RPL6B YLR448W GPI16 YHR188C YME1 YPR024W HIR2 Mitochondrial amino acid transporter, acts both as a glutamate uniporter and as an aspartate‐glutamate exchanger; involved in nitrogen metabolism and nitrogen compound biosynthesis 201 log2(mtr10. Ts/WT.TS) adj.p.v al Membrane protein involved in zinc ion homeostasis, member of the four‐protein IZH family, expression induced by zinc deficiency; deletion reduces sensitivity to elevated zinc and shortens lag phase, overexpression reduces Zap1p activity ‐0.7738 0.0003 S‐adenosylmethionine transporter of the mitochondrial inner membrane, member of the mitochondrial carrier family; required for biotin biosynthesis and respiratory growth Putative protein of unknown function; weak sequence similarity to bis (5'‐nucleotidyl)‐tetraphosphatases; (GFP)‐fusion protein localizes to the vacuole; null mutant is highly sensitive to azaserine and resistant to sodium‐O‐vandate Glucosyl transferase, involved in N‐linked glycosylation; adds glucose to the dolichol‐linked oligosaccharide precursor prior to transfer to protein during lipid‐linked oligosaccharide biosynthesis; similar to Alg6p Protein required for normal prospore membrane formation; interacts with Gip1p, which is the meiosis‐specific regulatory subunit of the Glc7p protein phosphatase; expressed specifically in spores and localizes to the prospore membrane ‐0.7728 0.0003 ‐0.7702 0.0006 ‐0.7697 0.0002 ‐0.7684 0.0002 ‐0.7670 0.0001 ‐0.7666 0.0002 ‐0.7659 0.0009 Putative protein of unknown function, predicted to contain a PINc domain Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the endoplasmic reticulum; YNR021W is not an essential gene ‐0.7623 0.0007 ‐0.7622 0.0002 YHR078W YGR097W High osmolarity‐regulated gene of unknown function Component of RNA polymerase II holoenzyme, phosphorylated in response to oxidative stress; has a role in destruction of Ssn8p; proposed to function in activation of the glycerol channel Fps1p; paralogous to Rgc1p ‐0.7619 ‐0.7610 0.0006 0.0002 HIP1 YGR191W ‐0.7601 0.0008 RPL16B YNL069C ‐0.7586 0.0005 UBP8 YMR223W ‐0.7583 0.0005 ESBP6 YNL125C ‐0.7573 0.0006 CIN2 YPL241C High‐affinity histidine permease, also involved in the transport of manganese ions N‐terminally acetylated protein component of the large (60S) ribosomal subunit, binds to 5.8 S rRNA; has similarity to Rpl16Ap, E. coli L13 and rat L13a ribosomal proteins; transcriptionally regulated by Rap1p Ubiquitin‐specific protease that is a component of the SAGA (Spt‐ Ada‐Gcn5‐Acetyltransferase) acetylation complex; required for SAGA‐mediated deubiquitination of histone H2B Protein with similarity to monocarboxylate permeases, appears not to be involved in transport of monocarboxylates such as lactate, pyruvate or acetate across the plasma membrane GTPase‐activating protein (GAP) for Cin4p; tubulin folding factor C involved in beta‐tubulin (Tub2p) folding; mutants display increased chromosome loss and benomyl sensitivity; deletion complemented by human GAP, retinitis pigmentosa 2 ‐0.7545 0.0002 Symbols ORF Description IZH3 YLR023C PET8 YNL003C NA YNL217W ALG8 YOR067C YSW1 YBR148W MNN5 YJL186W SWD1 YAR003W STO1 YMR125W NA YIL151C NA YNR021W NA ASK10 Alpha‐1,2‐mannosyltransferase, responsible for addition of the second alpha‐1,2‐linked mannose of the branches on the mannan backbone of oligosaccharides, localizes to an early Golgi compartment Subunit of the COMPASS (Set1C) complex, which methylates histone H3 on lysine 4 and is required in transcriptional silencing near telomeres; WD40 beta propeller superfamily member with similarity to mammalian Rbbp7 Large subunit of the nuclear mRNA cap‐binding protein complex, interacts with Npl3p to carry nuclear poly(A)+ mRNA to cytoplasm; also involved in nuclear mRNA degradation and telomere maintenance; orthologous to mammalian CBP80 202 Symbols ORF Description UBP13 YBL067C NAB6 YML117W Putative ubiquitin carboxyl‐terminal hydrolase, ubiquitin‐specific protease that cleaves ubiquitin‐protein fusions Putative RNA‐binding protein that associates with mRNAs encoding cell wall proteins in high‐throughput studies; deletion mutants display increased sensitivity to some cell wall disrupting agents; expression negatively regulated by cAMP NA YGR127W ICE2 YIL090W KSS1 YGR040W TIR1 YER011W NTH2 YBR001C BER1 YLR412W PRM8 YGL053W KSP1 YHR082C GDT1 YBR187W NA YKL091C AIM22 YJL046W HOG1 YLR113W TRZ1 YKR079C MST1 FAR11 YKL194C YNL127W Putative protein of unknown function; expression is regulated by Msn2p/Msn4p, indicating a possible role in stress response Integral ER membrane protein with type‐III transmembrane domains; mutations cause defects in cortical ER morphology in both the mother and daughter cells Mitogen‐activated protein kinase (MAPK) involved in signal transduction pathways that control filamentous growth and pheromone response; the KSS1 gene is nonfunctional in S288C strains and functional in W303 strains Cell wall mannoprotein of the Srp1p/Tip1p family of serine‐alanine‐ rich proteins; expression is downregulated at acidic pH and induced by cold shock and anaerobiosis; abundance is increased in cells cultured without shaking Putative neutral trehalase, required for thermotolerance and may mediate resistance to other cellular stresses Protein involved in microtubule‐related processes, N‐acetylation; GFP‐fusion protein localizes to the cytoplasm and is induced in response to the DNA‐damaging agent MMS; YLR412W is not an essential gene; similar to Arabidopsis SRR1 gene Pheromone‐regulated protein with 2 predicted transmembrane segments and an FF sequence, a motif involved in COPII binding; forms a complex with Prp9p in the ER; member of DUP240 gene family Ser/thr protein kinase; nuclear translocation required for haploid filamentous growth; regulates filamentous growth induced nuclear translocation of Bcy1p, Fus3p, and Sks1p; overproduction causes allele‐specific suppression of prp20‐10 Putative protein of unknown function; expression is reduced in a gcr1 null mutant; GFP‐fusion protein localizes to the vacuole; expression pattern and physical interactions suggest a possible role in ribosome biogenesis Putative homolog of Sec14p, which is a phosphatidylinositol/phosphatidylcholine transfer protein involved in lipid metabolism; localizes to the nucleus Putative lipoate‐protein ligase, required along with Lip2 and Lip5 for lipoylation of Lat1p and Kgd2p; similar to E. coli LplA; null mutant displays reduced frequency of mitochondrial genome loss Mitogen‐activated protein kinase involved in osmoregulation via three independent osmosensors; mediates the recruitment and activation of RNA Pol II at Hot1p‐dependent promoters; localization regulated by Ptp2p and Ptp3p tRNA 3'‐end processing endonuclease tRNase Z; also localized to mitochondria and interacts genetically with Rex2 exonuclease; homolog of the human candidate prostate cancer susceptibility gene ELAC2 Mitochondrial threonyl‐tRNA synthetase Protein involved in recovery from cell cycle arrest in response to pheromone, in a Far1p‐independent pathway; interacts with Far3p, Far7p, Far8p, Far9p, and Far10p; has similarity to the N‐ and C‐ termini of N. crassa HAM‐2 203 log2(mtr10. Ts/WT.TS) adj.p.v al ‐0.7540 0.0004 ‐0.7519 0.0006 ‐0.7517 0.0007 ‐0.7515 0.0004 ‐0.7505 0.0005 ‐0.7499 0.0004 ‐0.7498 0.0002 ‐0.7491 0.0003 ‐0.7490 0.0001 ‐0.7458 0.0006 ‐0.7451 0.0001 ‐0.7450 0.0007 ‐0.7440 0.0002 ‐0.7433 0.0003 ‐0.7416 0.0003 ‐0.7414 ‐0.7404 0.0004 0.0007 log2(mtr10. Ts/WT.TS) adj.p.v al Glutamine‐dependent NAD(+) synthetase, essential for the formation of NAD(+) from nicotinic acid adenine dinucleotide Rab escort protein, forms a complex with the Ras‐like small GTPase Ypt1p that is required for the prenylation of Ypt1p by protein geranylgeranyltransferase type II (Bet2p‐Bet4p); sequence similarity to mammalian choroideraemia gene Sterol regulatory element binding protein, regulates transcription of sterol biosynthetic genes; contains Zn[2]‐Cys[6] binuclear cluster; homologous to Upc2p; relocates from intracellular membranes to perinuclear foci on sterol depletion Plasma membrane ATP‐binding cassette (ABC) transporter, multidrug transporter mediates export of many different organic anions including oligomycin; similar to human cystic fibrosis transmembrane receptor (CFTR) Putative protein of unknown function Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm; expressed during copper starvation; YOR296W is not an essential gene Protein involved in regulation of nonhomologous end joining; interacts with DNA ligase IV components Dnl4p and Lif1p; repressed by MAT heterozygosity; regulates cellular distribution of Lif1p Protein component of the large (60S) ribosomal subunit, nearly identical to Rpl12Bp; rpl12a rpl12b double mutant exhibits slow growth and slow translation; has similarity to E. coli L11 and rat L12 ribosomal proteins Vacuolar membrane protein involved in the ATP‐dependent transport of arginine into the vacuole and possibly in balancing ion homeostasis; homolog of human CLN3 involved in Batten disease (juvenile onset neuronal ceroid lipofuscinosis) ‐0.7401 0.0003 ‐0.7383 0.0001 ‐0.7383 0.0006 ‐0.7374 0.0002 ‐0.7338 ‐0.7302 0.0002 0.0004 ‐0.7282 0.0003 ‐0.7282 0.0008 ‐0.7275 0.0005 YFR014C Calmodulin‐dependent protein kinase; may play a role in stress response, many CA++/calmodulan dependent phosphorylation substrates demonstrated in vitro, amino acid sequence similar to Cmk2p and mammalian Cam Kinase II ‐0.7269 0.0006 AIM18 YHR198C Putative protein of unknown function; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐ throughput studies; null mutant displays elevated frequency of mitochondrial genome loss ‐0.7253 0.0010 UTP22 YGR090W Possible U3 snoRNP protein involved in maturation of pre‐18S rRNA, based on computational analysis of large‐scale protein‐protein interaction data ‐0.7240 0.0002 MAL31 YBR298C Maltose permease, high‐affinity maltose transporter (alpha‐ glucoside transporter); encoded in the MAL3 complex locus; member of the 12 transmembrane domain superfamily of sugar transporters; functional in genomic reference strain S288C ‐0.7239 0.0006 NCR1 YPL006W Vacuolar membrane protein that transits through the biosynthetic vacuolar protein sorting pathway, involved in sphingolipid metabolism; glycoprotein and functional orthologue of human Niemann Pick C1 (NPC1) protein ‐0.7234 0.0009 PEX2 YJL210W RING‐finger peroxin and E3 ubiquitin ligase, peroxisomal membrane protein with a C‐terminal zinc‐binding RING domain, forms translocation subcomplex with Pex10p and Pex12p which functions in peroxisomal matrix protein import ‐0.7187 0.0003 AXL1 YPR122W Haploid specific endoprotease that performs one of two N‐terminal cleavages during maturation of a‐factor mating pheromone; required for axial budding pattern of haploid cells ‐0.7183 0.0007 Symbols ORF Description QNS1 YHR074W MRS6 YOR370C ECM22 YLR228C YOR1 YGR281W NA NA YOR390W YOR296W NEJ1 YLR265C RPL12A YEL054C YHC3 YJL059W CMK1 204 log2(mtr10. Ts/WT.TS) adj.p.v al Non‐essential tetra‐spanning membrane protein found mostly in the late Golgi, can suppress some sed5 alleles; may be part of the transport machinery, but precise function is unknown; similar to mammalian syntaxin 5 ‐0.7170 0.0003 Nicotinic acid mononucleotide adenylyltransferase, involved in pathways of NAD biosynthesis, including the de novo, NAD(+) salvage, and nicotinamide riboside salvage pathways Mitochondrial protein of unknown function; homozygous diploid deletion strain has a sporulation defect characterized by elevated dityrosine in the soluble fraction; expression induced by calcium shortage; YJL147W is a non‐essential gene ‐0.7166 0.0001 ‐0.7149 0.0008 Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm; potential Cdc28p substrate ‐0.7148 0.0003 YIL036W Basic leucine zipper (bZIP) transcription factor of the ATF/CREB family, proposed to be a regulator of oleate responsive genes; involved in utilization of non‐optimal carbon sources and chromosome stability ‐0.7148 0.0010 RFT1 YBL020W Essential integral membrane protein that is required for translocation of Man5GlcNac2‐PP‐Dol from the cytoplasmic side to the lumenal side of the ER membrane but is not the flippase; mutation is suppressed by expression of human p53 protein ‐0.7110 0.0002 PAU5 YFL020C Member of the seripauperin multigene family encoded mainly in subtelomeric regions; induced during alcoholic fermentation; induced by low temperature and also by anaerobic conditions; negatively regulated by oxygen and repressed by heme ‐0.7083 0.0009 NA YHR035W Putative protein of unknown function; not an essential gene ‐0.7073 0.0003 STP22 YCL008C Component of the ESCRT‐I complex, which is involved in ubiquitin‐ dependent sorting of proteins into the endosome; homologous to the mouse and human Tsg101 tumor susceptibility gene; mutants exhibit a Class E Vps phenotype ‐0.7068 0.0004 NUP157 YER105C Abundant subunit of the nuclear pore complex (NPC), present on both sides of the NPC; has similarity to Nup170p; essential role, with Nup170p, in NPC assembly ‐0.7064 0.0004 BTS1 YPL069C Geranylgeranyl diphosphate synthase, increases the intracellular pool of geranylgeranyl diphosphate, suppressor of bet2 mutation that causes defective geranylgeranylation of small GTP‐binding proteins that mediate vesicular traffic ‐0.7058 0.0005 TOK1 YJL093C Outward‐rectifier potassium channel of the plasma membrane with two pore domains in tandem, each of which forms a functional channel permeable to potassium; carboxy tail functions to prevent inner gate closures; target of K1 toxin ‐0.7051 0.0004 PCL8 YPL219W Cyclin, interacts with Pho85p cyclin‐dependent kinase (Cdk) to phosphorylate and regulate glycogen synthase, also activates Pho85p for Glc8p phosphorylation ‐0.7047 0.0003 TUL1 YKL034W Golgi‐localized RING‐finger ubiquitin ligase (E3), involved in ubiquitinating and sorting membrane proteins that contain polar transmembrane domains to multivesicular bodies for delivery to the vacuole for quality control purposes ‐0.7047 0.0009 RSF2 YJR127C Zinc‐finger protein involved in transcriptional control of both nuclear and mitochondrial genes, many of which specify products required for glycerol‐based growth, respiration, and other functions ‐0.7046 0.0006 Symbols ORF Description SFT2 YBL102W NMA1 YLR328W NA YJL147C NA YHR159W CST6 205 log2(mtr10. Ts/WT.TS) adj.p.v al ‐0.7045 0.0004 ‐0.7041 0.0004 ‐0.7027 0.0004 ‐0.7024 0.0009 Putative protein of unknown function Putative protein of unknown function; GFP‐fusion protein localizes to the cytoplasm; conserved in closely related Saccharomyces species ‐0.7022 ‐0.7015 0.0006 0.0002 NA YER150W NA GPI‐anchored cell wall protein involved in weak acid resistance; basal expression requires Msn2p/Msn4p; expression is induced under conditions of stress and during the diauxic shift; similar to Sed1p ‐0.7014 ‐0.7005 0.0007 0.0009 MSN1 YOL116W Transcriptional activator involved in regulation of invertase and glucoamylase expression, invasive growth and pseudohyphal differentiation, iron uptake, chromium accumulation, and response to osmotic stress; localizes to the nucleus ‐0.6982 0.0003 NA YKL027W ‐0.6977 0.0007 ARG2 YJL071W Protein of unknown function, localized to the mitochondrial outer membrane Acetylglutamate synthase (glutamate N‐acetyltransferase), mitochondrial enzyme that catalyzes the first step in the biosynthesis of the arginine precursor ornithine; forms a complex with Arg5,6p ‐0.6976 0.0006 PHO91 YNR013C Low‐affinity phosphate transporter of the vacuolar membrane; deletion of pho84, pho87, pho89, pho90, and pho91 causes synthetic lethality; transcription independent of Pi and Pho4p activity; overexpression results in vigorous growth ‐0.6970 0.0002 NA YJR015W Putative protein of unknown function; localizes to the endoplasmic reticulum and cytoplasm; predicted to encode a membrane transporter based on phylogenetic analysis; YJR015W is a non‐ essential gene ‐0.6967 0.0002 SUR1 YPL057C Probable catalytic subunit of a mannosylinositol phosphorylceramide (MIPC) synthase, forms a complex with probable regulatory subunit Csg2p; function in sphingolipid biosynthesis is overlapping with that of Csh1p ‐0.6954 0.0007 TED1 YIL039W Conserved phosphoesterase domain‐containing protein that acts together with Emp24p/Erv25p in cargo exit from the ER; deletion confers sensitivity to 4‐(N‐(S‐glutathionylacetyl)amino) phenylarsenoxide (GSAO) ‐0.6950 0.0002 SAS5 YOR213C Subunit of the SAS complex (Sas2p, Sas4p, Sas5p), which acetylates free histones and nucleosomes and regulates transcriptional silencing; stimulates Sas2p HAT activity ‐0.6915 0.0008 NA KEG1 YKR051W YFR042W Putative protein of unknown function Integral membrane protein of the ER; physically interacts with Kre6p; has a role in the synthesis of beta‐1,6‐glucan in the cell wall; required for cell viability ‐0.6914 ‐0.6877 0.0003 0.0007 Symbols ORF Description COT1 YOR316C SGF11 YPL047W Vacuolar transporter that mediates zinc transport into the vacuole; overexpression confers resistance to cobalt and rhodium Integral subunit of SAGA histone acetyltransferase complex, regulates transcription of a subset of SAGA‐regulated genes, required for the Ubp8p association with SAGA and for H2B deubiquitylation NA YIL092W AMN1 YBR158W NA NA YKR005C YJL016W NA SPI1 Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm and to the nucleus Protein required for daughter cell separation, multiple mitotic checkpoints, and chromosome stability; contains 12 degenerate leucine‐rich repeat motifs; expression is induced by the Mitotic Exit Network (MEN) 206 log2(mtr10. Ts/WT.TS) adj.p.v al Putative sensor/transporter protein involved in cell wall biogenesis; contains 14‐16 transmembrane segments and several putative glycosylation and phosphorylation sites; null mutation is synthetically lethal with pkc1 deletion ‐0.6876 0.0006 Cell‐cycle regulated activator of anaphase‐promoting complex/cyclosome (APC/C), which is required for metaphase/anaphase transition; directs ubiquitination of mitotic cyclins, Pds1p, and other anaphase inhibitors; potential Cdc28p substrate Protein functioning in transport of glycosylphosphatidylinositol intermediates into ER lumen; required for normal intracellular sterol distribution; human ARV1 required for normal cholesterol and bile acid homeostasis; similar to Nup120p ‐0.6843 0.0001 ‐0.6839 0.0010 YNL238W Subtilisin‐like protease (proprotein convertase), a calcium‐ dependent serine protease involved in the activation of proproteins of the secretory pathway ‐0.6821 0.0003 NSG1 YHR133C Protein involved in regulation of sterol biosynthesis; specifically stabilizes Hmg2p, one of two HMG‐CoA isoenzymes that catalyze the rate‐limiting step in sterol biosynthesis; homolog of mammalian INSIG proteins ‐0.6797 0.0002 RIM2 YBR192W ‐0.6788 0.0003 POR2 YIL114C ‐0.6786 0.0001 EST3 YIL009C‐A ‐0.6782 0.0007 NA YPR127W Mitochondrial pyrimidine nucleotide transporter; imports pyrimidine nucleoside triphosphates and exports pyrimidine nucleoside monophosphates; member of the mitochondrial carrier family Putative mitochondrial porin (voltage‐dependent anion channel), related to Por1p but not required for mitochondrial membrane permeability or mitochondrial osmotic stability Component of the telomerase holoenzyme, involved in telomere replication Protein of unknown function, differentially expressed during alcoholic fermentation; expression activated by transcription factor YRM1/YOR172W; green fluorescent protein (GFP)‐fusion protein localizes to both the cytoplasm and the nucleus ‐0.6777 0.0005 ERG24 YNL280C C‐14 sterol reductase, acts in ergosterol biosynthesis; mutants accumulate the abnormal sterol ignosterol (ergosta‐8,14 dienol), and are viable under anaerobic growth conditions but inviable on rich medium under aerobic conditions ‐0.6772 0.0008 RML2 YEL050C Mitochondrial ribosomal protein of the large subunit, has similarity to E. coli L2 ribosomal protein; fat21 mutant allele causes inability to utilize oleate and may interfere with activity of the Adr1p transcription factor ‐0.6771 0.0006 CLB4 YLR210W B‐type cyclin involved in cell cycle progression; activates Cdc28p to promote the G2/M transition; may be involved in DNA replication and spindle assembly; accumulates during S phase and G2, then targeted for ubiquitin‐mediated degradation ‐0.6756 0.0003 LSB3 YFR024C‐A ‐0.6733 0.0002 YUR1 YJL139C ‐0.6709 0.0006 TYW3 YGL050W Protein containing a C‐terminal SH3 domain; binds Las17p, which is a homolog of human Wiskott‐Aldrich Syndrome protein involved in actin patch assembly and actin polymerization Mannosyltransferase of the KTR1 family, involved in protein N‐ glycosylation; located in the Golgi apparatus tRNA methyltransferase required for synthesis of wybutosine, a modified guanosine found at the 3'‐position adjacent to the anticodon of phenylalanine tRNA which supports reading frame maintenance by stabilizing codon‐anticodon interactions ‐0.6693 0.0002 Symbols ORF Description CWH43 YCR017C CDC20 YGL116W ARV1 YLR242C KEX2 207 log2(mtr10. Ts/WT.TS) adj.p.v al Mannosyltransferase, involved in asparagine‐linked glycosylation in the endoplasmic reticulum (ER); essential for viability, mutation is functionally complemented by human ortholog ‐0.6684 0.0005 YMR258C Protein of unknown function with similarity to F‐box proteins; physically interacts with Skp1p; green fluorescent protein (GFP)‐ fusion protein localizes to the cytoplasm and nucleus; YMR258C is not an essential gene ‐0.6683 0.0004 NA YCR015C ‐0.6653 0.0002 SYP1 YCR030C Putative protein of unknown function; YCR015C is not an essential gene Protein of unknown function that is involved in endocytic site formation; may regulate assembly and disassembly of the septin ring; colocalizes and interacts with septin subunits; potential role in actin cytoskeletal organization ‐0.6649 0.0006 NA YOR019W ‐0.6644 0.0003 PSY2 YNL201C Protein of unknown function that may interact with ribosomes, based on co‐purification experiments Putative subunit of an evolutionarily conserved protein phosphatase complex containing the catalytic subunit Pph3p and the regulatory subunit Psy4p; required for cisplatin and oxaliplatin resistance; putative homolog of mammalian R3 ‐0.6640 0.0005 NA YFR035C ‐0.6639 0.0009 ADH4 YGL256W Putative protein of unknown function, deletion mutant exhibits synthetic phenotype with alpha‐synuclein Alcohol dehydrogenase isoenzyme type IV, dimeric enzyme demonstrated to be zinc‐dependent despite sequence similarity to iron‐activated alcohol dehydrogenases; transcription is induced in response to zinc deficiency ‐0.6615 0.0004 PRR2 YDL214C Serine/threonine protein kinase that inhibits pheromone induced signalling downstream of MAPK, possibly at the level of the Ste12p transcription factor; mutant has increased aneuploidy tolerance ‐0.6613 0.0003 MKK1 YOR231W Mitogen‐activated kinase kinase involved in protein kinase C signaling pathway that controls cell integrity; upon activation by Bck1p phosphorylates downstream target, Slt2p; functionally redundant with Mkk2p ‐0.6594 0.0002 RGD2 MDM38 YFL047W YOL027C GTPase‐activating protein (RhoGAP) for Cdc42p and Rho5p Mitochondrial inner membrane protein, involved in membrane integration of a subset of mitochondrial proteins; required for K+/H+ exchange; associates with mitochondrial ribosomes; human ortholog Letm1 implicated in Wolf‐Hirschhorn syndrome ‐0.6585 ‐0.6572 0.0002 0.0005 VID28 YIL017C Protein involved in proteasome‐dependent catabolite degradation of fructose‐1,6‐bisphosphatase (FBPase); localized to the nucleus and the cytoplasm ‐0.6572 0.0006 AFI1 YOR129C Arf3p polarization‐specific docking factor, required for the polarized distribution of the ADP‐ribosylation factor, Arf3p; participates in polarity development and maintenance of a normal haploid budding pattern; interacts with Cnm7p ‐0.6569 0.0009 PGA1 YNL158W Essential component of GPI‐mannosyltransferase II, responsible for second mannose addition to GPI precursors as a partner of Gpi18p; required for maturation of Gas1p and Pho8p; has synthetic genetic interations with secretory pathway genes ‐0.6565 0.0009 COX15 YER141W Protein required for the hydroxylation of heme O to form heme A, which is an essential prosthetic group for cytochrome c oxidase ‐0.6562 0.0005 Symbols ORF Description ALG1 YBR110W NA 208 log2(mtr10. Ts/WT.TS) adj.p.v al Plasma membrane H+‐ATPase, pumps protons out of the cell; major regulator of cytoplasmic pH and plasma membrane potential; part of the P2 subgroup of cation‐transporting ATPases; Hsp30p plays a role in Pma1p regulation ‐0.6516 0.0003 Protein of unknown function, contains a J‐domain, which is a region with homology to the E. coli DnaJ protein Type II phosphatidylinositol 4‐kinase that binds Las17p, which is a homolog of human Wiskott‐Aldrich Syndrome protein involved in actin patch assembly and actin polymerization ‐0.6508 0.0007 ‐0.6499 0.0006 RNA binding protein involved in the asymmetric localization of ASH1 mRNA; represses translation of ASH1 mRNA, an effect reversed by Yck1p‐dependent phosphoryation; regulates telomere position effect and length; similarity to hnRNP‐K ‐0.6497 0.0003 YNR028W Peptidyl‐prolyl cis‐trans isomerase (cyclophilin), catalyzes the cis‐ trans isomerization of peptide bonds N‐terminal to proline residues; similarity to Cpr4p suggests a potential role in the secretory pathway ‐0.6473 0.0003 NA YLR177W Putative protein of unknown function; phosphorylated by Dbf2p‐ Mob1p in vitro; some strains contain microsatellite polymophisms at this locus; YLR177W is not an essential gene ‐0.6469 0.0008 WRS1 YOL097C ‐0.6458 0.0006 AIM19 YIL087C ‐0.6412 0.0004 ATG7 YHR171W Cytoplasmic tryptophanyl‐tRNA synthetase, aminoacylates tryptophanyl‐tRNA Putative protein of unknown function; the authentic, non‐tagged protein is detected in purified mitochondria in high‐throughput studies; null mutant displays reduced respiratory growth Autophagy‐related protein and dual specificity member of the E1 family of ubiquitin‐activating enzymes; mediates the conjugation of Atg12p with Atg5p and Atg8p with phosphatidylethanolamine, required steps in autophagosome formation ‐0.6401 0.0010 DOG1 YHR044C 2‐deoxyglucose‐6‐phosphate phosphatase, similar to Dog2p, member of a family of low molecular weight phosphatases; confers 2‐deoxyglucose resistance when overexpressed, in vivo substrate has not yet been identified ‐0.6398 0.0005 PDR17 YNL264C Phosphatidylinositol transfer protein (PITP), downregulates Plb1p‐ mediated turnover of phosphatidylcholine, found in the cytosol and microsomes, homologous to Pdr16p, deletion affects phospholipid composition ‐0.6393 0.0003 YDC1 YPL087W ‐0.6369 0.0007 RKR1 YMR247C Alkaline dihydroceramidase, involved in sphingolipid metabolism; preferentially hydrolyzes dihydroceramide to a free fatty acid and dihydrosphingosine; has a minor reverse activity RING domain E3 ubiquitin ligase; involved in the ubiquitin‐mediated degradation of non‐stop proteins; functional connections to chromatin modification; nuclear protein that also co‐localizes with ribosomes; homolog of mouse listerin ‐0.6334 0.0002 IRC20 YLR247C Putative helicase; localizes to the mitochondrion and the nucleus; YLR247C is not an essential gene; null mutant displays increased levels of spontaneous Rad52p foci ‐0.6333 0.0008 PTK1 YKL198C Putative serine/threonine protein kinase that regulates spermine uptake; involved in polyamine transport; possible mitochondrial protein ‐0.6332 0.0003 GCD7 YLR291C Beta subunit of the translation initiation factor eIF2B, the guanine‐ nucleotide exchange factor for eIF2; activity subsequently regulated by phosphorylated eIF2; first identified as a negative regulator of GCN4 expression ‐0.6312 0.0003 Symbols ORF Description PMA1 YGL008C JJJ2 YJL162C LSB6 YJL100W HEK2 YBL032W CPR8 209 log2(mtr10. Ts/WT.TS) adj.p.v al ‐0.6312 0.0006 ‐0.6311 ‐0.6284 0.0010 0.0007 Transcriptional activator of the basic leucine zipper (bZIP) family, required for transcription of genes involved in resistance to arsenic compounds ‐0.6266 0.0006 YBR023C Chitin synthase III, catalyzes the transfer of N‐acetylglucosamine (GlcNAc) to chitin; required for synthesis of the majority of cell wall chitin, the chitin ring during bud emergence, and spore wall chitosan ‐0.6262 0.0004 LDB16 YCL005W Protein of unknown function; null mutants have decreased net negative cell surface charge; GFP‐fusion protein expression is induced in response to the DNA‐damaging agent MMS; native protein is detected in purified mitochondria ‐0.6218 0.0002 RPS18B YML026C Protein component of the small (40S) ribosomal subunit; nearly identical to Rps18Ap and has similarity to E. coli S13 and rat S18 ribosomal proteins ‐0.6213 0.0002 HVG1 CPT1 YER039C YNL130C Protein of unknown function, has homology to Vrg4p Cholinephosphotransferase, required for phosphatidylcholine biosynthesis and for inositol‐dependent regulation of EPT1 transcription ‐0.6209 ‐0.6207 0.0003 0.0004 NA YBR235W Putative ion transporter, similar to mammalian electroneutral Na(+)‐ (K+)‐C1‐ cotransporter family; YBR235W is not an essential gene ‐0.6202 0.0006 YMD8 YML038C Putative nucleotide sugar transporter, has similarity to Vrg4p ‐0.6174 0.0004 DFG10 YIL049W Protein of unknown function, involved in filamentous growth ‐0.6172 0.0004 NA YHR045W Putative protein of unknown function; possible role in iron metabolism and/or amino acid and carbohydrate metabolism; green fluorescent protein (GFP)‐fusion protein localizes to the endoplasmic reticulum ‐0.6166 0.0004 YCK2 YNL154C Palmitoylated plasma membrane‐bound casein kinase I isoform; shares redundant functions with Yck1p in morphogenesis, proper septin assembly, endocytic trafficking; provides an essential function overlapping with that of Yck1p ‐0.6162 0.0006 CAN1 YEL063C ‐0.6149 0.0002 ODC2 YOR222W Plasma membrane arginine permease, requires phosphatidyl ethanolamine (PE) for localization, exclusively associated with lipid rafts; mutation confers canavanine resistance Mitochondrial inner membrane transporter, exports 2‐oxoadipate and 2‐oxoglutarate from the mitochondrial matrix to the cytosol for use in lysine and glutamate biosynthesis and in lysine catabolism ‐0.6146 0.0002 LEU5 YHR002W Mitochondrial carrier protein involved in the accumulation of CoA in the mitochondrial matrix; homolog of human Graves disease protein; does not encode an isozyme of Leu4p, as first hypothesized ‐0.6127 0.0004 NA CIR2 YOL098C YOR356W Putative metalloprotease Putative ortholog of human electron transfer flavoprotein dehydrogenase (ETF‐dH); found in a large supramolecular complex with other mitochondrial dehydrogenases; may have a role in oxidative stress response ‐0.6120 ‐0.6118 0.0004 0.0006 Symbols ORF Description NA YBR219C NA RPL17A NA YKL180W Putative protein of unknown function; YBR219C is not an essential gene NA Protein component of the large (60S) ribosomal subunit, nearly identical to Rpl17Bp and has similarity to E. coli L22 and rat L17 ribosomal proteins; copurifies with the Dam1 complex (aka DASH complex) ARR1 YPR199C CHS3 210 log2(mtr10. Ts/WT.TS) adj.p.v al Uncharacterized protein of unknown function Cell wall mannoprotein of the Srp1p/Tip1p family of serine‐alanine‐ rich proteins; expressed under anaerobic conditions and required for anaerobic growth ‐0.6094 ‐0.6073 0.0008 0.0003 YPL202C Iron‐regulated transcriptional activator; activates genes involved in intracellular iron use and required for iron homeostasis and resistance to oxidative stress; similar to Aft1p ‐0.6049 0.0004 MDL2 YPL270W Mitochondrial inner membrane half‐type ATP‐binding cassette (ABC) transporter, required for respiratory growth at high temperature; similar to human TAP1 and TAP2 implicated in bare lymphocyte syndrome and Wegener‐like granulomatosis ‐0.6019 0.0010 SCS3 YGL126W Protein required for inositol prototrophy, identified as an ortholog of the FIT family of proteins involved in triglyceride droplet biosynthesis; disputed role in the synthesis of inositol phospholipids from inositol ‐0.6013 0.0010 Symbols ORF Description NA TIR3 YIL067C YIL011W AFT2 211 APPENDIX J: Up-regulated genes in mtr10Δ cells in amino acid starved condition This section includes the list of transcriptionally differentially expressed genes (cutoff conditions: log2 FC 0.6 and adjusted p-value < 0.001) in mtr10Δ cells in amino acid starved condition. Up-regulated genes in mtr10Δ cells in amino acid starved condition. log2(mtr1 0.Ts/WT.T s) adj.p.val Subunit of the Cop9 signalosome, which is required for deneddylation, or removal of the ubiquitin‐like protein Rub1p from Cdc53p (cullin); involved in adaptation to pheromone signaling 3.1021 0.0000 Glycerol proton symporter of the plasma membrane, subject to glucose‐induced inactivation, strongly but transiently induced when cells are subjected to osmotic shock Putative membrane protein of unknown function Predicted tail‐anchored plasma membrane protein containing a conserved CYSTM module; related proteins in other organisms may be involved in response to stress; green fluorescent protein (GFP)‐ fusion protein localizes to the cell periphery 2.4358 0.0000 2.4040 2.3777 0.0000 0.0000 Cis‐golgi localized protein involved in ER to Golgi transport; forms a complex with the mammalian GRASP65 homolog, Grh1p; mutants are compromised for the fusion of ER‐derived vesicles with Golgi membranes 2.1003 0.0000 Dubious open reading frame unlikely to encode a protein, based on available experimental and comparative sequence data; partially overlaps the dubious ORF YBR056C‐B Putative protein of unknown function with similarity to acyl‐carrier‐ protein reductases; YDL114W is not an essential gene 2.0108 0.0001 2.0075 0.0001 1.9835 0.0000 Symbols ORF Description CSN9 YDR179C STL1 YDR536W NA NA YNR062C YDR034W‐B BUG1 YDL099W NA YBR056W‐A NA YDL114W SDC1 YDR469W Subunit of the COMPASS (Set1C) complex, which methylates lysine 4 of histone H3 and is required in chromatin silencing at telomeres; contains a Dpy‐30 domain that mediates interaction with Bre2p; similar to C. elegans and human DPY‐30 212 log2(mtr1 0.Ts/WT.T s) adj.p.val Subunit of a kinetochore‐microtubule binding complex with Spc105p that bridges centromeric heterochromatin and kinetochore MAPs and motors, and is also required for sister chromatid bi‐ orientation and kinetochore binding of SAC components 1.9616 0.0000 YDR515W RNA binding protein that associates with polysomes; proposed to be involved in regulating mRNA translation; involved in the copper‐ dependent mineralization of copper sulfide complexes on cell surface in cells cultured in copper salts 1.9587 0.0000 NA YFR012W‐A Putative protein of unknown function; identified by homology 1.9170 0.0000 DON1 YDR273W Meiosis‐specific component of the spindle pole body, part of the leading edge protein (LEP) coat, forms a ring‐like structure at the leading edge of the prospore membrane during meiosis II 1.8897 0.0000 NA CSM4 YDR514C YPL200W Putative protein of unknown function Protein required for accurate chromosome segregation during meiosis; involved in meiotic telomere clustering (bouquet formation) and telomere‐led rapid prophase movements 1.8877 1.8867 0.0000 0.0001 SRT1 YMR101C Cis‐prenyltransferase involved in synthesis of long‐chain dolichols (19‐22 isoprene units; as opposed to Rer2p which synthesizes shorter‐chain dolichols); localizes to lipid bodies; transcription is induced during stationary phase 1.8689 0.0000 NA NSE4 YAR064W YDL105W 1.8530 1.8367 0.0000 0.0001 TFB5 YDR079C‐A Putative protein of unknown function Nuclear protein that plays a role in the function of the Smc5p‐ Rhc18p complex Component of the RNA polymerase II general transcription and DNA repair factor TFIIH; involved in transcription initiation and in nucleotide‐excision repair; homolog of Chlamydomonas reinhardtii REX1‐S protein involved in DNA repair 1.8357 0.0000 DAD4 YDR320C‐A Essential subunit of the Dam1 complex (aka DASH complex), couples kinetochores to the force produced by MT depolymerization thereby aiding in chromosome segregation; is transferred to the kinetochore prior to mitosis 1.8149 0.0000 PEX19 YDL065C Chaperone and import receptor for newly‐synthesized class I peroxisomal membrane proteins (PMPs), binds PMPs in the cytoplasm and delivers them to the peroxisome for subsequent insertion into the peroxisomal membrane 1.7924 0.0000 NOP6 YDL213C rRNA‐binding protein required for 40S ribosomal subunit biogenesis; contains an RNA recognition motif (RRM) and has similarity to hydrophilins; NOP6 may be a fungal‐specific gene as no homologs have been yet identified in higher eukaryotes 1.7819 0.0000 SNN1 YNL086W Putative protein of unknown function; likely member of BLOC complex involved in endosomal cargo sorting; green fluorescent protein (GFP)‐fusion protein localizes to endosomes 1.7769 0.0000 MSH6 YDR097C Protein required for mismatch repair in mitosis and meiosis, forms a complex with Msh2p to repair both single‐base & insertion‐deletion mispairs; potentially phosphorylated by Cdc28p 1.7721 0.0000 NA NA YDR374C YBR219C Putative protein of unknown function Putative protein of unknown function; YBR219C is not an essential gene 1.7619 1.7564 0.0000 0.0001 Symbols ORF Description KRE28 YDR532C SLF1 213 log2(mtr1 0.Ts/WT.T s) adj.p.val Putative protein of unknown function; mutant in a srs2 mutant background displays MMS hypersensitivity; ortholog of human MHF2, a component of the Fanconi anemia (FA) complex that is involved in maintaining genome stability 1.7098 0.0000 YDR412W Component of the pre‐60S pre‐ribosomal particle; required for cell viability under standard (aerobic) conditions but not under anaerobic conditions; exonuclease required for 5′ end processing of pre‐60S ribosomal RNA 1.7033 0.0001 RAD34 YDR314C 1.6958 0.0000 TAF10 YDR167W Protein involved in nucleotide excision repair (NER); homologous to RAD4 Subunit (145 kDa) of TFIID and SAGA complexes, involved in RNA polymerase II transcription initiation and in chromatin modification 1.6930 0.0000 PES4 YFR023W 1.6841 0.0000 JAC1 YGL018C 1.6789 0.0000 ZIP1 YDR285W Transverse filament protein of the synaptonemal complex; required for normal levels of meiotic recombination and pairing between homologous chromosome during meiosis; potential Cdc28p substrate 1.6739 0.0000 DYN2 YDR424C Cytoplasmic light chain dynein, microtubule motor protein; proposed to be involved in the assembly of the nuclear pore complex 1.6724 0.0000 HXT9 YJL219W Putative hexose transporter that is nearly identical to Hxt11p, has similarity to major facilitator superfamily (MFS) transporters, expression of HXT9 is regulated by transcription factors Pdr1p and Pdr3p 1.6722 0.0001 NA YOR072W‐B 1.6602 0.0006 RAD28 YDR030C Putative protein of unknown function; identified by expression profiling and mass spectrometry Protein involved in DNA repair, related to the human CSA protein that is involved in transcription‐coupled repair nucleotide excision repair 1.6540 0.0000 CWC15 YDR163W Non‐essential protein involved in pre‐mRNA splicing, component of a complex containing Cef1p; has similarity to S. pombe Cwf15p 1.6455 0.0000 MGT1 YDL200C DNA repair methyltransferase (6‐O‐methylguanine‐DNA methylase) involved in protection against DNA alkylation damage 1.6327 0.0000 TMA23 YMR269W Nucleolar protein of unknown function implicated in ribosome biogenesis; TMA23 may be a fungal‐specific gene as no homologs have been yet identified in higher eukaryotes 1.6318 0.0006 URM1 YIL008W Ubiquitin‐like protein involved in thiolation of cytoplasmic tRNAs; receives sulfur from the E1‐like enzyme Uba4p and transfers it to tRNA; also functions as a protein tag with roles in nutrient sensing and oxidative stress response 1.6288 0.0001 RPT2 YDL007W One of six ATPases of the 19S regulatory particle of the 26S proteasome involved in the degradation of ubiquitinated substrates; required for normal peptide hydrolysis by the core 20S particle 1.6213 0.0000 Symbols ORF Description NA YDL160C‐A RRP17 Poly(A) binding protein, suppressor of DNA polymerase epsilon mutation, similar to Mip6p Specialized J‐protein that functions with Hsp70 in Fe‐S cluster biogenesis in mitochondria, involved in iron metabolism; contains a J domain typical to J‐type chaperones; localizes to the mitochondrial matrix 214 log2(mtr1 0.Ts/WT.T s) adj.p.val Hsp90 chaperone required for pheromone signaling and negative regulation of Hsf1p; docks with Tom70p for mitochondrial preprotein delivery; promotes telomerase DNA binding and nucleotide addition; interacts with Cns1p, Cpr6p, Cpr7p, Sti1p 1.6164 0.0002 YGR146C‐A YNL210W Putative protein of unknown function Protein with RNA‐binding motifs required for meiosis‐specific mRNA splicing; required for chromosome pairing and meiotic recombination 1.6157 1.6117 0.0000 0.0000 FAP7 YDL166C Essential NTPase required for small ribosome subunit synthesis, mediates processing of the 20S pre‐rRNA at site D in the cytoplasm but associates only transiently with 43S preribosomes via Rps14p, may be the endonuclease for site D 1.5956 0.0001 YPS3 YLR121C Aspartic protease, member of the yapsin family of proteases involved in cell wall growth and maintenance; attached to the plasma membrane via a glycosylphosphatidylinositol (GPI) anchor 1.5860 0.0002 CNL1 YDR357C Protein of unknown function; likely member of BLOC complex involved in endosomal cargo sorting; null mutant is sensitive to drug inducing secretion of vacuolar cargo; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm 1.5805 0.0001 MBP1 YDL056W Transcription factor involved in regulation of cell cycle progression from G1 to S phase, forms a complex with Swi6p that binds to MluI cell cycle box regulatory element in promoters of DNA synthesis genes 1.5753 0.0001 SAS10 YDL153C Essential subunit of U3‐containing Small Subunit (SSU) processome complex involved in production of 18S rRNA and assembly of small ribosomal subunit; disrupts silencing when overproduced; mutant has increased aneuploidy tolerance 1.5740 0.0007 CAR1 YPL111W 1.5642 0.0001 RPC53 ESF1 YDL150W YDR365C 1.5616 1.5547 0.0003 0.0001 NA YDL177C 1.5505 0.0000 VPS64 YDR200C Arginase, responsible for arginine degradation, expression responds to both induction by arginine and nitrogen catabolite repression; disruption enhances freeze tolerance RNA polymerase III subunit C53 Nucleolar protein involved in pre‐rRNA processing; depletion causes severely decreased 18S rRNA levels Putative protein of unknown function; similar to the mouse IMPACT gene; YDL177C is not an essential gene Protein required for cytoplasm to vacuole targeting of proteins; forms a complex with Far3p and Far7p to Far11p involved in recovery from pheromone‐induced cell cycle arrest; mutant has increased aneuploidy tolerance 1.5494 0.0002 NA YLR154C‐H 1.5435 0.0003 SEM1 YDR363W‐A Putative protein of unknown function identified by fungal homology comparisons and RT‐PCR; this ORF partially overlaps RND5‐2 Component of the lid subcomplex of the regulatory subunit of the 26S proteasome; involved in mRNA export mediated by the TREX‐2 complex (Sac3p‐Thp1p); ortholog of human DSS1 1.5263 0.0000 SSA2 YLL024C ATP binding protein involved in protein folding and vacuolar import of proteins; member of heat shock protein 70 (HSP70) family; associated with the chaperonin‐containing T‐complex; present in the cytoplasm, vacuolar membrane and cell wall 1.5248 0.0001 RAD9 YDR217C DNA damage‐dependent checkpoint protein, required for cell‐cycle arrest in G1/S, intra‐S, and G2/M; transmits checkpoint signal by activating Rad53p and Chk1p; hyperphosphorylated by Mec1p and Tel1p; potential Cdc28p substrate 1.5213 0.0001 Symbols ORF Description HSP82 YPL240C NA MER1 215 log2(mtr1 0.Ts/WT.T s) adj.p.val Component of the EKC/KEOPS protein complex with Kae1p, Gon7p, Bud32p, and Cgi121p; EKC/KEOPS complex is required for t6A tRNA modification and may have roles in telomere maintenance and transcription Nucleolar protein involved in rRNA processing, pre‐rRNA cleavage at site A2; also involved in telomere maintenance; mutation is synthetically lethal with a gar1 mutation 1.5068 0.0000 1.5057 0.0007 Nucleolar protein required for maturation of 18S rRNA, member of the eIF4A subfamily of DEAD‐box ATP‐dependent RNA helicases Putative protein of unknown function Protein involved in RNA splicing by the spliceosome; component of a complex containing Cef1p; interacts genetically with ISY1 and BUD13; may bind RNA; has similarity to S. pombe Cwf21p 1.5047 0.0006 1.5011 1.4971 0.0000 0.0000 YCL069W YDL104C Permease of basic amino acids in the vacuolar membrane Highly conserved mitochondrial protein, essential for t6A modification of mitochondrial tRNAs that decode ANN codons; similar to Kae1p and E. coli YgjD, both of which are also required for tRNA t6A modification 1.4959 1.4790 0.0001 0.0001 NA YCR108C 1.4760 0.0000 SMD3 YLR147C 1.4698 0.0000 YOS9 YDR057W Putative protein of unknown function; identified by fungal homology and RT‐PCR Core Sm protein Sm D3; part of heteroheptameric complex (with Smb1p, Smd1p, Smd2p, Sme1p, Smx3p, and Smx2p) that is part of the spliceosomal U1, U2, U4, and U5 snRNPs; homolog of human Sm D3 ER quality‐control lectin; integral subunit of the HRD ligase; binds to glycans with terminal alpha‐1,6 linked mannose on misfolded N‐ glycosylated proteins and participates in targeting proteins to ERAD; member of the OS‐9 protein family 1.4680 0.0001 BUR6 YER159C Subunit of a heterodimeric NC2 transcription regulator complex with Ncb2p; complex binds to TBP and can repress transcription by preventing preinitiation complex assembly or stimulate activated transcription; homologous to human NC2alpha 1.4676 0.0000 NA YDR379C‐A Protein involved in the assembly of the mitochondrial succinate dehydrogenase complex; mutations in human ortholog SDHAF1 are associated with infantile leukoencephalopathy 1.4671 0.0002 NA YFL067W Protein of unknown function, down‐regulated at low calcium levels 1.4639 0.0000 BTN2 YGR142W v‐SNARE binding protein that facilitates specific protein retrieval from a late endosome to the Golgi; modulates arginine uptake, possible role in mediating pH homeostasis between the vacuole and plasma membrane H(+)‐ATPase 1.4614 0.0001 GIR2 YDR152W Highly‐acidic cytoplasmic RWD domain‐containing protein of unknown function; forms a complex with Rbg2p; interacts with Rbg1p and Gcn1p; associates with translating ribosomes; putative intrinsically unstructured protein 1.4579 0.0000 SSA1 YAL005C ATPase involved in protein folding and nuclear localization signal (NLS)‐directed nuclear transport; member of heat shock protein 70 (HSP70) family; forms a chaperone complex with Ydj1p; localized to the nucleus, cytoplasm, and cell wall 1.4578 0.0001 PDS1 YDR113C Securin, inhibits anaphase by binding separin Esp1p; blocks cyclin destruction and mitotic exit, essential for meiotic progression and mitotic cell cycle arrest; localization is cell‐cycle dependent and regulated by Cdc28p phosphorylation 1.4565 0.0000 Symbols ORF Description PCC1 YKR095W‐A RRP8 YDR083W FAL1 YDR021W NA CWC21 YLR361C‐A YDR482C VBA3 QRI7 216 log2(mtr1 0.Ts/WT.T s) adj.p.val Putative protein of unknown function; non‐essential gene; proposed function in rRNA and ribosome biosynthesis based on transcriptional co‐regulation; genetic interactions suggest a role in ER‐associated protein degradation (ERAD) 1.4502 0.0005 YLR194C Structural constituent of the cell wall attached to the plasma membrane by a GPI‐anchor; expression is upregulated in response to cell wall stress 1.4448 0.0000 NA YGR226C Dubious open reading frame, unlikely to encode a protein; not conserved in closely related Saccharomyces species; overlaps significantly with a verified ORF, AMA1/YGR225W 1.4351 0.0000 RNH202 YDR279W 1.4311 0.0000 NA YDL157C Ribonuclease H2 subunit, required for RNase H2 activity; related to human AGS2 that causes Aicardi‐Goutieres syndrome Putative protein of unknown function; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐ throughput studies 1.4202 0.0000 NA YLR363W‐A 1.4082 0.0003 DAD1 YDR016C Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the nucleus Essential subunit of the Dam1 complex (aka DASH complex), couples kinetochores to the force produced by MT depolymerization thereby aiding in chromosome segregation; is transferred to the kinetochore prior to mitosis 1.4068 0.0000 PET100 YDR079W Chaperone that specifically facilitates the assembly of cytochrome c oxidase, integral to the mitochondrial inner membrane; interacts with a subcomplex of subunits VII, VIIa, and VIII (Cox7p, Cox9p, and Cox8p) but not with the holoenzyme 1.4037 0.0000 SPC19 YDR201W Essential subunit of the Dam1 complex (aka DASH complex), couples kinetochores to the force produced by MT depolymerization thereby aiding in chromosome segregation; also localized to nuclear side of spindle pole body 1.3975 0.0000 VPS60 YDR486C Cytoplasmic and vacuolar membrane protein involved in late endosome to vacuole transport; required for normal filament maturation during pseudohyphal growth; may function in targeting cargo proteins for degradation; interacts with Vta1p 1.3818 0.0000 NA YOL155W‐A 1.3807 0.0000 RTT103 YDR289C Putative protein of unknown function; identified by expression profiling and mass spectrometry Protein that interacts with exonuclease Rat1p and Rai1p and plays a role in transcription termination by RNA polymerase II, has an RPR domain (carboxy‐terminal domain interacting domain); also involved in regulation of Ty1 transposition 1.3805 0.0000 NA YHR214C‐E Putative protein of unknown function; identified by gene‐trapping, microarray‐based expression analysis, and genome‐wide homology searching 1.3786 0.0000 NA YLR154W‐E 1.3693 0.0001 IRC18 YJL037W Dubious open reading frame unlikely to encode a protein; encoded within the the 35S rRNA gene on the opposite strand Putative protein of unknown function; expression induced in respiratory‐deficient cells and in carbon‐limited chemostat cultures; similar to adjacent ORF, YJL038C; null mutant displays increased levels of spontaneous Rad52p foci 1.3632 0.0002 NA EMI1 YDR132C YDR512C Putative protein of unknown function Non‐essential protein required for transcriptional induction of the early meiotic‐specific transcription factor IME1, also required for sporulation; contains twin cysteine‐x9‐cysteine motifs 1.3617 1.3548 0.0000 0.0001 Symbols ORF Description NA YDR161W NA 217 log2(mtr1 0.Ts/WT.T s) adj.p.val Essential protein required for the DNA integrity checkpoint pathways; interacts physically with Mec1p; putative homolog of S. pombe Rad26 and human ATRIP 1.3532 0.0002 Mitochondrial ribosomal protein of the small subunit; contains twin cysteine‐x9‐cysteine motifs Putative ATP‐dependent RNA helicase of the DEAD‐box protein family, constituent of 66S pre‐ribosomal particles; essential protein involved in ribosome biogenesis 1.3525 0.0000 1.3512 0.0001 tRNA‐specific 2‐thiouridylase, responsible for 2‐thiolation of the wobble base of mitochondrial tRNAs; human ortholog is implicated in myoclonus epilepsy associated with ragged red fibers (MERRF) 1.3455 0.0001 YDR088C RNA splicing factor, required for ATP‐independent portion of 2nd catalytic step of spliceosomal RNA splicing; interacts with Prp18p; contains zinc knuckle domain 1.3448 0.0001 FYV7 YLR068W 1.3426 0.0009 NA YDL233W Essential protein required for maturation of 18S rRNA; required for survival upon exposure to K1 killer toxin Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm and nucleus; YDL233W is not an essential gene 1.3378 0.0000 SPR3 YGR059W Sporulation‐specific homolog of the yeast CDC3/10/11/12 family of bud neck microfilament genes; septin protein involved in sporulation; regulated by ABFI 1.3268 0.0004 HUB1 YNR032C‐A Ubiquitin‐like protein modifier, may function in modification of Sph1p and Hbt1p, functionally complemented by the human or S. pombe ortholog; mechanism of Hub1p adduct formation not yet clear 1.3228 0.0002 SHS1 YDL225W One of five related septins (Cdc3p, Cdc10p, Cdc11p, Cdc12p, Shs1p) that form a cortical filamentous collar at the mother‐bud neck which is necessary for normal morphogenesis and cytokinesis 1.3215 0.0002 QRI1 YDL103C UDP‐N‐acetylglucosamine pyrophosphorylase, catalyzes the formation of UDP‐N‐acetylglucosamine (UDP‐GlcNAc), which is important in cell wall biosynthesis, protein N‐glycosylation, and GPI anchor biosynthesis 1.3194 0.0000 MCD1 YDL003W Essential subunit of the cohesin complex required for sister chromatid cohesion in mitosis and meiosis; apoptosis induces cleavage and translocation of a C‐terminal fragment to mitochondria; expression peaks in S phase 1.3111 0.0001 SSP1 YHR184W Protein involved in the control of meiotic nuclear division and coordination of meiosis with spore formation; transcription is induced midway through meiosis 1.3097 0.0000 PAR32 YDL173W Putative protein of unknown function; hyperphosphorylated upon rapamycin treatment in a Tap42p‐dependent manner; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm; PAR32 is not an essential gene 1.3079 0.0000 HNT2 YDR305C Dinucleoside triphosphate hydrolase; has similarity to the tumor suppressor FHIT and belongs to the histidine triad (HIT) superfamily of nucleotide‐binding proteins 1.3061 0.0000 MTC3 YGL226W Protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the mitochondrion; mtc3 is synthetically sick with cdc13‐1 1.3041 0.0000 Symbols ORF Description LCD1 YDR499W MRP10 YDL045W‐A DBP10 YDL031W SLM3 YDL033C SLU7 218 log2(mtr1 0.Ts/WT.T s) adj.p.val Core Sm protein Sm E; part of heteroheptameric complex (with Smb1p, Smd1p, Smd2p, Smd3p, Smx3p, and Smx2p) that is part of the spliceosomal U1, U2, U4, and U5 snRNPs; homolog of human Sm E 1.3010 0.0000 YJR022W Lsm (Like Sm) protein; forms heteroheptameric complex (with Lsm2p, Lsm3p, Lsm4p, Lsm5p, Lsm6p, and Lsm7p) that is part of spliceosomal U6 snRNP and is also implicated in processing of pre‐ tRNA, pre‐snoRNA, and pre‐rRNA 1.2949 0.0001 TRS23 YDR246W One of 10 subunits of the transport protein particle (TRAPP) complex of the cis‐Golgi which mediates vesicle docking and fusion; involved in endoplasmic reticulum (ER) to Golgi membrane traffic; human homolog is TRAPPC4 1.2870 0.0000 PLP1 YDR183W Protein that interacts with CCT (chaperonin containing TCP‐1) complex and has a role in actin and tubulin folding; has weak similarity to phosducins, which are G‐protein regulators 1.2869 0.0000 NPL3 YDR432W RNA‐binding protein that promotes elongation, regulates termination, and carries poly(A) mRNA from nucleus to cytoplasm; required for pre‐mRNA splicing; dissociation from mRNAs promoted by Mtr10p; phosphorylated by Sky1p in the cytoplasm 1.2866 0.0000 OMS1 YDR316W Protein integral to the mitochondrial membrane; has a conserved methyltransferase motif; multicopy suppressor of respiratory defects caused by OXA1 mutations 1.2856 0.0002 EAF7 YNL136W Subunit of the NuA4 histone acetyltransferase complex, which acetylates the N‐terminal tails of histones H4 and H2A 1.2851 0.0000 NA YNR064C 1.2835 0.0000 MKC7 YDR144C Epoxide hydrolase, member of the alpha/beta hydrolase fold family; may have a role in detoxification of epoxides GPI‐anchored aspartyl protease, member of the yapsin family of proteases involved in cell wall growth and maintenance; shares functions with Yap3p and Kex2p 1.2790 0.0001 CWC2 YDL209C Member of the NineTeen Complex (NTC) that contains Prp19p and stabilizes U6 snRNA in catalytic forms of the spliceosome containing U2, U5, and U6 snRNAs; binds directly to U6 snRNA; similar to S. pombe Cwf2 1.2785 0.0000 BDF2 YDL070W Protein involved in transcription initiation at TATA‐containing promoters; associates with the basal transcription factor TFIID; contains two bromodomains; corresponds to the C‐terminal region of mammalian TAF1; redundant with Bdf1p 1.2756 0.0002 ERV2 YPR037C Flavin‐linked sulfhydryl oxidase localized to the endoplasmic reticulum lumen, involved in disulfide bond formation within the ER 1.2755 0.0000 DPB4 YDR121W Shared subunit of DNA polymerase (II) epsilon and of ISW2/yCHRAC chromatin accessibility complex; involved in both chromosomal DNA replication and in inheritance of telomeric silencing 1.2686 0.0001 CDC37 YDR168W Essential Hsp90p co‐chaperone; necessary for passage through the START phase of the cell cycle; stabilizes protein kinase nascent chains and participates along with Hsp90p in their folding 1.2674 0.0001 NA YFL065C Putative protein of unknown function; induced by treatment with 8‐ methoxypsoralen and UVA irradiation 1.2670 0.0006 Symbols ORF Description SME1 YOR159C LSM8 219 log2(mtr1 0.Ts/WT.T s) adj.p.val Homocitrate synthase isozyme, catalyzes the condensation of acetyl‐CoA and alpha‐ketoglutarate to form homocitrate, which is the first step in the lysine biosynthesis pathway; highly similar to the other isozyme, Lys20p 1.2639 0.0003 YDR123C Component of the heteromeric Ino2p/Ino4p basic helix‐loop‐helix transcription activator that binds inositol/choline‐responsive elements (ICREs), required for derepression of phospholipid biosynthetic genes in response to inositol depletion 1.2629 0.0002 NA GAL3 YDR506C YDR009W Possible membrane‐localized protein Transcriptional regulator involved in activation of the GAL genes in response to galactose; forms a complex with Gal80p to relieve Gal80p inhibition of Gal4p; binds galactose and ATP but does not have galactokinase activity 1.2483 1.2480 0.0002 0.0000 UBC1 YDR177W Ubiquitin‐conjugating enzyme that mediates selective degradation of short‐lived and abnormal proteins; plays a role in vesicle biogenesis and ER‐associated protein degradation (ERAD); component of the cellular stress response 1.2349 0.0000 NA YDL218W Putative protein of unknown function; YDL218W transcription is regulated by Azf1p and induced by starvation and aerobic conditions; expression also induced in cells treated with the mycotoxin patulin 1.2347 0.0002 DAL4 YIR028W Allantoin permease; expression sensitive to nitrogen catabolite repression and induced by allophanate, an intermediate in allantoin degradation 1.2322 0.0001 NCB2 YDR397C Subunit of a heterodimeric NC2 transcription regulator complex with Bur6p; complex binds to TBP and can repress transcription by preventing preinitiation complex assembly or stimulate activated transcription; homologous to human NC2beta 1.2308 0.0002 DUN1 YDL101C Cell‐cycle checkpoint serine‐threonine kinase required for DNA damage‐induced transcription of certain target genes, phosphorylation of Rad55p and Sml1p, and transient G2/M arrest after DNA damage; also regulates postreplicative DNA repair 1.2291 0.0000 NA YHR213W‐A Putative protein of unknown function; identified by gene‐trapping, microarray‐based expression analysis, and genome‐wide homology searching 1.2277 0.0002 CPR6 YLR216C Peptidyl‐prolyl cis‐trans isomerase (cyclophilin), catalyzes the cis‐ trans isomerization of peptide bonds N‐terminal to proline residues; binds to Hsp82p and contributes to chaperone activity 1.2266 0.0000 CRP1 COX17 YHR146W YLL009C Protein that binds to cruciform DNA structures Copper metallochaperone that transfers copper to Sco1p and Cox11p for eventual delivery to cytochrome c oxidase; contains twin cysteine‐x9‐cysteine motifs 1.2205 1.2191 0.0000 0.0001 KCC4 YCL024W Protein kinase of the bud neck involved in the septin checkpoint, associates with septin proteins, negatively regulates Swe1p by phosphorylation, shows structural homology to bud neck kinases Gin4p and Hsl1p 1.2177 0.0001 DOS2 YDR068W 1.2162 0.0000 NA YER137C Protein of unknown function, green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm Putative protein of unknown function 1.2108 0.0006 Symbols ORF Description LYS21 YDL131W INO2 220 log2(mtr1 0.Ts/WT.T s) adj.p.val Catalytic core subunit of the Anaphase‐Promoting Complex/Cyclosome (APC/C), which is a ubiquitin‐protein ligase required for degradation of anaphase inhibitors, including mitotic cyclins, during the metaphase/anaphase transition 1.2096 0.0000 Protein of unknown function; null mutant is resistant to 2‐deoxy‐D‐ glucose and displays abnormally elongated buds D‐Tyr‐tRNA(Tyr) deacylase, functions in protein translation, may affect nonsense suppression via alteration of the protein synthesis machinery; ubiquitous among eukaryotes Mitochondrial intermembrane space protein, required for normal oxygen consumption; contains twin cysteine‐x9‐cysteine motifs 1.2082 0.0001 1.2068 0.0000 1.2065 0.0000 Nucleolar protein of unknown function, positive regulator of mitotic exit; involved in regulating release of Cdc14p from the nucleolus in early anaphase, may play similar role in meiosis Phospholipid‐binding protein that interacts with both Ypt7p and Vps33p, may partially counteract the action of Vps33p and vice versa, localizes to the rim of the vacuole as cells approach stationary phase Nucleosomal histone H3‐Lys79 methylase; methylation is required for telomeric silencing, meiotic checkpoint control, and DNA damage response 1.2051 0.0000 1.2039 0.0000 1.1991 0.0001 Galactose permease, required for utilization of galactose; also able to transport glucose Chitin deacetylase, together with Cda2p involved in the biosynthesis ascospore wall component, chitosan; required for proper rigidity of the ascospore wall 1.1990 0.0009 1.1976 0.0007 YDR325W Subunit of the condensin complex; required for establishment and maintenance of chromosome condensation, chromosome segregation and chromatin binding of the condensin complex; required for clustering of tRNA genes at the nucleolus 1.1965 0.0001 NA YDR239C 1.1939 0.0000 STF1 YDL130W‐A Protein of unknown function that may interact with ribosomes, based on co‐purification experiments Protein involved in regulation of the mitochondrial F1F0‐ATP synthase; Stf1p and Stf2p may act as stabilizing factors that enhance inhibitory action of the Inh1p protein 1.1886 0.0001 EKI1 YDR147W Ethanolamine kinase, primarily responsible for phosphatidylethanolamine synthesis via the CDP‐ethanolamine pathway; exhibits some choline kinase activity, thus contributing to phosphatidylcholine synthesis via the CDP‐choline pathway 1.1874 0.0000 RUB1 YDR139C Ubiquitin‐like protein with similarity to mammalian NEDD8; conjugation (neddylation) substrates include the cullins Cdc53p, Rtt101p, and Cul3p; activated by Ula1p and Uba3p (E1 enzyme pair); conjugation mediated by Ubc12p (E2 enzyme) 1.1848 0.0001 NA YDR042C 1.1844 0.0000 SPO22 YIL073C Putative protein of unknown function; expression is increased in ssu72‐ts69 mutant Meiosis‐specific protein essential for chromosome synapsis, involved in completion of nuclear divisions during meiosis; induced early in meiosis 1.1840 0.0001 YSY6 YBR162W‐A Protein whose expression suppresses a secretory pathway mutation in E. coli; has similarity to the mammalian RAMP4 protein involved in secretion 1.1787 0.0001 SNU23 YDL098C Component of U4/U6.U5 snRNP involved in mRNA splicing via spliceosome 1.1786 0.0001 Symbols ORF Description APC11 YDL008W DGR2 YKL121W DTD1 YDL219W MIC14 YDR031W SPO12 YHR152W IVY1 YDR229W DOT1 YDR440W GAL2 YLR081W CDA1 YLR307W YCG1 221 log2(mtr1 0.Ts/WT.T s) adj.p.val Site‐specific endonuclease required for gene conversion at the MAT locus (homothallic switching) through the generation of a ds DNA break; expression restricted to mother cells in late G1 as controlled by Swi4p‐Swi6p, Swi5p and Ash1p 1.1763 0.0007 YLR412C‐A YMR263W Putative protein of unknown function Subunit of a histone deacetylase complex, along with Rpd3p and Sin3p, that is involved in silencing at telomeres, rDNA, and silent mating‐type loci; involved in telomere maintenance 1.1741 1.1732 0.0002 0.0001 GLE1 YDL207W Cytoplasmic nucleoporin required for polyadenylated RNA export but not for protein import; component of Nup82p nuclear pore subcomplex; contains a nuclear export signal 1.1727 0.0000 ECO1 YFR027W Acetyltransferase required for sister chromatid cohesion; modifies Smc3p at DNA replication forks during S‐phase; modifies Mcd1p in response to double‐strand DNA breaks during G2/M; mutations in human homolog ESCO2 cause Roberts syndrome 1.1724 0.0000 STI1 YOR027W Hsp90 cochaperone, interacts with the Ssa group of the cytosolic Hsp70 chaperones and activates Ssa1p ATPase activity; interacts with Hsp90 chaperones and inhibits their ATPase activity; homolog of mammalian Hop 1.1722 0.0001 NA YDR246W‐A 1.1708 0.0001 PUF2 YPR042C Putative protein of unknown function; identified by fungal homology and RT‐PCR Member of the PUF protein family, which is defined by the presence of Pumilio homology domains that confer RNA binding activity; preferentially binds mRNAs encoding membrane‐associated proteins 1.1686 0.0000 CDC7 YDL017W DDK (Dbf4‐dependent kinase) catalytic subunit required for firing origins and replication fork progression in S phase through phosphorylation of Mcm2‐7p complexes and Cdc45p; kinase activity correlates with cyclical DBF4 expression 1.1602 0.0000 ACM1 YPL267W Pseudosubstrate inhibitor of the anaphase‐promoting complex/cyclosome (APC/C), that suppresses APC/C [Cdh1]‐ mediated proteolysis of mitotic cyclins; associates with Cdh1p, Bmh1p and Bmh2p; cell cycle regulated protein 1.1601 0.0001 MED2 YDL005C Subunit of the RNA polymerase II mediator complex; associates with core polymerase subunits to form the RNA polymerase II holoenzyme; essential for transcriptional regulation 1.1594 0.0001 JNM1 YMR294W Component of the yeast dynactin complex, consisting of Nip100p, Jnm1p, and Arp1p; required for proper nuclear migration and spindle partitioning during mitotic anaphase B 1.1592 0.0001 NA YDR056C Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the endoplasmic reticulum; YDR056C is not an essential protein 1.1568 0.0000 DIA1 YMR316W Protein of unknown function, involved in invasive and pseudohyphal growth; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm in a punctate pattern 1.1550 0.0008 LSM7 YNL147W Lsm (Like Sm) protein; part of heteroheptameric complexes (Lsm2p‐ 7p and either Lsm1p or 8p): cytoplasmic Lsm1p complex involved in mRNA decay; nuclear Lsm8p complex part of U6 snRNP and possibly involved in processing tRNA, snoRNA, and rRNA 1.1490 0.0002 MRPL28 YDR462W Mitochondrial ribosomal protein of the large subunit 1.1477 0.0000 Symbols ORF Description HO YDL227C NA SAP30 222 log2(mtr1 0.Ts/WT.T s) adj.p.val Nucleolar protein that forms a complex with Csm1p, and then Mam1p at kinetochores during meiosis I to mediate accurate homolog segregation; required for condensin recruitment to the replication fork barrier site and rDNA repeat segregation 1.1447 0.0001 YMR186W Cytoplasmic chaperone of the Hsp90 family, redundant in function and nearly identical with Hsp82p, and together they are essential; expressed constitutively at 10‐fold higher basal levels than HSP82 and induced 2‐3 fold by heat shock 1.1445 0.0002 NGG1 YDR176W Transcriptional regulator involved in glucose repression of Gal4p‐ regulated genes; component of transcriptional adaptor and histone acetyltransferase complexes, the ADA complex, the SAGA complex, and the SLIK complex 1.1441 0.0002 PAU4 YLR461W Member of the seripauperin multigene family encoded mainly in subtelomeric regions; active during alcoholic fermentation, regulated by anaerobiosis, negatively regulated by oxygen, repressed by heme 1.1436 0.0000 PIB1 YDR313C RING‐type ubiquitin ligase of the endosomal and vacuolar membranes, binds phosphatidylinositol(3)‐phosphate; contains a FYVE finger domain 1.1433 0.0001 DIT1 YDR403W Sporulation‐specific enzyme required for spore wall maturation, involved in the production of a soluble LL‐dityrosine‐containing precursor of the spore wall; transcripts accumulate at the time of prospore enclosure 1.1419 0.0003 HPF1 YOL155C 1.1410 0.0001 GPR1 YDL035C Haze‐protective mannoprotein that reduces the particle size of aggregated proteins in white wines Plasma membrane G protein coupled receptor (GPCR) that interacts with the heterotrimeric G protein alpha subunit, Gpa2p, and with Plc1p; sensor that integrates nutritional signals with the modulation of cell fate via PKA and cAMP synthesis 1.1394 0.0000 MRPL7 SRB7 YDR237W YDR308C Mitochondrial ribosomal protein of the large subunit Subunit of the RNA polymerase II mediator complex; associates with core polymerase subunits to form the RNA polymerase II holoenzyme; essential for transcriptional regulation; target of the global repressor Tup1p 1.1389 1.1340 0.0001 0.0000 NA YCR075W‐A 1.1325 0.0001 SPT3 YDR392W Putative protein of unknown function; identified by homology to Ashbya gossypii Subunit of the SAGA and SAGA‐like transcriptional regulatory complexes, interacts with Spt15p to activate transcription of some RNA polymerase II‐dependent genes, also functions to inhibit transcription at some promoters 1.1291 0.0001 NA YDR348C Protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cell periphery and bud neck; potential Cdc28p substrate 1.1278 0.0000 PLM2 YDR501W Forkhead Associated domain containing protein and putative transcription factor found associated with chromatin; target of SBF transcription factor; induced in response to DNA damaging agents and deletion of telomerase; similar to TOS4 1.1275 0.0001 RRP42 YDL111C Exosome non‐catalytic core component; involved in 3'‐5' RNA processing and degradation in both the nucleus and the cytoplasm; has similarity to E. coli RNase PH and to human hRrp42p (EXOSC7) 1.1274 0.0000 Symbols ORF Description LRS4 YDR439W HSC82 223 log2(mtr1 0.Ts/WT.T s) adj.p.val Nucleolar protein that forms a complex with Lrs4p and then Mam1p at kinetochores during meiosis I to mediate accurate homolog segregation; required for condensin recruitment to the replication fork barrier site and rDNA repeat segregation 1.1273 0.0002 YLR298C Component of the U1 snRNP complex required for pre‐mRNA splicing; putative ortholog of human U1C protein, which is involved in formation of a complex between U1 snRNP and the pre‐mRNA 5' splice site 1.1248 0.0001 NA RNH203 YER188C‐A YLR154C Putative protein of unknown function Ribonuclease H2 subunit, required for RNase H2 activity; related to human AGS3 that causes Aicardi‐Goutieres syndrome 1.1247 1.1238 0.0005 0.0000 LSM3 YLR438C‐A Lsm (Like Sm) protein; part of heteroheptameric complexes (Lsm2p‐ 7p and either Lsm1p or 8p): cytoplasmic Lsm1p complex involved in mRNA decay; nuclear Lsm8p complex part of U6 snRNP and possibly involved in processing tRNA, snoRNA, and rRNA 1.1232 0.0001 RRP14 YKL082C Essential protein, constituent of 66S pre‐ribosomal particles; interacts with proteins involved in ribosomal biogenesis and cell polarity; member of the SURF‐6 family 1.1230 0.0005 TLG1 YDR468C Essential t‐SNARE that forms a complex with Tlg2p and Vti1p and mediates fusion of endosome‐derived vesicles with the late Golgi; binds the docking complex VFT (Vps fifty‐three) through interaction with Vps51p 1.1220 0.0001 AHA1 YDR214W Co‐chaperone that binds to Hsp82p and activates its ATPase activity; similar to Hch1p; expression is regulated by stresses such as heat shock 1.1212 0.0002 TOA1 YOR194C TFIIA large subunit; involved in transcriptional activation, acts as antirepressor or as coactivator; homologous to largest and second largest subunits of human and Drosophila TFIIA 1.1157 0.0004 SCM3 YDL139C Nonhistone component of centromeric chromatin that binds stoichiometrically to CenH3‐H4 histones, required for kinetochore assembly; required for G2/M progression and localization of Cse4p; may protect Cse4p from ubiquitylation 1.1153 0.0000 NA YNL146W Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the endoplasmic reticulum; YNL146W is not an essential gene 1.1144 0.0000 PUT1 YLR142W Proline oxidase, nuclear‐encoded mitochondrial protein involved in utilization of proline as sole nitrogen source; PUT1 transcription is induced by Put3p in the presence of proline and the absence of a preferred nitrogen source 1.1121 0.0002 ENT1 YDL161W Epsin‐like protein involved in endocytosis and actin patch assembly and functionally redundant with Ent2p; binds clathrin via a clathrin‐ binding domain motif at C‐terminus 1.1117 0.0000 OGG1 YML060W Mitochondrial glycosylase/lyase that specifically excises 7,8‐dihydro‐ 8‐oxoguanine residues located opposite cytosine or thymine residues in DNA, repairs oxidative damage to mitochondrial DNA, contributes to UVA resistance 1.1106 0.0005 SPC29 YPL124W Inner plaque spindle pole body (SPB) component, links the central plaque component Spc42p to the inner plaque component Spc110p; required for SPB duplication 1.1105 0.0000 Symbols ORF Description CSM1 YCR086W YHC1 224 log2(mtr1 0.Ts/WT.T s) adj.p.val Non‐essential kinetochore protein, subunit of the Ctf19 central kinetochore complex (Ctf19p‐Mcm21p‐Okp1p‐Mcm22p‐Mcm16p‐ Ctf3p‐Chl4p‐Mcm19p‐Nkp1p‐Nkp2p‐Ame1p‐Mtw1p) 1.1092 0.0000 YDR346C Protein with a potential role in cell survival pathways, required for the diauxic growth shift; expression in mammalian cells increases survival under conditions inducing apoptosis; mutant has increased aneuploidy tolerance 1.1080 0.0002 SLK19 YOR195W Kinetochore‐associated protein required for normal segregation of chromosomes in meiosis and mitosis; component of the FEAR regulatory network, which promotes Cdc14p release from the nucleolus during anaphase; potential Cdc28p substrate 1.1078 0.0005 SEN34 YAR008W Subunit of the tRNA splicing endonuclease, which is composed of Sen2p, Sen15p, Sen34p, and Sen54p; Sen34p contains the active site for tRNA 3' splice site cleavage and has similarity to Sen2p and to Archaeal tRNA splicing endonuclease 1.1054 0.0000 SWI5 YDR146C Transcription factor that activates transcription of genes expressed at the M/G1 phase boundary and in G1 phase; localization to the nucleus occurs during G1 and appears to be regulated by phosphorylation by Cdc28p kinase 1.1050 0.0005 GIN4 YDR507C Protein kinase involved in bud growth and assembly of the septin ring, proposed to have kinase‐dependent and kinase‐independent activities; undergoes autophosphorylation; similar to Kcc4p and Hsl1p 1.1048 0.0002 SNM1 YDR478W Subunit of RNase MRP, which cleaves pre‐rRNA and has a role in cell cycle‐regulated degradation of daughter cell‐specific mRNAs; binds to the NME1 RNA subunit of RNase MRP 1.0996 0.0000 NA NA MHR1 NA YIL014C‐A YDR296W NA Putative protein of unknown function Protein involved in homologous recombination in mitochondria; required for recombination‐dependent mtDNA partitioning; involved in stimulation of mitochondrial DNA replication in response to oxidative stress 1.0981 1.0964 1.0960 0.0001 0.0002 0.0001 MPS1 YDL028C Dual‐specificity kinase required for spindle pole body (SPB) duplication and spindle checkpoint function; substrates include SPB proteins Spc42p, Spc110p, and Spc98p, mitotic exit network protein Mob1p, and checkpoint protein Mad1p 1.0936 0.0001 SWA2 YDR320C Auxilin‐like protein involved in vesicular transport; clathrin‐binding protein required for uncoating of clathrin‐coated vesicles 1.0901 0.0001 CDC31 YOR257W Calcium‐binding component of the spindle pole body (SPB) half‐ bridge, required for SPB duplication in mitosis and meiosis II; homolog of mammalian centrin; binds multiubiquitinated proteins and is involved in proteasomal protein degradation 1.0887 0.0001 ATG10 YLL042C Conserved E2‐like conjugating enzyme that mediates formation of the Atg12p‐Atg5p conjugate, which is a critical step in autophagy 1.0870 0.0001 UGX2 YDL169C 1.0848 0.0004 NA YDL012C Protein of unknown function, transcript accumulates in response to any combination of stress conditions Tail‐anchored plasma membrane protein containing a conserved CYSTM module, possibly involved in response to stress; may contribute to non‐homologous end‐joining (NHEJ) based on ydl012c htz1 double null phenotype 1.0845 0.0000 NA YFR012W Putative protein of unknown function 1.0824 0.0007 Symbols ORF Description NKP1 YDR383C SVF1 225 log2(mtr1 0.Ts/WT.T s) adj.p.val Protein with similarity to P‐type ATPase sodium pumps, member of the Na+ efflux ATPase family Inner plaque spindle pole body (SPB) component, ortholog of human kendrin; involved in connecting nuclear microtubules to SPB; interacts with Tub4p‐complex and calmodulin; phosphorylated by Mps1p in cell cycle‐dependent manner 1.0798 0.0002 1.0789 0.0001 YDR386W Subunit of the structure‐specific Mms4p‐Mus81p endonuclease that cleaves branched DNA; involved in DNA repair, replication fork stability, and joint molecule formation/resolution during meiotic recombination; helix‐hairpin‐helix protein 1.0781 0.0001 HXT14 YNL318C Protein with similarity to hexose transporter family members, expression is induced in low glucose and repressed in high glucose; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐throughput studies 1.0773 0.0005 FCF1 YDR339C Putative PINc domain nuclease required for early cleavages of 35S pre‐rRNA and maturation of 18S rRNA; component of the SSU (small subunit) processome involved in 40S ribosomal subunit biogenesis; copurifies with Faf1p 1.0764 0.0003 NHP2 YDL208W Nuclear protein related to mammalian high mobility group (HMG) proteins, essential for function of H/ACA‐type snoRNPs, which are involved in 18S rRNA processing 1.0750 0.0009 CDC9 YDL164C DNA ligase found in the nucleus and mitochondria, an essential enzyme that joins Okazaki fragments during DNA replication; also acts in nucleotide excision repair, base excision repair, and recombination 1.0744 0.0002 TFB3 YDR460W Subunit of TFIIH and nucleotide excision repair factor 3 complexes, involved in transcription initiation, required for nucleotide excision repair; ring finger protein similar to mammalian CAK and TFIIH subunit 1.0731 0.0001 NA YDR248C Putative protein of unknown function; sequence similarity to bacterial and human gluconokinase; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm; upregulated by deletion of the RNAP‐II associated factor, PAF1 1.0724 0.0001 PGM3 YMR278W Phosphoglucomutase, catalyzes interconversion of glucose‐1‐ phosphate and glucose‐6‐phospate; transcription induced in response to stress; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm and nucleus; non‐essential 1.0717 0.0001 NA YBR196C‐A 1.0686 0.0002 SSF2 YDR312W Putative protein of unknown function; identified by fungal homology and RT‐PCR Protein required for ribosomal large subunit maturation, functionally redundant with Ssf1p; member of the Brix family 1.0678 0.0001 RKM2 YDR198C Ribosomal protein lysine methyltransferase, responsible for trimethylation of the lysine residue at position 3 of Rpl12Ap and Rpl12Bp 1.0671 0.0001 RMD1 UGA4 YDL001W YDL210W Cytoplasmic protein required for sporulation Permease that serves as a gamma‐aminobutyrate (GABA) transport protein involved in the utilization of GABA as a nitrogen source; catalyzes the transport of putrescine and delta‐aminolevulinic acid (ALA); localized to the vacuolar membrane 1.0655 1.0625 0.0001 0.0002 TMA7 YLR262C‐A Protein of unknown that associates with ribosomes; null mutant exhibits translation defects, altered polyribosome profiles, and resistance to the translation inhibitor anisomcyin 1.0617 0.0002 Symbols ORF Description ENA5 YDR038C SPC110 YDR356W MUS81 226 log2(mtr1 0.Ts/WT.T s) adj.p.val Sm‐like protein involved in docking and fusion of exocytic vesicles through binding to assembled SNARE complexes at the membrane; localization to sites of secretion (bud neck and bud tip) is dependent on SNARE function 1.0606 0.0000 YDL036C Mitochondrial tRNA:pseudouridine synthase, catalyzes the formation of pseudouridine at position 32 in mitochondrial tRNAs; contains an N‐terminal mitochondrial targeting sequence 1.0593 0.0000 NA YDR115W Putative mitochondrial ribosomal protein of the large subunit, has similarity to E. coli L34 ribosomal protein; required for respiratory growth, as are most mitochondrial ribosomal proteins 1.0590 0.0001 NA YHL048C‐A 1.0550 0.0001 LIF1 YGL090W Putative protein of unknown function; identified by expression profiling and mass spectrometry Component of the DNA ligase IV complex that mediates nonhomologous end joining in DNA double‐strand break repair; physically interacts with Dnl4p and Nej1p; homologous to mammalian XRCC4 protein 1.0515 0.0002 ARO80 YDR421W Zinc finger transcriptional activator of the Zn2Cys6 family; activates transcription of aromatic amino acid catabolic genes in the presence of aromatic amino acids 1.0499 0.0000 GYP7 YDL234C GTPase‐activating protein for yeast Rab family members including: Ypt7p (most effective), Ypt1p, Ypt31p, and Ypt32p (in vitro); involved in vesicle mediated protein trafficking 1.0489 0.0001 CIN4 YMR138W GTP‐binding protein involved in beta‐tubulin (Tub2p) folding; isolated as mutant with increased chromosome loss and sensitivity to benomyl; regulated by the GTPase‐activating protein, Cin2p, the human retinitis pigmentosa 2 (RP2) homolog 1.0480 0.0004 MZM1 YDR493W Mitochondrial matrix protein with a role in maintaining the labile mitochondrial zinc pool; null mutant has a respiratory growth defect and an elevated frequency of mitochondrial genome loss; overexpression causes cell cycle delay or arrest 1.0463 0.0000 GPI11 YDR302W ER membrane protein involved in a late step of glycosylphosphatidylinositol (GPI) anchor assembly; involved in the addition of phosphoethanolamine to the multiply mannosylated GPI intermediate; human PIG‐Fp is a functional homolog 1.0453 0.0002 CSE4 YKL049C Centromere protein that resembles histone H3, required for proper kinetochore function; homolog of human CENP‐A; levels are regulated by E3 ubiquitin ligase Psh1p 1.0420 0.0002 SEC20 YDR498C Membrane glycoprotein v‐SNARE involved in retrograde transport from the Golgi to the ER; required for N‐ and O‐glycosylation in the Golgi but not in the ER; interacts with the Dsl1p complex through Tip20p 1.0418 0.0001 RTT102 YGR275W Component of both the SWI/SNF and RSC chromatin remodeling complexes, suggested role in chromosome maintenance; possible weak regulator of Ty1 transposition 1.0416 0.0001 NA YMR244C‐A Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to both the cytoplasm and nucleus and is induced in response to the DNA‐damaging agent MMS; YMR244C‐A is not an essential gene 1.0404 0.0001 SVS1 YPL163C Cell wall and vacuolar protein, required for wild‐type resistance to vanadate 1.0397 0.0001 Symbols ORF Description SEC1 YDR164C PUS9 227 log2(mtr1 0.Ts/WT.T s) adj.p.val Nucleosome assembly factor, involved in chromatin assembly and disassembly, anti‐silencing protein that causes derepression of silent loci when overexpressed; plays a role in regulating Ty1 transposition 1.0389 0.0000 YMR256C Subunit VII of cytochrome c oxidase, which is the terminal member of the mitochondrial inner membrane electron transport chain 1.0300 0.0001 NA YBR071W Putative protein of unknown function; (GFP)‐fusion and epitope‐ tagged proteins localize to the cytoplasm; mRNA expression may be regulated by the cell cycle and/or cell wall stress 1.0222 0.0001 SGT2 YOR007C Glutamine‐rich cytoplasmic protein that serves as a scaffold for binding Get4/5p and other proteins required to mediate posttranslational insertion of tail‐anchored proteins into the ER membrane; has similarity to human cochaperone SGT 1.0178 0.0001 SMT3 YDR510W Ubiquitin‐like protein of the SUMO family, conjugated to lysine residues of target proteins; regulates chromatid cohesion, chromosome segregation, APC‐mediated proteolysis, DNA replication and septin ring dynamics; phosphorylated at Ser2 1.0156 0.0001 UBC9 YDL064W SUMO‐conjugating enzyme involved in the Smt3p conjugation pathway; nuclear protein required for S‐ and M‐phase cyclin degradation and mitotic control; involved in proteolysis mediated by the anaphase‐promoting complex cyclosome (APCC) 1.0136 0.0000 RPC11 YDR045C RNA polymerase III subunit C11; mediates pol III RNA cleavage activity and is important for termination of transcription; homologous to TFIIS 1.0118 0.0001 RGA2 YDR379W GTPase‐activating protein for the polarity‐establishment protein Cdc42p; implicated in control of septin organization, pheromone response, and haploid invasive growth; regulated by Pho85p and Cdc28p 1.0105 0.0002 PBI2 YNL015W Cytosolic inhibitor of vacuolar proteinase B (PRB1), required for efficient vacuole inheritance; with thioredoxin forms protein complex LMA1, which assists in priming SNARE molecules and promotes vacuole fusion 1.0099 0.0002 NUP42 YDR192C Subunit of the nuclear pore complex (NPC) that localizes exclusively to the cytoplasmic side; involved in RNA export, most likely at a terminal step; interacts with Gle1p 1.0084 0.0003 SLY1 YDR189W Hydrophilic protein involved in vesicle trafficking between the ER and Golgi; SM (Sec1/Munc‐18) family protein that binds the tSNARE Sed5p and stimulates its assembly into a trans‐SNARE membrane‐ protein complex 1.0084 0.0001 GCD6 YDR211W Catalytic epsilon subunit of the translation initiation factor eIF2B, the guanine‐nucleotide exchange factor for eIF2; activity subsequently regulated by phosphorylated eIF2; first identified as a negative regulator of GCN4 expression 1.0055 0.0001 PRP3 ENT5 YDR473C YDR153C Splicing factor, component of the U4/U6‐U5 snRNP complex Protein containing an N‐terminal epsin‐like domain involved in clathrin recruitment and traffic between the Golgi and endosomes; associates with the clathrin adaptor Gga2p, clathrin adaptor complex AP‐1, and clathrin 1.0039 1.0031 0.0000 0.0001 CDC36 YDL165W Component of the CCR4‐NOT complex, which has multiple roles in regulating mRNA levels including regulation of transcription and destabilizing mRNAs by deadenylation; basal transcription factor 1.0030 0.0001 Symbols ORF Description ASF1 YJL115W COX7 228 log2(mtr1 0.Ts/WT.T s) adj.p.val eIF3g subunit of the core complex of translation initiation factor 3 (eIF3), which is essential for translation; stimulates resumption of ribosomal scanning during translation reinitiation 1.0028 0.0008 YDR049W Zinc finger protein, forms a mitochondrially‐associated complex with Cdc48p and Npl4p under oxidative stress that is required for ubiquitin‐mediated mitochondria‐associated protein degradation (MAD); conserved in C. elegans and human 1.0025 0.0000 MOD5 YOR274W Delta 2‐isopentenyl pyrophosphate:tRNA isopentenyl transferase, required for biosynthesis of the modified base isopentenyladenosine in mitochondrial and cytoplasmic tRNAs; gene is nuclear and encodes two isozymic forms 0.9998 0.0005 TEN1 YLR010C 0.9983 0.0001 PTP1 YDL230W Protein that regulates telomeric length; protects telomeric ends in a complex with Cdc13p and Stn1p Phosphotyrosine‐specific protein phosphatase that dephosphorylates a broad range of substrates in vivo, including Fpr3p; localized to the cytoplasm and the mitochondria 0.9980 0.0000 MRPS16 ACN9 YPL013C YDR511W Mitochondrial ribosomal protein of the small subunit Protein of the mitochondrial intermembrane space, required for acetate utilization and gluconeogenesis; has orthologs in higher eukaryotes 0.9975 0.9966 0.0003 0.0001 PDC2 YDR081C Transcription factor required for the synthesis of the glycolytic enzyme pyruvate decarboxylase, required for high level expression of both the THI and the PDC genes 0.9940 0.0000 MAD1 YGL086W Coiled‐coil protein involved in the spindle‐assembly checkpoint; phosphorylated by Mps1p upon checkpoint activation which leads to inhibition of the activity of the anaphase promoting complex; forms a complex with Mad2p 0.9920 0.0002 PEX3 YDR329C Peroxisomal membrane protein (PMP) required for proper localization and stability of PMPs; anchors peroxisome retention factor Inp1p at the peroxisomal membrane; interacts with Pex19p 0.9917 0.0000 HSP104 YLL026W Heat shock protein that cooperates with Ydj1p (Hsp40) and Ssa1p (Hsp70) to refold and reactivate previously denatured, aggregated proteins; responsive to stresses including: heat, ethanol, and sodium arsenite; involved in [PSI+] propagation 0.9916 0.0004 IZH1 YDR492W Membrane protein involved in zinc ion homeostasis, member of the four‐protein IZH family; transcription is regulated directly by Zap1p, expression induced by zinc deficiency and fatty acids; deletion increases sensitivity to elevated zinc 0.9907 0.0002 YFH1 YDL120W Mitochondrial matrix iron chaperone; oxidizes and stores iron; interacts with Isu1p to promote Fe‐S cluster assembly; mutation results in multiple Fe/S‐dependent enzyme deficiencies; human frataxin homolog is mutated in Friedrich's ataxia 0.9823 0.0002 SFA1 YDL168W Bifunctional enzyme containing both alcohol dehydrogenase and glutathione‐dependent formaldehyde dehydrogenase activities, functions in formaldehyde detoxification and formation of long chain and complex alcohols, regulated by Hog1p‐Sko1p 0.9756 0.0000 UBA2 YDR390C Subunit of a heterodimeric nuclear SUMO activating enzyme (E1) with Aos1p; activates Smt3p (SUMO) before its conjugation to proteins (sumoylation), which may play a role in protein targeting; essential for viability 0.9726 0.0000 Symbols ORF Description TIF35 YDR429C VMS1 229 log2(mtr1 0.Ts/WT.T s) adj.p.val Putative protein of unknown function; may interact with ribosomes, based on co‐purification experiments; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm 0.9718 0.0001 YDL178W D‐lactate dehydrogenase, located in the mitochondrial matrix 0.9696 0.0002 GTR2 YGR163W Putative GTP binding protein that negatively regulates Ran/Tc4 GTPase cycle; activates transcription; subunit of EGO and GSE complexes; required for sorting of Gap1p; localizes to cytoplasm and to chromatin; homolog of human RagC and RagD 0.9679 0.0007 MTQ2 YDR140W S‐adenosylmethionine‐dependent methyltransferase of the seven beta‐strand family; subunit of complex with Trm112p that methylates translation release factor Sup45p (eRF1) in the ternary complex eRF1‐eRF3‐GTP; similar to E.coli PrmC 0.9671 0.0002 BLS1 YLR408C Putative protein of unknown function; likely member of BLOC complex involved in endosomal cargo sorting; green fluorescent protein (GFP)‐fusion protein localizes to the endosome; YLR408C is not an essential gene 0.9668 0.0000 NA YHR138C Putative protein of unknown function; has similarity to Pbi2p; double null mutant lacking Pbi2p and Yhr138p exhibits highly fragmented vacuoles 0.9634 0.0002 NA YJL144W Cytoplasmic hydrophilin of unknown function, possibly involved in the dessication response; expression induced by osmotic stress, starvation and during stationary phase; GFP‐fusion protein is induced by the DNA‐damaging agent MMS 0.9623 0.0001 HEM13 YDR044W Coproporphyrinogen III oxidase, an oxygen requiring enzyme that catalyzes the sixth step in the heme biosynthetic pathway; transcription is repressed by oxygen and heme (via Rox1p and Hap1p) 0.9613 0.0001 PRP9 YDL030W Subunit of the SF3a splicing factor complex, required for spliceosome assembly; acts after the formation of the U1 snRNP‐ pre‐mRNA complex 0.9609 0.0000 CRH1 YGR189C Chitin transglycosylase that functions in the transfer of chitin to beta(1‐6) and beta(1‐3) glucans in the cell wall; similar and functionally redundant to Utr2; localizes to sites of polarized growth; expression induced by cell wall stress 0.9581 0.0001 JEM1 YJL073W DnaJ‐like chaperone required for nuclear membrane fusion during mating, localizes to the ER membrane; exhibits genetic interactions with KAR2 0.9574 0.0001 NA YLL066W‐B 0.9551 0.0008 PRO1 YDR300C 0.9528 0.0005 NA YLL066W‐B 0.9523 0.0006 DFM1 YDR411C Putative protein of unknown function; overexpression causes a cell cycle delay or arrest Gamma‐glutamyl kinase, catalyzes the first step in proline biosynthesis Putative protein of unknown function; overexpression causes a cell cycle delay or arrest Endoplasmic reticulum (ER) localized protein involved in ER‐ associated protein degradation (ERAD), ER stress and homeostasis; interacts with components of ERAD‐L and ERAD‐C and Cdc48p; derlin‐like family member similar to Der1p 0.9511 0.0001 PAU18 YLL064C Protein of unknown function, member of the seripauperin multigene family encoded mainly in subtelomeric regions; identical to Pau6p 0.9495 0.0001 Symbols ORF Description NA YDR186C DLD2 230 log2(mtr1 0.Ts/WT.T s) adj.p.val Subunit of the SAS complex (Sas2p, Sas4p, Sas5p), which acetylates free histones and nucleosomes and regulates transcriptional silencing; required for the HAT activity of Sas2p 0.9494 0.0001 YPL192C Pheromone‐regulated protein required for nuclear envelope fusion during karyogamy; localizes to the outer face of the nuclear membrane; interacts with Kar5p at the spindle pole body 0.9489 0.0001 TIM22 YDL217C Essential core component of the mitochondrial TIM22 complex involved in insertion of polytopic proteins into the inner membrane; forms the channel through which proteins are imported 0.9480 0.0001 RTR2 YDR066C 0.9477 0.0001 MRPL51 POL3 YPR100W YDL102W Protein of unknown function with high similarity to Rtr1p; exhibits genetic interactions with Rtr1p; green fluorescent protein (GFP)‐ fusion protein localizes to the cytoplasm; YDR066C is not an essential gene Mitochondrial ribosomal protein of the large subunit Catalytic subunit of DNA polymerase delta; required for chromosomal DNA replication during mitosis and meiosis, intragenic recombination, repair of double strand DNA breaks, and DNA replication during nucleotide excision repair (NER) 0.9476 0.9453 0.0002 0.0000 NA YDL211C 0.9435 0.0003 ALG14 YBR070C Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the vacuole Component of UDP‐GlcNAc transferase required for the second step of dolichyl‐linked oligosaccharide synthesis; anchors the catalytic subunit Alg13p to the ER membrane; similar to bacterial and human glycosyltransferases 0.9365 0.0001 GRX3 YDR098C Hydroperoxide and superoxide‐radical responsive glutathione‐ dependent oxidoreductase; monothiol glutaredoxin subfamily member along with Grx4p and Grx5p; protects cells from oxidative damage 0.9362 0.0000 NRM1 YNR009W Transcriptional co‐repressor of MBF (MCB binding factor)‐regulated gene expression; Nrm1p associates stably with promoters via MBF to repress transcription upon exit from G1 phase 0.9335 0.0005 OST5 YGL226C‐A Zeta subunit of the oligosaccharyltransferase complex of the ER lumen, which catalyzes asparagine‐linked glycosylation of newly synthesized proteins 0.9335 0.0001 NA RPB11 YDR282C YOL005C 0.9304 0.9296 0.0001 0.0001 SHE9 YDR393W Putative protein of unknown function RNA polymerase II subunit B12.5; part of central core; similar to Rpc19p and bacterial alpha subunit Mitochondrial inner membrane protein required for normal mitochondrial morphology, may be involved in fission of the inner membrane; forms a homo‐oligomeric complex 0.9295 0.0002 PAM16 YJL104W Constituent of the import motor (PAM complex) component of the Translocase of the Inner Mitochondrial membrane (TIM23 complex); forms a 1:1 subcomplex with Pam18p and inhibits its cochaperone activity; contains a J‐like domain 0.9265 0.0001 MUD2 YKL074C Protein involved in early pre‐mRNA splicing; component of the pre‐ mRNA‐U1 snRNP complex, the commitment complex; interacts with Msl5p/BBP splicing factor and Sub2p; similar to metazoan splicing factor U2AF65 0.9256 0.0002 Symbols ORF Description SAS4 YDR181C PRM3 231 log2(mtr1 0.Ts/WT.T s) adj.p.val Peripheral membrane protein that is required for vacuolar H+‐ ATPase (V‐ATPase) function, although not an actual component of the V‐ATPase complex; functions in the assembly of the V‐ATPase; localized to the yeast endoplasmic reticulum (ER) 0.9246 0.0001 YKR078W Cytoplasmic protein of unknown function, has similarity to Vps5p; potential Cdc28p substrate; contains a Phox homology (PX) domain and specifically binds phosphatidylinositol 3‐phosphate (PtdIns‐3‐P) 0.9242 0.0003 MSS2 YDL107W Peripherally bound inner membrane protein of the mitochondrial matrix involved in membrane insertion of C‐terminus of Cox2p, interacts genetically and physically with Cox18p 0.9220 0.0001 SPC72 YAL047C Component of the cytoplasmic Tub4p (gamma‐tubulin) complex, binds spindle pole bodies and links them to microtubules; has roles in astral microtubule formation and stabilization 0.9199 0.0001 GET3 YDL100C Guanine nucleotide exchange factor for Gpa1p; amplifies G protein signaling; subunit of the GET complex, which is involved in Golgi to ER trafficking and insertion of proteins into the ER membrane; has low‐level ATPase activity 0.9188 0.0000 NA YDR124W 0.9173 0.0002 NA ASF2 YDR366C YDL197C 0.9171 0.9165 0.0001 0.0001 NA YNR075C‐A 0.9165 0.0002 ATP16 YDL004W Putative protein of unknown function; non‐essential gene; expression is strongly induced by alpha factor Putative protein of unknown function Anti‐silencing protein that causes derepression of silent loci when overexpressed Identified by gene‐trapping, microarray‐based expression analysis, and genome‐wide homology searching Delta subunit of the central stalk of mitochondrial F1F0 ATP synthase, which is a large, evolutionarily conserved enzyme complex required for ATP synthesis; phosphorylated 0.9148 0.0001 NA YPR108W‐A 0.9125 0.0006 RVB1 YDR190C Putative protein of unknown function; identified by fungal homology and RT‐PCR Essential protein involved in transcription regulation; component of chromatin remodeling complexes; required for assembly and function of the INO80 complex; also referred to as pontin; member of the RUVB‐like protein family 0.9094 0.0002 CDC1 YDR182W Putative lipid phosphatase of the endoplasmic reticulum; shows Mn2+ dependence and may affect Ca2+ signaling; mutants display actin and general growth defects and pleiotropic defects in cell cycle progression and organelle distribution 0.9069 0.0000 LSM6 YDR378C Lsm (Like Sm) protein; part of heteroheptameric complexes (Lsm2p‐ 7p and either Lsm1p or 8p): cytoplasmic Lsm1p complex involved in mRNA decay; nuclear Lsm8p complex part of U6 snRNP and possibly involved in processing tRNA, snoRNA, and rRNA 0.9065 0.0001 RPT3 YDR394W One of six ATPases of the 19S regulatory particle of the 26S proteasome involved in the degradation of ubiquitinated substrates; substrate of N‐acetyltransferase B 0.9025 0.0000 TAH1 YCR060W 0.9018 0.0002 THI13 YDL244W HSP90 cofactor; interacts with Hsp82p, Pih1p, Rvb1 and Rvb2, contains a single TPR domain with at least two TPR motifs Protein involved in synthesis of the thiamine precursor hydroxymethylpyrimidine (HMP); member of a subtelomeric gene family including THI5, THI11, THI12, and THI13 0.9012 0.0002 Symbols ORF Description VMA22 YHR060W NA 232 log2(mtr1 0.Ts/WT.T s) adj.p.val Subunit of the Sec61p translocation complex (Sec61p‐Sss1p‐Sbh1p) that forms a channel for passage of secretory proteins through the endoplasmic reticulum membrane, and of the Ssh1p complex (Ssh1p‐Sbh2p‐Sss1p); interacts with Ost4p and Wbp1p 0.9007 0.0005 YDR195W RNA‐binding protein involved in the cleavage step of mRNA 3'‐end formation prior to polyadenylation, and in snoRNA maturation; part of holo‐CPF subcomplex APT, which associates with 3'‐ends of snoRNA‐ and mRNA‐encoding genes 0.8998 0.0001 UBX3 YDL091C UBX (ubiquitin regulatory X) domain‐containing protein that interacts with Cdc48p, green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm in a punctate pattern 0.8993 0.0003 RSM19 YNR037C 0.8988 0.0004 COA2 YPL189C‐A Mitochondrial ribosomal protein of the small subunit, has similarity to E. coli S19 ribosomal protein Cytochrome oxidase assembly factor; null mutation results in respiratory deficiency with specific loss of cytochrome oxidase activity; functions downstream of assembly factors Mss51p and Coa1p and interacts with assembly factor Shy1p 0.8930 0.0007 MTW1 YAL034W‐A Essential component of the MIND kinetochore complex (Mtw1p Including Nnf1p‐Nsl1p‐Dsn1p) which joins kinetochore subunits contacting DNA to those contacting microtubules; critical to kinetochore assembly 0.8921 0.0004 NA YDL121C Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the endoplasmic reticulum; YDL121C is not an essential protein 0.8914 0.0003 YPD1 YDL235C Phosphorelay intermediate protein, phosphorylated by the plasma membrane sensor Sln1p in response to osmotic stress and then in turn phosphorylates the response regulators Ssk1p in the cytosol and Skn7p in the nucleus 0.8907 0.0001 TAF1 YGR274C TFIID subunit (145 kDa), involved in RNA polymerase II transcription initiation; possesses in vitro histone acetyltransferase activity but its role in vivo appears to be minor; involved in promoter binding and G1/S progression 0.8886 0.0004 SRN2 YLR119W Component of the ESCRT‐I complex, which is involved in ubiquitin‐ dependent sorting of proteins into the endosome; suppressor of rna1‐1 mutation; may be involved in RNA export from nucleus 0.8881 0.0005 DET1 YDR051C Acid phosphatase involved in the non‐vesicular transport of sterols in both directions between the endoplasmic reticulum and plasma membrane; deletion confers sensitivity to nickel 0.8868 0.0001 NA YDL176W Protein of unknown function, predicted by computational methods to be involved in fructose‐1,6‐bisphosphatase (Fbp1p) degradation; interacts with components of the GID complex; YDL176W is not an essential gene 0.8862 0.0002 NA YDR131C 0.8859 0.0001 ARG82 YDR173C F‐box protein, substrate‐specific adaptor subunit that recruits substrates to a core ubiquitination complex Inositol polyphosphate multikinase (IPMK), sequentially phosphorylates Ins(1,4,5)P3 to form Ins(1,3,4,5,6)P5; also has diphosphoinositol polyphosphate synthase activity; regulates arginine‐, phosphate‐, and nitrogen‐responsive genes 0.8848 0.0002 NA YDL086W Putative protein of unknown function; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐ throughput studies; YDL086W is not an essential gene 0.8846 0.0001 Symbols ORF Description SSS1 YDR086C REF2 233 log2(mtr1 0.Ts/WT.T s) adj.p.val Meiosis‐specific transcription factor required for exit from pachytene and for full meiotic recombination; activates middle sporulation genes; competes with Sum1p for binding to promoters containing middle sporulation elements (MSE) 0.8806 0.0003 YPL170W Heme‐binding protein involved in regulation of cytochrome P450 protein Erg11p; damage response protein, related to mammalian membrane progesterone receptors; mutations lead to defects in telomeres, mitochondria, and sterol synthesis 0.8762 0.0001 RLF2 YPR018W Largest subunit (p90) of the Chromatin Assembly Complex (CAF‐1) with Cac2p and Msi1p that assembles newly synthesized histones onto recently replicated DNA; involved in the maintenance of transcriptionally silent chromatin 0.8760 0.0002 CAB5 YDR196C Probable dephospho‐CoA kinase (DPCK) that catalyzes the last step in coenzyme A biosynthesis; null mutant lethality is complemented by E. coli coaE (encoding DPCK); detected in purified mitochondria in high‐throughput studies 0.8751 0.0002 COG8 YML071C Component of the conserved oligomeric Golgi complex (Cog1p through Cog8p), a cytosolic tethering complex that functions in protein trafficking to mediate fusion of transport vesicles to Golgi compartments 0.8701 0.0001 DAL2 YIR029W Allantoicase, converts allantoate to urea and ureidoglycolate in the second step of allantoin degradation; expression sensitive to nitrogen catabolite repression and induced by allophanate, an intermediate in allantoin degradation 0.8698 0.0001 DUR3 YHL016C Plasma membrane transporter for both urea and polyamines, expression is highly sensitive to nitrogen catabolite repression and induced by allophanate, the last intermediate of the allantoin degradative pathway 0.8693 0.0001 BCS1 YDR375C Mitochondrial protein of the AAA ATPase family; has ATP‐ dependent chaperone activity; required for assembly of Rip1p and Qcr10p into cytochrome bc(1) complex; mutations in human homolog BCS1L are linked to neonatal mitochondrial diseases 0.8677 0.0002 SNA4 YDL123W 0.8662 0.0002 MRPL33 YAE1 YMR286W YJR067C 0.8660 0.8654 0.0002 0.0002 SKP1 YDR328C Protein of unknown function, localized to the vacuolar outer membrane; predicted to be palmitoylated Mitochondrial ribosomal protein of the large subunit Protein of unknown function, essential for growth under standard (aerobic) conditions but not under anaerobic conditions Evolutionarily conserved kinetochore protein that is part of multiple protein complexes, including the SCF ubiquitin ligase complex, the CBF3 complex that binds centromeric DNA, and the RAVE complex that regulates assembly of the V‐ATPase 0.8642 0.0001 NA YML083C Putative protein of unknown function; strong increase in transcript abundance during anaerobic growth compared to aerobic growth; cells deleted for YML083C do not exhibit growth defects in anerobic or anaerobic conditions 0.8642 0.0006 SIR4 YDR227W Silent information regulator that, together with SIR2 and SIR3, is involved in assembly of silent chromatin domains at telomeres and the silent mating‐type loci; potentially phosphorylated by Cdc28p; some alleles of SIR4 prolong lifespan 0.8635 0.0001 TIM8 YJR135W‐A Mitochondrial intermembrane space protein, forms a complex with Tim13p that delivers a subset of hydrophobic proteins to the TIM22 complex for inner membrane insertion; homolog of human TIMM8A, implicated in Mohr‐Tranebjaerg syndrome 0.8624 0.0004 Symbols ORF Description NDT80 YHR124W DAP1 234 log2(mtr1 0.Ts/WT.T s) adj.p.val Conserved nuclear protein that interacts with GTP‐Gsp1p, which is a Ran homolog of the Ras GTPase family, and stimulates nucleotide release, involved in nuclear protein import, nucleotide release is inhibited by Yrb1p 0.8607 0.0002 YDR295C Subunit of a possibly tetrameric trichostatin A‐sensitive class II histone deacetylase complex containing an Hda1p homodimer and an Hda2p‐Hda3p heterodimer; involved in telomere maintenance 0.8599 0.0001 SMC3 YJL074C Subunit of the multiprotein cohesin complex required for sister chromatid cohesion in mitotic cells; also required, with Rec8p, for cohesion and recombination during meiosis; phylogenetically conserved SMC chromosomal ATPase family member 0.8591 0.0001 ZIM17 YNL310C Heat shock protein with a zinc finger motif; essential for protein import into mitochondria; may act with Pam18p to facilitate recognition and folding of imported proteins by Ssc1p (mtHSP70) in the mitochondrial matrix 0.8583 0.0001 PRP28 YDR243C 0.8575 0.0001 PEX29 YDR479C RNA helicase in the DEAD‐box family, involved in RNA isomerization at the 5' splice site Peroxisomal integral membrane peroxin, involved in the regulation of peroxisomal size, number and distribution; genetic interactions suggest that Pex28p and Pex29p act at steps upstream of those mediated by Pex30p, Pex31p, and Pex32p 0.8559 0.0004 GPI8 YDR331W ER membrane glycoprotein subunit of the glycosylphosphatidylinositol transamidase complex that adds glycosylphosphatidylinositol (GPI) anchors to newly synthesized proteins; human PIG‐K protein is a functional homolog 0.8556 0.0002 YRB1 YDR002W Ran GTPase binding protein; involved in nuclear protein import and RNA export, ubiquitin‐mediated protein degradation during the cell cycle; shuttles between the nucleus and cytoplasm; is essential; homolog of human RanBP1 0.8546 0.0001 TGL2 YDR058C Triacylglycerol lipase that is localized to the mitochondria; has lipolytic activity towards triacylglycerols and diacylglycerols when expressed in E. coli 0.8540 0.0003 RPN6 YDL097C Essential, non‐ATPase regulatory subunit of the 26S proteasome lid required for the assembly and activity of the 26S proteasome; the human homolog (S9 protein) partially rescues Rpn6p depletion 0.8524 0.0001 EAF6 YJR082C Subunit of the NuA4 acetyltransferase complex that acetylates histone H4 and NuA3 acetyltransferase complex that acetylates histone H3 0.8514 0.0007 FCF2 YLR051C Essential nucleolar protein involved in the early steps of 35S rRNA processing; interacts with Faf1p; member of a transcriptionally co‐ regulated set of genes called the RRB regulon 0.8499 0.0007 NA YDR003W‐A 0.8497 0.0006 FRQ1 YDR373W Putative protein of unknown function; identified by expression profiling and mass spectrometry N‐myristoylated calcium‐binding protein that may have a role in intracellular signaling through its regulation of the phosphatidylinositol 4‐kinase Pik1p; member of the recoverin/frequenin branch of the EF‐hand superfamily 0.8497 0.0001 XRS2 YDR369C Protein required for DNA repair; component of the Mre11 complex, which is involved in double strand breaks, meiotic recombination, telomere maintenance, and checkpoint signaling 0.8488 0.0002 Symbols ORF Description MOG1 YJR074W HDA2 235 log2(mtr1 0.Ts/WT.T s) adj.p.val Component of U1 snRNP required for mRNA splicing via spliceosome; yeast specific, no metazoan counterpart; interacts with mRNA in commitment complex 0.8478 0.0001 NA YIL169C NA Putative protein of unknown function; serine/threonine rich and highly similar to YOL155C, a putative glucan alpha‐1,4‐glucosidase; transcript is induced in both high and low pH environments; YIL169C is a non‐essential gene 0.8457 0.8448 0.0003 0.0003 RBS1 YDL189W Protein of unknown function, identified as a high copy suppressor of psk1 psk2 mutations that confer temperature‐sensitivity for galactose utilization; proposed to bind single‐stranded nucleic acids via its R3H domain 0.8436 0.0001 SIS1 YNL007C Type II HSP40 co‐chaperone that interacts with the HSP70 protein Ssa1p; not functionally redundant with Ydj1p due to due to substrate specificity; shares similarity with bacterial DnaJ proteins 0.8408 0.0001 RTT107 YHR154W Protein implicated in Mms22‐dependent DNA repair during S phase, DNA damage induces phosphorylation by Mec1p at one or more SQ/TQ motifs; interacts with Mms22p and Slx4p; has four BRCT domains; has a role in regulation of Ty1 transposition 0.8398 0.0008 KIN28 YDL108W Serine/threonine protein kinase, subunit of the transcription factor TFIIH; involved in transcription initiation at RNA polymerase II promoters 0.8393 0.0001 NA YCR102C Putative protein of unknown function, involved in copper metabolism; similar to C. carbonum toxD gene; member of the quinone oxidoreductase family 0.8392 0.0003 NA Putative protein of unknown function 0.8378 0.0006 CDC21 YMR030W‐ A YOR074C Thymidylate synthase, required for de novo biosynthesis of pyrimidine deoxyribonucleotides; expression is induced at G1/S 0.8367 0.0004 MIM1 YOL026C Mitochondrial outer membrane protein, required for assembly of the translocase of the outer membrane (TOM) complex and thereby for mitochondrial protein import; N terminus is exposed to the cytosol: transmembrane segment is highly conserved 0.8354 0.0001 MRP1 YDR347W Mitochondrial ribosomal protein of the small subunit; MRP1 exhibits genetic interactions with PET122, encoding a COX3‐specific translational activator, and with PET123, encoding a small subunit mitochondrial ribosomal protein 0.8317 0.0002 TAF7 YMR227C 0.8290 0.0003 IWR1 YDL115C TFIID subunit (67 kDa), involved in RNA polymerase II transcription initiation Protein involved in both basal and regulated transcription from RNA polymerase II (RNAP II) promoters; interacts with most of the RNAP II subunits; nucleo‐cytoplasmic shuttling protein; deletion causes hypersensitivity to K1 killer toxin 0.8280 0.0001 YCF1 YDR135C Vacuolar glutathione S‐conjugate transporter of the ATP‐binding cassette family, has a role in detoxifying metals such as cadmium, mercury, and arsenite; also transports unconjugated bilirubin; similar to human cystic fibrosis protein CFTR 0.8248 0.0002 VAM6 YDL077C Vacuolar protein that plays a critical role in the tethering steps of vacuolar membrane fusion by facilitating guanine nucleotide exchange on small guanosine triphosphatase Ypt7p 0.8236 0.0003 Symbols ORF Description SNU56 YDR240C NA NA 236 log2(mtr1 0.Ts/WT.T s) adj.p.val Protein involved in bud‐site selection and required for axial budding pattern; localizes with septins to bud neck in mitosis and may constitute an axial landmark for next round of budding 0.8235 0.0002 YNL166C Protein involved in organization of septins at the mother‐bud neck, may interact directly with the Cdc11p septin, localizes to bud neck in a septin‐dependent manner 0.8230 0.0001 CBS2 YDR197W Mitochondrial translational activator of the COB mRNA; interacts with translating ribosomes, acts on the COB mRNA 5'‐untranslated leader 0.8212 0.0001 FPR2 YDR519W Membrane‐bound peptidyl‐prolyl cis‐trans isomerase (PPIase), binds to the drugs FK506 and rapamycin; expression pattern suggests possible involvement in ER protein trafficking 0.8184 0.0002 NA YDL073W 0.8161 0.0002 PAU8 YAL068C 0.8158 0.0002 DIF1 YLR437C Putative protein of unknown function; YDL073W is not an essential gene Protein of unknown function, member of the seripauperin multigene family encoded mainly in subtelomeric regions Protein that regulates the nuclear localization of ribonucleotide reductase Rnr2p and Rnr4p subunits; phosphorylated by Dun1p in response to DNA damage and degraded; N‐terminal half has similarity to S. pombe Spd1 protein 0.8156 0.0002 RPN9 YDR427W Non‐ATPase regulatory subunit of the 26S proteasome, has similarity to putative proteasomal subunits in other species; null mutant is temperature sensitive and exhibits cell cycle and proteasome assembly defects 0.8151 0.0002 SDH4 YDR178W Membrane anchor subunit of succinate dehydrogenase (Sdh1p, Sdh2p, Sdh3p, Sdh4p), which couples the oxidation of succinate to the transfer of electrons to ubiquinone as part of the TCA cycle and the mitochondrial respiratory chain 0.8145 0.0003 CHA1 YCL064C Catabolic L‐serine (L‐threonine) deaminase, catalyzes the degradation of both L‐serine and L‐threonine; required to use serine or threonine as the sole nitrogen source, transcriptionally induced by serine and threonine 0.8137 0.0003 PST1 YDR055W Cell wall protein that contains a putative GPI‐attachment site; secreted by regenerating protoplasts; up‐regulated by activation of the cell integrity pathway, as mediated by Rlm1p; upregulated by cell wall damage via disruption of FKS1 0.8131 0.0008 SAN1 YDR143C 0.8129 0.0004 NA YDR319C Ubiquitin‐protein ligase, involved in the proteasome‐dependent degradation of aberrant nuclear proteins; contains intrinsically disordered regions that contribute to substrate recognition Putative protein of unknown function, identified as an ortholog of the highly conserved FIT family of proteins involved in triglyceride droplet biosynthesis; interacts with Sst2p and Hsp82p in high‐ throughput two‐hybrid screens 0.8108 0.0001 TBS1 YBR150C Putative protein of unknown function; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐ throughput studies 0.8096 0.0009 PSF1 YDR013W Subunit of the GINS complex (Sld5p, Psf1p, Psf2p, Psf3p), which is localized to DNA replication origins and implicated in assembly of the DNA replication machinery 0.8095 0.0001 MPP6 YNR024W Nuclear exosome‐associated RNA binding protein; involved in surveillance of pre‐rRNAs and pre‐mRNAs, and the degradation of cryptic non‐coding RNAs (ncRNA); copurifies with ribosomes 0.8084 0.0003 Symbols ORF Description BUD3 YCL014W BNI5 237 log2(mtr1 0.Ts/WT.T s) adj.p.val Peptidyl‐prolyl cis‐trans isomerase (cyclophilin), catalyzes the cis‐ trans isomerization of peptide bonds N‐terminal to proline residues; binds to Hsp82p and contributes to chaperone activity 0.8053 0.0003 YMR273C Protein with a role in regulating Swe1p‐dependent polarized growth; interacts with silencing proteins at the telomere; has a role in Bcy1p localization; implicated in mRNA nuclear export; involved in mitotic exit through Cdc14p regulation 0.8035 0.0004 GIC2 YDR309C Redundant rho‐like GTPase Cdc42p effector; homolog of Gic1p; involved in initiation of budding and cellular polarization; interacts with Cdc42p via the Cdc42/Rac‐interactive binding (CRIB) domain and with PI(4,5)P2 via a polybasic region 0.8020 0.0001 FYV6 YNL133C Protein of unknown function, required for survival upon exposure to K1 killer toxin; proposed to regulate double‐strand break repair via non‐homologous end‐joining 0.8011 0.0002 YKE2 YLR200W Subunit of the heterohexameric Gim/prefoldin protein complex involved in the folding of alpha‐tubulin, beta‐tubulin, and actin 0.8010 0.0004 SYF1 YDR416W Member of the NineTeen Complex (NTC) that contains Prp19p and stabilizes U6 snRNA in catalytic forms of the spliceosome containing U2, U5, and U6 snRNAs; null mutant has splicing defect and arrests in G2/M; homologs in human and C. elegans 0.7998 0.0002 IES2 YNL215W Protein that associates with the INO80 chromatin remodeling complex under low‐salt conditions; essential for growth under anaerobic conditions 0.7976 0.0002 GCS1 YDL226C ADP‐ribosylation factor GTPase activating protein (ARF GAP), involved in ER‐Golgi transport; shares functional similarity with Glo3p 0.7975 0.0004 MRPL37 ASP1 YBR268W YDR321W Mitochondrial ribosomal protein of the large subunit Cytosolic L‐asparaginase, involved in asparagine catabolism 0.7958 0.7957 0.0003 0.0006 PHO2 YDL106C Homeobox transcription factor; regulatory targets include genes involved in phosphate metabolism; binds cooperatively with Pho4p to the PHO5 promoter; phosphorylation of Pho2p facilitates interaction with Pho4p 0.7956 0.0006 NA YOR389W 0.7938 0.0005 NA KCS1 YDR415C YDR017C Putative protein of unknown function; expression regulated by copper levels Putative protein of unknown function Inositol hexakisphosphate (IP6) and inositol heptakisphosphate (IP7) kinase; generation of high energy inositol pyrophosphates by Kcs1p is required for many processes such as vacuolar biogenesis, stress response and telomere maintenance 0.7904 0.7900 0.0004 0.0006 PFD1 YJL179W Subunit of heterohexameric prefoldin, which binds cytosolic chaperonin and transfers target proteins to it; involved in the biogenesis of actin and of alpha‐ and gamma‐tubulin 0.7899 0.0002 RNH70 YGR276C 0.7897 0.0004 PMS1 YNL082W 3'‐5' exoribonuclease; required for maturation of 3' ends of 5S rRNA and tRNA‐Arg3 from dicistronic transcripts ATP‐binding protein required for mismatch repair in mitosis and meiosis; functions as a heterodimer with Mlh1p, binds double‐ and single‐stranded DNA via its N‐terminal domain, similar to E. coli MutL 0.7893 0.0003 Symbols ORF Description CPR7 YJR032W ZDS1 238 log2(mtr1 0.Ts/WT.T s) adj.p.val Protein involved in minichromosome maintenance; component of the COMA complex (Ctf19p, Okp1p, Mcm21p, Ame1p) that bridges kinetochore subunits that are in contact with centromeric DNA and the subunits bound to microtubules 0.7881 0.0004 YHR129C Actin‐related protein of the dynactin complex; required for spindle orientation and nuclear migration; putative ortholog of mammalian centractin 0.7879 0.0006 RFA3 YJL173C Subunit of heterotrimeric Replication Protein A (RPA), which is a highly conserved single‐stranded DNA binding protein involved in DNA replication, repair, and recombination 0.7878 0.0002 CPR5 YDR304C Peptidyl‐prolyl cis‐trans isomerase (cyclophilin) of the endoplasmic reticulum, catalyzes the cis‐trans isomerization of peptide bonds N‐ terminal to proline residues; transcriptionally induced in response to unfolded proteins in the ER 0.7860 0.0001 RAM1 YDL090C Beta subunit of the CAAX farnesyltransferase (FTase) that prenylates the a‐factor mating pheromone and Ras proteins; required for the membrane localization of Ras proteins and a‐factor; homolog of the mammalian FTase beta subunit 0.7854 0.0002 NA OST4 NA YDL232W NA Subunit of the oligosaccharyltransferase complex of the ER lumen, which catalyzes protein asparagine‐linked glycosylation; type I membrane protein required for incorporation of Ost3p or Ost6p into the OST complex 0.7854 0.7845 0.0004 0.0001 GRX1 YCL035C Hydroperoxide and superoxide‐radical responsive heat‐stable glutathione‐dependent disulfide oxidoreductase with active site cysteine pair; protects cells from oxidative damage 0.7839 0.0009 HST4 YDR191W Member of the Sir2 family of NAD(+)‐dependent protein deacetylases; involved along with Hst3p in silencing at telomeres, cell cycle progression, radiation resistance, genomic stability and short‐chain fatty acid metabolism 0.7830 0.0003 SNA2 YDR525W‐A Protein of unknown function, has similarity to Pmp3p, which is involved in cation transport; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm in a punctate pattern 0.7827 0.0002 NUM1 YDR150W Protein required for nuclear migration, localizes to the mother cell cortex and the bud tip; may mediate interactions of dynein and cytoplasmic microtubules with the cell cortex 0.7800 0.0002 SLX5 YDL013W Subunit of the Slx5‐Slx8 SUMO‐targeted ubiquitin ligase (STUbL) complex, stimulated by SUMO‐modified substrates; contains a RING domain and two SIMs (SUMO‐interacting motifs); forms SUMO‐ dependent nuclear foci, including DNA repair centers 0.7800 0.0002 SHU2 YDR078C Protein involved in a Rad51p‐, Rad54p‐dependent pathway for homologous recombination repair, important for error‐free repair of spontaneous and induced DNA lesions to protect the genome from mutation; associates with Shu1p, Psy3p, and Csm2p 0.7789 0.0002 UBC13 YDR092W Ubiquitin‐conjugating enzyme involved in the error‐free DNA postreplication repair pathway; interacts with Mms2p to assemble ubiquitin chains at the Ub Lys‐63 residue; DNA damage triggers redistribution from the cytoplasm to the nucleus 0.7785 0.0001 CUS2 YNL286W Protein that binds to U2 snRNA and Prp11p, may be involved in U2 snRNA folding; contains two RNA recognition motifs (RRMs) 0.7766 0.0003 Symbols ORF Description MCM21 YDR318W ARP1 239 log2(mtr1 0.Ts/WT.T s) adj.p.val Protein with similarity to ATP‐binding cassette (ABC) transporter family members; lacks predicted membrane‐spanning regions; transcriptionally activated by Yrm1p along with genes involved in multidrug resistance 0.7763 0.0002 YDR128W Protein of unknown function; mtc5 is synthetically sick with cdc13‐1 0.7763 0.0001 GPI17 YDR434W Transmembrane protein subunit of the glycosylphosphatidylinositol transamidase complex that adds GPIs to newly synthesized proteins; human PIG‐Sp homolog 0.7762 0.0008 GIM5 YML094W Subunit of the heterohexameric cochaperone prefoldin complex which binds specifically to cytosolic chaperonin and transfers target proteins to it 0.7755 0.0002 SPS1 YDR523C Putative protein serine/threonine kinase expressed at the end of meiosis and localized to the prospore membrane, required for correct localization of enzymes involved in spore wall synthesis 0.7754 0.0008 FMP10 YER182W Putative protein of unknown function; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐ throughput studies 0.7751 0.0007 NA YDR026C Protein of unknown function that may interact with ribosomes, based on co‐purification experiments; Myb‐like DNA‐binding protein that may bind to the Ter region of rDNA; interacts physically with Fob1p 0.7749 0.0002 NA HIF1 YDR444W YLL022C Putative protein of unknown function Non‐essential component of the HAT‐B histone acetyltransferase complex (Hat1p‐Hat2p‐Hif1p), localized to the nucleus; has a role in telomeric silencing 0.7732 0.7725 0.0007 0.0003 UMP1 YBR173C Short‐lived chaperone required for correct maturation of the 20S proteasome; may inhibit premature dimerization of proteasome half‐mers; degraded by proteasome upon completion of its assembly 0.7723 0.0002 SWR1 YDR334W Swi2/Snf2‐related ATPase that is the structural component of the SWR1 complex, which exchanges histone variant H2AZ (Htz1p) for chromatin‐bound histone H2A 0.7709 0.0004 TRM9 YML014W tRNA methyltransferase, catalyzes esterification of modified uridine nucleotides in tRNA(Arg3) and tRNA(Glu), likely as part of a complex with Trm112p; deletion confers resistance to zymocin 0.7693 0.0006 VTA1 YLR181C Multivesicular body (MVB) protein involved in endosomal protein sorting; regulates Vps4p activity by promoting its oligomerization; has an N‐terminal Vps60‐ and Did2‐ binding domain, a linker region, and a C‐terminal Vps4p binding domain 0.7691 0.0003 SRP14 YDL092W Signal recognition particle (SRP) subunit, interacts with the RNA component of SRP to form the Alu domain, which is the region of SRP responsible for arrest of nascent chain elongation during membrane targeting; homolog of mammalian SRP14 0.7685 0.0003 MRPL44 NA PSH1 YMR225C YDR109C YOL054W Mitochondrial ribosomal protein of the large subunit Putative kinase E3 ubiquitin ligase that mediates poyubiquitination and degradation of centromere‐binding protein Cse4p and prevents Cse4p from mislocalizing to euchromatin; ubiquitylation of Cse4p may be antagonized by Scm3p 0.7677 0.7675 0.7665 0.0004 0.0001 0.0003 NA YLR099W‐A Putative protein of unknown function 0.7665 0.0003 Symbols ORF Description NA YDR061W MTC5 240 log2(mtr1 0.Ts/WT.T s) adj.p.val Component of the SF3b subcomplex of the U2 snRNP, essential protein required for for splicing and for assembly of SF3b 0.7634 0.0003 YDR226W Adenylate kinase, required for purine metabolism; localized to the cytoplasm and the mitochondria; lacks cleavable signal sequence 0.7628 0.0001 PCF11 YDR228C mRNA 3' end processing factor, essential component of cleavage and polyadenylation factor IA (CF IA), involved in pre‐mRNA 3' end processing and in transcription termination; binds C‐terminal domain of largest subunit of RNA pol II (Rpo21p) 0.7620 0.0001 PKH3 YDR466W Protein kinase with similarity to mammalian phosphoinositide‐ dependent kinase 1 (PDK1) and yeast Pkh1p and Pkh2p, two redundant upstream activators of Pkc1p; identified as a multicopy suppressor of a pkh1 pkh2 double mutant 0.7617 0.0003 VPS3 YDR495C Component of CORVET tethering complex; cytoplasmic protein required for the sorting and processing of soluble vacuolar proteins, acidification of the vacuolar lumen, and assembly of the vacuolar H+‐ATPase 0.7615 0.0006 PPH22 YDL188C Catalytic subunit of protein phosphatase 2A (PP2A), functionally redundant with Pph21p; methylated at C terminus; forms alternate complexes with several regulatory subunits; involved in signal transduction and regulation of mitosis 0.7611 0.0006 CTF8 YHR191C Subunit of a complex with Ctf18p that shares some subunits with Replication Factor C and is required for sister chromatid cohesion 0.7600 0.0002 DPM1 YPR183W Dolichol phosphate mannose (Dol‐P‐Man) synthase of the ER membrane, catalyzes the formation of Dol‐P‐Man from Dol‐P and GDP‐Man; required for glycosyl phosphatidylinositol membrane anchoring, O mannosylation, and protein glycosylation 0.7600 0.0002 TIM11 YDR322C‐A Subunit e of mitochondrial F1F0‐ATPase, which is a large, evolutionarily conserved enzyme complex required for ATP synthesis; essential for the dimeric and oligomeric state of ATP synthase 0.7569 0.0001 MSA2 YKR077W Putative transcriptional activator, that interacts with G1‐specific transcription factor, MBF and G1‐specific promoters; ortholog of Msa2p, an MBF and SBF activator that regulates G1‐specific transcription and cell cycle initiation 0.7569 0.0005 YOS1 YER074W‐A Integral membrane protein required for ER to Golgi transport; localized to the Golgi, the ER, and COPII vesicles; interacts with Yip1p and Yif1p 0.7552 0.0002 RSM28 YDR494W Mitochondrial ribosomal protein of the small subunit; genetic interactions suggest a possible role in promoting translation initiation 0.7534 0.0002 NA YPL071C Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to both the cytoplasm and the nucleus 0.7522 0.0004 IPK1 YDR315C Inositol 1,3,4,5,6‐pentakisphosphate 2‐kinase, nuclear protein required for synthesis of 1,2,3,4,5,6‐hexakisphosphate (phytate), which is integral to cell function; has 2 motifs conserved in other fungi; ipk1 gle1 double mutant is inviable 0.7518 0.0007 MFB1 YDR219C Mitochondria‐associated F‐box protein involved in maintenance of normal mitochondrial morphology; interacts with Skp1p through the F‐box motif; preferentially localizes to the mother cell during budding 0.7492 0.0004 Symbols ORF Description YSF3 YNL138W‐A ADK1 241 log2(mtr1 0.Ts/WT.T s) adj.p.val Sulfiredoxin, contributes to oxidative stress resistance by reducing cysteine‐sulfinic acid groups in the peroxiredoxin Tsa1p, which is formed upon exposure to oxidants; conserved in higher eukaryotes 0.7469 0.0004 YLL021W Component of the polarisome, which functions in actin cytoskeletal organization during polarized growth; acts as a scaffold for Mkk1p and Mpk1p cell wall integrity signaling components; potential Cdc28p substrate 0.7452 0.0003 RPN5 YDL147W Essential, non‐ATPase regulatory subunit of the 26S proteasome lid, similar to mammalian p55 subunit and to another S. cerevisiae regulatory subunit, Rpn7p 0.7425 0.0005 RPC10 YHR143W‐A 0.7423 0.0006 NBP2 YDR162C RNA polymerase subunit ABC10‐alpha, found in RNA polymerase complexes I, II, and III Protein involved in the HOG (high osmolarity glycerol) pathway, negatively regulates Hog1p by recruitment of phosphatase Ptc1p the Pbs2p‐Hog1p complex, found in the nucleus and cytoplasm, contains an SH3 domain that binds Pbs2p 0.7419 0.0002 ELF1 YKL160W Transcription elongation factor that contains a conserved zinc finger domain; implicated in the maintenance of proper chromatin structure in actively transcribed regions; deletion inhibits Brome mosaic virus (BMV) gene expression 0.7394 0.0002 GNA1 YFL017C Evolutionarily conserved glucosamine‐6‐phosphate acetyltransferase required for multiple cell cycle events including passage through START, DNA synthesis, and mitosis; involved in UDP‐N‐acetylglucosamine synthesis, forms GlcNAc6P from AcCoA 0.7393 0.0001 RAD30 YDR419W DNA polymerase eta, involved in translesion synthesis during post‐ replication repair; catalyzes the synthesis of DNA opposite cyclobutane pyrimidine dimers and other lesions; mutations in human pol eta are responsible for XPV 0.7390 0.0002 NA YDL144C Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm and nucleus; YDL144C is not an essential gene. 0.7381 0.0002 NA YOR385W Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm; YOR385W is not an essential gene 0.7376 0.0002 NA YDR338C Putative protein of unknown function, member of the multi‐drug and toxin extrusion (MATE) family of the multidrug/oligosaccharidyl‐ lipid/polysaccharide (MOP) exporter superfamily 0.7371 0.0006 AIR2 YDL175C Zinc knuckle protein, involved in nuclear RNA processing and degredation as a component of the TRAMP complex; stimulates the poly(A) polymerase activity of Pap2p in vitro; functionally redundant with Air1p 0.7362 0.0003 CYK3 YDL117W SH3‐domain protein located in the mother‐bud neck and the cytokinetic actin ring; mutant phenotype and genetic interactions suggest a role in cytokinesis 0.7350 0.0001 SPO71 YDR104C Meiosis‐specific protein of unknown function, required for spore wall formation during sporulation; dispensable for both nuclear divisions during meiosis 0.7347 0.0001 NA NA NA YDR381C‐A NA Protein of unknown function, localized to the mitochondrial outer membrane 0.7347 0.7286 0.0007 0.0003 Symbols ORF Description SRX1 YKL086W SPA2 242 log2(mtr1 0.Ts/WT.T s) adj.p.val ADP‐ribosylation factor, GTPase of the Ras superfamily involved in regulation of coated formation vesicles in intracellular trafficking within the Golgi; functionally interchangeable with Arf1p 0.7280 0.0003 YDR179W‐A NA YMR305C Putative protein of unknown function NA Cell wall protein with similarity to glucanases; may play a role in conjugation during mating based on mutant phenotype and its regulation by Ste12p 0.7263 0.7245 0.7244 0.0005 0.0004 0.0001 LEO1 YOR123C Component of the Paf1 complex, which associates with RNA polymerase II and is involved in histone methylation; plays a role in regulating Ty1 transposition 0.7208 0.0003 NA YBL059W Putative protein of unknown function; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐ throughput studies 0.7196 0.0005 NA NA NA YIL161W NA Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm; mRNA is enriched in Scp160p‐associated mRNPs; YIL161W is a non‐essential gene 0.7188 0.7183 0.0007 0.0002 VPS72 YDR485C Htz1p‐binding component of the SWR1 complex, which exchanges histone variant H2AZ (Htz1p) for chromatin‐bound histone H2A; required for vacuolar protein sorting 0.7163 0.0003 DIG2 YDR480W MAP kinase‐responsive inhibitor of the Ste12p transcription factor, involved in the regulation of mating‐specific genes and the invasive growth pathway; related regulators Dig1p and Dig2p bind to Ste12p 0.7151 0.0004 MRPL11 SRM1 YDL202W YGL097W Mitochondrial ribosomal protein of the large subunit Nucleotide exchange factor for Gsp1p, localizes to the nucleus, required for nucleocytoplasmic trafficking of macromolecules; suppressor of the pheromone response pathway; potentially phosphorylated by Cdc28p 0.7145 0.7129 0.0002 0.0008 FMC1 YIL098C Mitochondrial matrix protein, required for assembly or stability at high temperature of the F1 sector of mitochondrial F1F0 ATP synthase; null mutant temperature sensitive growth on glycerol is suppressed by multicopy expression of Odc1p 0.7106 0.0001 AAD4 YDL243C Putative aryl‐alcohol dehydrogenase with similarity to P. chrysosporium aryl‐alcohol dehydrogenase, involved in the oxidative stress response; expression induced in cells treated with the mycotoxin patulin 0.7101 0.0004 NA YEL073C Putative protein of unknown function; located adjacent to ARS503 and the telomere on the left arm of chromosome V; regulated by inositol/choline 0.7079 0.0004 GTB1 YDR221W Glucosidase II beta subunit, forms a complex with alpha subunit Rot2p, involved in removal of two glucose residues from N‐linked glycans during glycoprotein biogenesis in the ER 0.7077 0.0004 RDH54 YBR073W DNA‐dependent ATPase, stimulates strand exchange by modifying the topology of double‐stranded DNA; involved in recombinational repair of DNA double‐strand breaks during mitosis and meiosis; proposed to be involved in crossover interference 0.7068 0.0010 SMB1 YER029C Core Sm protein Sm B; part of heteroheptameric complex (with Smd1p, Smd2p, Smd3p, Sme1p, Smx3p, and Smx2p) that is part of the spliceosomal U1, U2, U4, and U5 snRNPs; homolog of human Sm B and Sm B' 0.7043 0.0003 Symbols ORF Description ARF2 YDL137W NA NA SCW10 243 log2(mtr1 0.Ts/WT.T s) adj.p.val Putative protein of unknown function, deletion confers reduced fitness in saline Plasma membrane low glucose sensor that regulates glucose transport; contains 12 predicted transmembrane segments and a long C‐terminal tail required for induction of hexose transporters; also senses fructose and mannose; similar to Rgt2p 0.7042 0.0006 0.7036 0.0003 YDR006C Protein whose overexpression suppresses the growth defect of mutants lacking protein kinase A activity; involved in cAMP‐ mediated signaling; localized to the nucleus; similar to the mouse testis‐specific protein PBS13 0.7025 0.0002 SAC3 YDR159W Nuclear pore‐associated protein, required for biogenesis of the small ribosomal subunit; forms a complex with Thp1p that is involved in transcription and in mRNA export from the nucleus 0.7015 0.0002 YPS7 YDR349C Putative GPI‐anchored aspartic protease, member of the yapsin family of proteases involved in cell wall growth and maintenance; located in the cytoplasm and endoplasmic reticulum 0.6982 0.0001 STP1 YDR463W Transcription factor, undergoes proteolytic processing by SPS (Ssy1p‐Ptr3p‐Ssy5p)‐sensor component Ssy5p in response to extracellular amino acids; activates transcription of amino acid permease genes and may have a role in tRNA processing 0.6970 0.0003 NA YKL018C‐A Putative protein of unknown function; identified by homology; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm 0.6955 0.0005 NA YMR114C Protein of unknown function; may interact with ribosomes, based on co‐purification experiments; green fluorescent protein (GFP)‐ fusion protein localizes to the nucleus and cytoplasm; YMR114C is not an essential gene 0.6948 0.0009 NA YOL114C Putative protein of unknown function with similarity to human ICT1 and prokaryotic factors that may function in translation termination; YOL114C is not an essential gene 0.6945 0.0004 CLB3 YDL155W B‐type cyclin involved in cell cycle progression; activates Cdc28p to promote the G2/M transition; may be involved in DNA replication and spindle assembly; accumulates during S phase and G2, then targeted for ubiquitin‐mediated degradation 0.6924 0.0002 TIF11 YMR260C Translation initiation factor eIF1A, essential protein that forms a complex with Sui1p (eIF1) and the 40S ribosomal subunit and scans for the start codon; C‐terminus associates with Fun12p (eIF5B); N terminus interacts with eIF2 and eIF3 0.6888 0.0006 SRP101 YDR292C Signal recognition particle (SRP) receptor alpha subunit; contain GTPase domains; involved in SRP‐dependent protein targeting; interacts with the beta subunit, Srp102p 0.6882 0.0003 NA CLN2 YDR307W YPL256C Putative protein of unknown function G1 cyclin involved in regulation of the cell cycle; activates Cdc28p kinase to promote the G1 to S phase transition; late G1 specific expression depends on transcription factor complexes, MBF (Swi6p‐ Mbp1p) and SBF (Swi6p‐Swi4p) 0.6867 0.6842 0.0005 0.0003 MED11 YMR112C Subunit of the RNA polymerase II mediator complex; associates with core polymerase subunits to form the RNA polymerase II holoenzyme; essential protein 0.6826 0.0005 Symbols ORF Description NA YNR029C SNF3 YDL194W SOK1 244 log2(mtr1 0.Ts/WT.T s) adj.p.val Essential protein required for maturation of Gas1p and Pho8p; involved in protein trafficking; GFP‐fusion protein localizes to the ER and YFP‐fusion protein to the nuclear envelope‐ER network; null mutants have a cell separation defect 0.6819 0.0005 YDR326C Protein involved in programmed cell death; mutant shows resistance to cell death induced by amiodarone or intracellular acidification 0.6813 0.0006 SNT2 YGL131C DNA binding protein with similarity to the S. pombe Snt2 protein; computational analysis suggests a role in regulation of expression of genes encoding amine transporters 0.6811 0.0003 OCA6 YDR067C Cytoplasmic protein required for replication of Brome mosaic virus in S. cerevisiae, which is a model system for studying positive‐strand RNA virus replication; null mutation confers sensitivity to tunicamycin and DTT 0.6811 0.0002 FPR3 YML074C Nucleolar peptidyl‐prolyl cis‐trans isomerase (PPIase); FK506 binding protein; phosphorylated by casein kinase II (Cka1p‐Cka2p‐Ckb1p‐ Ckb2p) and dephosphorylated by Ptp1p 0.6804 0.0002 DBF4 YDR052C Regulatory subunit of Cdc7p‐Dbf4p kinase complex, required for Cdc7p kinase activity and initiation of DNA replication; phosphorylates the Mcm2‐7 family of proteins; cell cycle regulated 0.6798 0.0006 DSK2 YMR276W Nuclear‐enriched ubiquitin‐like polyubiquitin‐binding protein, required for spindle pole body (SPB) duplication and for transit through the G2/M phase of the cell cycle, involved in proteolysis, interacts with the proteasome 0.6792 0.0009 TAF12 YDR145W Subunit (61/68 kDa) of TFIID and SAGA complexes, involved in RNA polymerase II transcription initiation and in chromatin modification, similar to histone H2A 0.6775 0.0005 ACK1 YDL203C Protein that functions upstream of Pkc1p in the cell wall integrity pathway; GFP‐fusion protein expression is induced in response to the DNA‐damaging agent MMS; non‐tagged Ack1p is detected in purified mitochondria 0.6768 0.0003 NA TRX3 YGR121W‐A YCR083W Putative protein of unknown function Mitochondrial thioredoxin, highly conserved oxidoreductase required to maintain the redox homeostasis of the cell, forms the mitochondrial thioredoxin system with Trr2p, redox state is maintained by both Trr2p and Glr1p 0.6766 0.6736 0.0010 0.0003 MNN10 YDR245W Subunit of a Golgi mannosyltransferase complex also containing Anp1p, Mnn9p, Mnn11p, and Hoc1p that mediates elongation of the polysaccharide mannan backbone; membrane protein of the mannosyltransferase family 0.6725 0.0002 PCL7 YIL050W Pho85p cyclin of the Pho80p subfamily, forms a functional kinase complex with Pho85p which phosphorylates Mmr1p and is regulated by Pho81p; involved in glycogen metabolism, expression is cell‐cycle regulated 0.6708 0.0005 TPM1 YNL079C Major isoform of tropomyosin; binds to and stabilizes actin cables and filaments, which direct polarized cell growth and the distribution of several organelles; acetylated by the NatB complex and acetylated form binds actin most efficiently 0.6704 0.0005 TRM3 YDL112W 2'‐O‐ribose methyltransferase, catalyzes the ribose methylation of the guanosine nucleotide at position 18 of tRNAs 0.6703 0.0005 Symbols ORF Description PGA2 YNL149C YSP2 245 log2(mtr1 0.Ts/WT.T s) adj.p.val Essential subunit of the Dam1 complex (aka DASH complex), couples kinetochores to the force produced by MT depolymerization thereby aiding in chromosome segregation; is transferred to the kinetochore prior to mitosis 0.6701 0.0003 YCR071C YLR246W Mitochondrial ribosomal protein of the large subunit Subunit of a palmitoyltransferase, composed of Erf2p and Shr5p, that adds a palmitoyl lipid moiety to heterolipidated substrates such as Ras1p and Ras2p through a thioester linkage; mutants partially mislocalize Ras2p to the vacuole 0.6699 0.6696 0.0006 0.0002 NTG2 YOL043C DNA N‐glycosylase and apurinic/apyrimidinic (AP) lyase involved in base excision repair, localizes to the nucleus; sumoylated 0.6687 0.0002 NA YLR460C 0.6678 0.0008 PEX5 YDR244W Member of the quinone oxidoreductase family, up‐regulated in response to the fungicide mancozeb; possibly up‐regulated by iodine Peroxisomal membrane signal receptor for the C‐terminal tripeptide signal sequence (PTS1) of peroxisomal matrix proteins, required for peroxisomal matrix protein import; also proposed to have PTS1‐ receptor independent functions 0.6657 0.0005 PKR1 YMR123W V‐ATPase assembly factor, functions with other V‐ATPase assembly factors in the ER to efficiently assemble the V‐ATPase membrane sector (V0) 0.6642 0.0006 RAV2 YDR202C 0.6642 0.0003 NA TMA64 NA YDR117C Subunit of RAVE (Rav1p, Rav2p, Skp1p), a complex that associates with the V1 domain of the vacuolar membrane (H+)‐ATPase (V‐ ATPase) and promotes assembly and reassembly of the holoenzyme NA Protein of unknown function that associates with ribosomes; has a putative RNA binding domain; in mammals the corresponding protein, eIF2D, has been shown to possess translation initiation factor activity 0.6639 0.6615 0.0007 0.0003 MRPL35 SPO13 YDR322W YHR014W Mitochondrial ribosomal protein of the large subunit Meiosis‐specific protein, involved in maintaining sister chromatid cohesion during meiosis I as well as promoting proper attachment of kinetochores to the spindle during meiosis I and meiosis II 0.6590 0.6582 0.0006 0.0009 NUP2 YLR335W Nucleoporin involved in nucleocytoplasmic transport, binds to either the nucleoplasmic or cytoplasmic faces of the nuclear pore complex depending on Ran‐GTP levels; also has a role in chromatin organization 0.6577 0.0006 PTR3 YFR029W Component of the SPS plasma membrane amino acid sensor system (Ssy1p‐Ptr3p‐Ssy5p), which senses external amino acid concentration and transmits intracellular signals that result in regulation of expression of amino acid permease genes 0.6571 0.0002 PRP11 YDL043C 0.6559 0.0005 NOP10 YHR072W‐A Subunit of the SF3a splicing factor complex, required for spliceosome assembly Constituent of small nucleolar ribonucleoprotein particles containing H/ACA‐type snoRNAs, which are required for pseudouridylation and processing of pre‐18S rRNA 0.6540 0.0007 SSY1 YDR160W Component of the SPS plasma membrane amino acid sensor system (Ssy1p‐Ptr3p‐Ssy5p), which senses external amino acid concentration and transmits intracellular signals that result in regulation of expression of amino acid permease genes 0.6535 0.0006 Symbols ORF Description DAD3 YBR233W‐A IMG2 ERF2 246 log2(mtr1 0.Ts/WT.T s) adj.p.val Rab family GTPase, involved in the ER‐to‐Golgi step of the secretory pathway; complex formation with the Rab escort protein Mrs6p is required for prenylation of Ypt1p by protein geranylgeranyltransferase type II (Bet2p‐Bet4p) 0.6532 0.0002 YGL061C Essential subunit of the Dam1 complex (aka DASH complex), couples kinetochores to the force produced by MT depolymerization thereby aiding in chromosome segregation; is transferred to the kinetochore prior to mitosis 0.6529 0.0003 SNA3 YJL151C Integral membrane protein localized to vacuolar intralumenal vesicles, computational analysis of large‐scale protein‐protein interaction data suggests a possible role in either cell wall synthesis or protein‐vacuolar targeting 0.6519 0.0002 FUN19 YAL034C 0.6513 0.0002 ECM18 BRE2 YDR125C YLR015W Non‐essential protein of unknown function; expression induced in response to heat stress Protein of unknown function, similar to Rlp24p Subunit of COMPASS (Set1C) complex, which methylates Lys4 of histone H3 and functions in silencing at telomeres; has a C‐terminal Sdc1 Dpy‐30 Interaction (SDI) domain that mediates binding to Sdc1p; similar to trithorax‐group protein ASH2L 0.6510 0.6506 0.0003 0.0007 UBC5 YDR059C Ubiquitin‐conjugating enzyme that mediates selective degradation of short‐lived, abnormal, or excess proteins, including histone H3; central component of the cellular stress response; expression is heat inducible 0.6492 0.0008 SLD2 YKL108W Protein required for DNA replication, phosphorylated in S phase by S‐phase cyclin‐dependent kinases (Cdks), phosphorylation is essential for DNA replication and for complex formation with Dpb11p; potential Cdc28p substrate 0.6456 0.0005 CDC48 YDL126C ATPase involved in ubiquitin‐mediated protein degradation; Cdc48p‐Npl4p‐Ufd1p complex participates in ER‐associated degradation (ERAD) while Cdc48p‐Npl4p‐Vms1p complex participates in mitochondria‐associated degradation (MAD) 0.6447 0.0004 NA YCR076C 0.6415 0.0006 CHO1 YER026C Putative protein of unknown function; YCR076C is not an essential gene Phosphatidylserine synthase, functions in phospholipid biosynthesis; catalyzes the reaction CDP‐diaclyglycerol + L‐serine = CMP + L‐1‐ phosphatidylserine, transcriptionally repressed by myo‐inositol and choline 0.6403 0.0002 CSR1 YLR380W Phosphatidylinositol transfer protein with a potential role in regulating lipid and fatty acid metabolism under heme‐depleted conditions; interacts specifically with thioredoxin peroxidase; may have a role in oxidative stress resistance 0.6401 0.0004 SLC1 YDL052C 1‐acyl‐sn‐glycerol‐3‐phosphate acyltransferase, catalyzes the acylation of lysophosphatidic acid to form phosphatidic acid, a key intermediate in lipid metabolism; enzymatic activity detected in lipid particles and microsomes 0.6386 0.0004 CKS1 YBR135W Cyclin‐dependent protein kinase regulatory subunit and adaptor; modulates proteolysis of M‐phase targets through interactions with the proteasome; role in transcriptional regulation, recruiting proteasomal subunits to target gene promoters 0.6365 0.0005 SIZ1 YDR409W SUMO/Smt3 ligase that promotes the attachment of sumo (Smt3p; small ubiquitin‐related modifier) to proteins; binds Ubc9p and may bind septins; specifically required for sumoylation of septins in vivo; localized to the septin ring 0.6361 0.0002 Symbols ORF Description YPT1 YFL038C DUO1 247 log2(mtr1 0.Ts/WT.T s) adj.p.val E3 ubiquitin ligase of the hect‐domain class; has a role in mRNA export from the nucleus and may regulate transcriptional coactivators; involved in degradation of excess histones 0.6355 0.0004 Phospholipid hydroperoxide glutathione peroxidase induced by glucose starvation that protects cells from phospholipid hydroperoxides and nonphospholipid peroxides during oxidative stress Dethiobiotin synthetase, catalyzes the third step in the biotin biosynthesis pathway; BIO4 is in a cluster of 3 genes (BIO3, BIO4, and BIO5) that mediate biotin synthesis; expression appears to be repressed at low iron levels 0.6329 0.0004 0.6329 0.0006 Mevalonate pyrophosphate decarboxylase, essential enzyme involved in the biosynthesis of isoprenoids and sterols, including ergosterol; acts as a homodimer 0.6312 0.0003 YDR391C Putative protein of unknown function, possibly involved in zinc homeostasis; Bdf1p‐dependent transcription induced by salt stress; green fluorescent protein (GFP)‐fusion protein localizes to both the cytoplasm and the nucleus 0.6311 0.0003 TRR1 YDR353W Cytoplasmic thioredoxin reductase, key regulatory enzyme that determines the redox state of the thioredoxin system, which acts as a disulfide reductase system and protects cells against both oxidative and reductive stress 0.6296 0.0003 HHO1 YPL127C Histone H1, a linker histone required for nucleosome packaging at restricted sites; suppresses DNA repair involving homologous recombination; not required for telomeric silencing, basal transcriptional repression, or efficient sporulation 0.6288 0.0005 IGO1 YNL157W Protein required for initiation of G0 program; prevents degradation of nutrient‐regulated mRNAs via the 5'‐3' mRNA decay pathway; phosphorylated by Rim15p; GFP protein localizes to the cytoplasm and nucleus; similar to Igo2p 0.6280 0.0004 MSS4 YDR208W Phosphatidylinositol‐4‐phosphate 5‐kinase, involved in actin cytoskeleton organization and cell morphogenesis; multicopy suppressor of stt4 mutation 0.6275 0.0006 NTR2 YKR022C Essential protein that forms a dimer with Ntr1p; also forms a trimer, with Ntr2p and the DExD/H‐box RNA helicase Prp43p, that is involved in spliceosome disassembly 0.6274 0.0003 GUK1 YDR454C Guanylate kinase, converts GMP to GDP; required for growth and mannose outer chain elongation of cell wall N‐linked glycoproteins 0.6263 0.0006 NA YDL119C Putative mitochondrial transport protein; GFP‐fusion protein is induced in response to the DNA‐damaging agent MMS; the authentic, non‐tagged protein is detected in purified mitochondria 0.6262 0.0003 RMP1 YLR145W Subunit of RNase MRP, which processes pre‐rRNA and has a role in cell cycle‐regulated degradation of daughter cell‐specific mRNAs; unlike most subunits, not shared between RNase MRP and nuclear RNase P 0.6259 0.0010 RPC17 YJL011C RNA polymerase III subunit C17; physically interacts with C31, C11, and TFIIIB70; may be involved in the recruitment of pol III by the preinitiation complex 0.6258 0.0010 Symbols ORF Description TOM1 YDR457W GPX2 YBR244W BIO4 YNR057C MVD1 YNR043W NA 248 log2(mtr1 0.Ts/WT.T s) adj.p.val Spindle pole body (SPB) component, required for the insertion of the duplication plaque into the nuclear membrane during SPB duplication; essential for bipolar spindle formation; component of the Mps2p‐Bbp1p complex 0.6249 0.0003 YLR165C Pseudouridine synthase, catalyzes only the formation of pseudouridine (Psi)‐2819 in mitochondrial 21S rRNA; not essential for viability 0.6229 0.0006 TOS6 YNL300W Glycosylphosphatidylinositol‐dependent cell wall protein, expression is periodic and decreases in respone to ergosterol perturbation or upon entry into stationary phase; depletion increases resistance to lactic acid 0.6224 0.0003 ITR1 YDR497C Myo‐inositol transporter with strong similarity to the minor myo‐ inositol transporter Itr2p, member of the sugar transporter superfamily; expression is repressed by inositol and choline via Opi1p and derepressed via Ino2p and Ino4p 0.6222 0.0002 PAN5 YHR063C 0.6221 0.0008 SPP2 YOR148C 2‐dehydropantoate 2‐reductase, part of the pantothenic acid pathway, structurally homologous to E. coli panE Essential protein that promotes the first step of splicing and is required for the final stages of spliceosome maturation; interacts with Prp2p, which may release Spp2p from the spliceosome following the first cleavage reaction 0.6206 0.0008 DAL3 YIR032C Ureidoglycolate hydrolase, converts ureidoglycolate to glyoxylate and urea in the third step of allantoin degradation; expression sensitive to nitrogen catabolite repression 0.6199 0.0009 ERG25 YGR060W C‐4 methyl sterol oxidase, catalyzes the first of three steps required to remove two C‐4 methyl groups from an intermediate in ergosterol biosynthesis; mutants accumulate the sterol intermediate 4,4‐dimethylzymosterol 0.6199 0.0007 RDS3 YPR094W Component of the SF3b subcomplex of the U2 snRNP, zinc cluster protein involved in pre‐mRNA splicing and cycloheximide resistance 0.6197 0.0010 MLC2 YPR188C Regulatory light chain for the type II myosin, Myo1p; binds to an IQ motif of Myo1p, localization to the bud neck depends on Myo1p; involved in the disassembly of the Myo1p ring 0.6186 0.0005 CCS1 YMR038C Copper chaperone for superoxide dismutase Sod1p, involved in oxidative stress protection; Met‐X‐Cys‐X2‐Cys motif within the N‐ terminal portion is involved in insertion of copper into Sod1p under conditions of copper deprivation 0.6173 0.0003 PEX22 YAL055W Putative peroxisomal membrane protein required for import of peroxisomal proteins, functionally complements a Pichia pastoris pex22 mutation 0.6138 0.0009 NA YGL041C‐B 0.6133 0.0009 NA YAL044W‐A 0.6123 0.0003 CCT6 YDR188W Putative protein of unknown function; identified by fungal homology and RT‐PCR Putative protein of unknown function; similar to S. pombe uvi31 which is a putative DNA repair protein Subunit of the cytosolic chaperonin Cct ring complex, related to Tcp1p, essential protein that is required for the assembly of actin and tubulins in vivo; contains an ATP‐binding motif 0.6099 0.0002 MDJ1 YFL016C Co‐chaperone that stimulates the ATPase activity of the HSP70 protein Ssc1p; involved in protein folding/refolding in the mitochodrial matrix; required for proteolysis of misfolded proteins; member of the HSP40 (DnaJ) family of chaperones 0.6067 0.0003 Symbols ORF Description NBP1 YLR457C PUS5 249 log2(mtr1 0.Ts/WT.T s) adj.p.val Protein that interacts physically and genetically with Tap42p, which regulates protein phosphatase 2A; component of the TOR (target of rapamycin) signaling pathway 0.6055 0.0004 YOR281C Essential protein that interacts with the CCT (chaperonin containing TCP‐1) complex to stimulate actin folding; has similarity to phosducins; null mutant lethality is complemented by mouse phosducin‐like protein MgcPhLP 0.6055 0.0005 RGT2 YDL138W Plasma membrane high glucose sensor that regulates glucose transport; contains 12 predicted transmembrane segments and a long C‐terminal tail required for induction of hexose transporters; highly similar to Snf3p 0.6055 0.0004 ATP5 YDR298C Subunit 5 of the stator stalk of mitochondrial F1F0 ATP synthase, which is an evolutionarily conserved enzyme complex required for ATP synthesis; homologous to bovine subunit OSCP (oligomycin sensitivity‐conferring protein); phosphorylated 0.6031 0.0007 NUP53 YMR153W Subunit of the nuclear pore complex (NPC), interacts with karyopherin Kap121p or with Nup170p via overlapping regions of Nup53p, involved in activation of the spindle checkpoint mediated by the Mad1p‐Mad2p complex 0.6016 0.0010 REG1 YDR028C Regulatory subunit of type 1 protein phosphatase Glc7p, involved in negative regulation of glucose‐repressible genes 0.6004 0.0007 RSM10 YDR041W Mitochondrial ribosomal protein of the small subunit, has similarity to E. coli S10 ribosomal protein; essential for viability, unlike most other mitoribosomal proteins 0.6004 0.0006 Symbols ORF Description TIP41 YPR040W PLP2 250 APPENDIX K: Summary tables of functional catalogue analysis for affected transcripts in msn5Δ cells in fed or amino acid starvation conditions This section includes four summary results of functional catalogue (FunCat) analysis of transcriptionally differentially expressed genes (cutoff conditions: log2 FC FC 0.6 or log2 0.6 and adjusted p-value < 0.05) in msn5Δ cells in fed or amino acid starvation conditions. The FunCat analysis was executed in the website: http://mips.helmholtzmuenchen.de/proj/funcatDB/ (Ruepp et al., 2004) Table K. 1. FunCat analysis of down-regulated transcripts in msn5Δ cells in fed condition. FUNCTIONAL CATALOGUE phospholipid metabolism mitotic cell cycle RNA synthesis mRNA synthesis transcriptional control transcription activation transcription repression REGULATION OF METABOLISM AND PROTEIN FUNCTION regulation of protein activity enzymatic activity regulation / enzyme regulator CELLULAR COMMUNICATION/SIGNAL TRANSDUCTION MECHANISM G‐protein mediated signal transduction small GTPase mediated signal transduction 251 numbers of gene matches 4 7 20 20 20 3 3 10 9 8 9 5 4 p-value 0.04032 0.037809 0.016061 0.005821 0.000976 0.045394 0.015685 0.022164 0.040119 0.021278 0.034185 0.009891 0.024253 continued Table. K. 1. continued FUNCTIONAL CATALOGUE transmembrane signal transduction non‐enzymatic receptor mediated signalling G‐protein coupled receptor signalling pathway catalase reaction INTERACTION WITH THE ENVIRONMENT cellular sensing and response to external stimulus chemoperception and response pheromone response, mating‐type determination, sex‐specific proteins CELL FATE cell growth / morphogenesis growth regulators / regulation of cell size DEVELOPMENT (Systemic) fungal/microorganismic development mating (fertilization) cytoskeleton/structural proteins centrosome CELL TYPE DIFFERENTIATION fungal/microorganismic cell type differentiation fungal and other eukaryotic cell type differentiation budding, cell polarity and filament formation hyphae formation UNCLASSIFIED PROTEINS numbers of gene matches 5 2 2 1 25 22 20 0.001254 0.007125 0.007125 0.037812 1.39E‐06 1.27E‐08 1.86E‐08 16 11 11 3 11 11 11 9 1 19 19 19 14 1 18 4.63E‐07 0.014673 0.005503 0.001315 5.48E‐08 5.48E‐08 5.48E‐08 0.050802 0.037812 0.000824 0.000824 0.000824 0.002384 0.037812 0.982084 p-value Table K. 2. Up-regulated transcripts in msn5Δ cells in fed condition. FUNCTIONAL CATALOGUE METABOLISM assimilation of ammonia, metabolism of the glutamate group metabolism of proline metabolism of arginine degradation of arginine metabolism of urea cycle, creatine and polyamines metabolism of urea (urea cycle) nitrogen, sulfur and selenium metabolism nitrogen metabolism catabolism of nitrogenous compounds urea catabolism (not urea cycle) phosphate metabolism C‐compound and carbohydrate metabolism 252 numbers of gene matches 64 4 2 3 3 4 3 9 3 3 1 20 34 p-value 0.000215 0.043414 0.015316 0.020528 2.25E‐05 0.00067 0.000422 0.001626 0.000723 0.000723 0.028385 0.014327 1.33E‐06 continued. Table K. 2. continued. FUNCTIONAL CATALOGUE sugar, glucoside, polyol and carboxylate metabolism sugar, glucoside, polyol and carboxylate anabolism sugar, glucoside, polyol and carboxylate catabolism glycogen metabolism glycogen anabolism regulation of C‐compound and carbohydrate metabolism ENERGY glycolysis and gluconeogenesis regulation of glycolysis and gluconeogenesis electron transport and membrane‐associated energy conservation respiration aerobic respiration metabolism of energy reserves (e.g. glycogen, trehalose) lysosomal and vacuolar protein degradation vacuolar protein degradation biotin binding transported compounds (substrates) anion transport phosphate transport C‐compound and carbohydrate transport 20.01.03.01 sugar transport 20.01.07 amino acid/amino acid derivatives transport 20.01.15 electron transport 20.03.02 carrier (electrochemical potential‐driven transport) 20.03.02.02 symporter 20.03.02.02.01 proton driven symporter 20.03.02.02.02 sodium driven symporter 20.03.02.03 antiporter 32.01.01 oxidative stress response 32.07.07 oxygen and radical detoxification 32.07.07.05 peroxidase reaction 34.01.03.03 homeostasis of phosphate 42.25 vacuole or lysosome 99 UNCLASSIFIED PROTEINS 253 numbers of gene matches 9 5 8 2 2 8 31 6 3 8 11 6 10 3 2 1 28 3 3 10 5 4 8 6 3 2 1 3 5 3 2 2 5 55 p-value 0.000438 0.00283 0.00168 0.007572 0.007572 0.027488 2.74E‐08 0.006218 0.015587 0.000292 0.001774 0.021576 3.02E‐06 0.036207 0.037236 0.028385 0.003816 0.036207 0.003135 9.11E‐06 0.001188 0.037865 0.002307 6.71E‐06 8.81E‐05 0.002359 0.028385 0.006496 0.019263 0.036207 0.015316 0.025294 0.007713 0.003908 Table K. 3. FunCat analysis of down-regulated transcripts in msn5Δ cells in amino acid starvation condition. FUNCTIONAL CATALOGUE 10.03 cell cycle 10.03.01 mitotic cell cycle and cell cycle control 10.03.02 meiosis 10.03.02.01 meiosis I 10.03.05 cell cycle dependent cytoskeleton reorganization 10.03.05.01 spindle pole body/centrosome and microtubule cycle 18 REGULATION OF METABOLISM AND PROTEIN FUNCTION 18.02 regulation of protein activity 18.02.01 enzymatic activity regulation / enzyme regulator 18.02.05 regulator of G‐protein signalling 20.01.17 nucleotide/nucleoside/nucleobase transport 20.01.23 allantoin and allantoate transport 20.01.27 drug/toxin transport 20.03 transport facilities 20.03.25 ABC transporters 30 CELLULAR COMMUNICATION/SIGNAL TRANSDUCTION MECHANISM 30.01.05 enzyme mediated signal transduction 30.01.05.05 G‐protein mediated signal transduction 30.01.05.05.01 small GTPase mediated signal transduction 30.05 transmembrane signal transduction 30.05.02 non‐enzymatic receptor mediated signalling 30.05.02.24 G‐protein coupled receptor signalling pathway 34 INTERACTION WITH THE ENVIRONMENT 34.11 cellular sensing and response to external stimulus 34.11.03 chemoperception and response 34.11.03.07 pheromone response, mating‐type determination, sex‐specific proteins 41 DEVELOPMENT (Systemic) 41.01 fungal/microorganismic development 41.01.01 mating (fertilization) 42.04.05 microtubule cytoskeleton 43 CELL TYPE DIFFERENTIATION 43.01 fungal/microorganismic cell type differentiation 43.01.03 fungal and other eukaryotic cell type differentiation 43.01.03.05 budding, cell polarity and filament formation 43.01.03.09 development of asco‐ basidio‐ or zygospore 99 UNCLASSIFIED PROTEINS 254 numbers of gene matches 43 28 13 3 6 6 19 16 13 3 4 3 6 17 7 19 11 7 6 10 3 3 40 35 33 0.001358 0.018237 0.016221 0.014924 0.015233 0.007273 0.008736 0.042612 0.036475 0.022331 0.008366 0.00341 0.005018 0.003107 0.000111 0.003804 0.022151 0.023786 0.032409 7.40E‐06 0.002199 0.002199 5.59E‐06 4.22E‐09 7.20E‐10 30 18 18 18 7 43 43 43 26 17 50 1.30E‐10 1.97E‐10 1.97E‐10 1.97E‐10 0.002998 1.64E‐07 1.64E‐07 1.64E‐07 0.00052 0.00053 0.902999 p-value Table K. 4. FunCat analysis of up-regulated transcripts in msn5Δ cells in amino acid starvation condition. FUNCTIONAL CATALOGUE degradation of arginine degradation of serine nitrogen metabolism catabolism of nitrogenous compounds ENERGY electron transport and membrane‐associated energy conservation respiration aerobic respiration protein folding and stabilization enzyme inhibitor protease inhibitor anion transport phosphate transport C‐compound and carbohydrate transport electron transport carrier (electrochemical potential‐driven transport) symporter CELL RESCUE, DEFENSE AND VIRULENCE stress response oxidative stress response oxygen and radical detoxification peroxidase reaction homeostasis of anions homeostasis of phosphate vacuole or lysosome UNCLASSIFIED PROTEINS 255 numbers of gene matches 2 1 2 2 23 6 12 6 8 5 2 3 3 5 6 4 2 25 23 8 4 2 3 3 7 61 p-value 0.002838 0.031158 0.018292 0.018292 0.000962 0.011323 0.001161 0.032216 0.008134 0.004229 0.009077 0.04575 0.004084 0.045681 0.044096 0.001983 0.00556 0.036729 0.012182 0.000266 0.007976 0.018292 0.010274 0.002177 0.000368 0.001865 APPENDIX L: Down-regulated genes in msn5Δ cells in fed condition This section includes the list of transcriptionally differentially expressed genes (cutoff conditions: log2 FC 0.6 and adjusted p-value < 0.05) in msn5Δ cells in fed condition. Down-regulated genes in msn5Δ cells in fed condition log2(msn5 .Tf/WT.Tf) adj.p. val Karyopherin involved in nuclear import and export of proteins, including import of replication protein A and export of Swi6p, Far1p, and Pho4p; required for re‐export of mature tRNAs after their retrograde import from the cytoplasm ‐7.1037 0.0000 YML058W‐A Protein involved in the Mec1p‐mediated checkpoint pathway that responds to DNA damage or replication arrest, transcription is induced by DNA damage ‐2.4244 0.0013 PRM1 YNL279W Pheromone‐regulated multispanning membrane protein involved in membrane fusion during mating; predicted to have 5 transmembrane segments and a coiled coil domain; localizes to the shmoo tip; regulated by Ste12p ‐2.2513 0.0000 FIG1 YBR040W Integral membrane protein required for efficient mating; may participate in or regulate the low affinity Ca2+ influx system, which affects intracellular signaling and cell‐cell fusion during mating ‐2.2108 0.0085 DAN1 YJR150C Cell wall mannoprotein with similarity to Tir1p, Tir2p, Tir3p, and Tir4p; expressed under anaerobic conditions, completely repressed during aerobic growth ‐2.1252 0.0041 PRM6 YML047C Pheromone‐regulated protein, predicted to have 2 transmembrane segments; regulated by Ste12p during mating ‐2.0349 0.0150 FUS1 YCL027W Membrane protein localized to the shmoo tip, required for cell fusion; expression regulated by mating pheromone; proposed to coordinate signaling, fusion, and polarization events required for fusion; potential Cdc28p substrate ‐1.7866 0.0014 WSC4 YHL028W ER membrane protein involved in the translocation of soluble secretory proteins and insertion of membrane proteins into the ER membrane; may also have a role in the stress response but has only partial functional overlap with WSC1‐3 ‐1.6054 0.0162 Symbols ORF Description MSN5 YDR335W HUG1 256 log2(msn5 .Tf/WT.Tf) adj.p. val Nonfunctional protein with homology to IMP dehydrogenase; probable pseudogene, located close to the telomere; is not expressed at detectable levels; YAR073W and YAR075W comprise a continuous reading frame in some strains of S. cerevisiae ‐1.5849 0.0096 YDL227C Site‐specific endonuclease required for gene conversion at the MAT locus (homothallic switching) through the generation of a ds DNA break; expression restricted to mother cells in late G1 as controlled by Swi4p‐Swi6p, Swi5p and Ash1p ‐1.5549 0.0032 AGA1 YNR044W ‐1.5162 0.0013 MF(ALPHA )2 YGL089C Anchorage subunit of a‐agglutinin of a‐cells, highly O‐glycosylated protein with N‐terminal secretion signal and C‐terminal signal for addition of GPI anchor to cell wall, linked to adhesion subunit Aga2p via two disulfide bonds Mating pheromone alpha‐factor, made by alpha cells; interacts with mating type a cells to induce cell cycle arrest and other responses leading to mating; also encoded by MF(ALPHA)1, which is more highly expressed than MF(ALPHA)2 ‐1.4937 0.0045 NA TIR1 YAR066W YER011W Putative GPI protein Cell wall mannoprotein of the Srp1p/Tip1p family of serine‐alanine‐ rich proteins; expression is downregulated at acidic pH and induced by cold shock and anaerobiosis; abundance is increased in cells cultured without shaking ‐1.4805 ‐1.4492 0.0017 0.0015 NA YIL082W‐A Retrotransposon TYA Gag and TYB Pol genes; transcribed/translated as one unit; polyprotein is processed to make a nucleocapsid‐like protein (Gag), reverse transcriptase (RT), protease (PR), and integrase (IN); similar to retroviral genes ‐1.3888 0.0005 NA YDR222W ‐1.3702 0.0276 PLB2 YMR006C Protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm in a punctate pattern Phospholipase B (lysophospholipase) involved in phospholipid metabolism; displays transacylase activity in vitro; overproduction confers resistance to lysophosphatidylcholine ‐1.3209 0.0026 NA YLR042C ‐1.3205 0.0015 STR3 YGL184C ‐1.3066 0.0341 SRO77 YBL106C ‐1.2967 0.0219 PPM2 YOL141W AdoMet‐dependent tRNA methyltransferase also involved in methoxycarbonylation; required for the synthesis of wybutosine (yW), a modified guanosine found at the 3'‐position adjacent to the anticodon of phe‐tRNA; similarity to Ppm1p ‐1.2951 0.0029 PRM2 YIL037C Pheromone‐regulated protein, predicted to have 4 transmembrane segments and a coiled coil domain; regulated by Ste12p; required for efficient nuclear fusion ‐1.2774 0.0231 SST2 YLR452C GTPase‐activating protein for Gpa1p, regulates desensitization to alpha factor pheromone; also required to prevent receptor‐ independent signaling of the mating pathway; member of the RGS (regulator of G‐protein signaling) family ‐1.1922 0.0015 SFG1 YOR315W Nuclear protein, putative transcription factor required for growth of superficial pseudohyphae (which do not invade the agar substrate) but not for invasive pseudohyphal growth; may act together with Phd1p; potential Cdc28p substrate ‐1.1781 0.0159 Symbols ORF Description IMD1 YAR073W HO Protein of unknown function; localizes to the cytoplasm; YLL042C is not an essential gene Peroxisomal cystathionine beta‐lyase, converts cystathionine into homocysteine; may be redox regulated by Gto1p Protein with roles in exocytosis and cation homeostasis; functions in docking and fusion of post‐Golgi vesicles with plasma membrane; homolog of Sro7p and Drosophila lethal giant larvae tumor suppressor; interacts with SNARE protein Sec9p 257 log2(msn5 .Tf/WT.Tf) adj.p. val Inosine monophosphate dehydrogenase, catalyzes the rate‐limiting step in GTP biosynthesis, expression is induced by mycophenolic acid resulting in resistance to the drug, expression is repressed by nutrient limitatio ‐1.1711 0.0031 YJR004C Alpha‐agglutinin of alpha‐cells, binds to Aga1p during agglutination, N‐ terminal half is homologous to the immunoglobulin superfamily and contains binding site for a‐agglutinin, C‐terminal half is highly glycosylated and contains GPI anchor ‐1.1666 0.0009 CRR1 YLR213C Putative glycoside hydrolase of the spore wall envelope; required for normal spore wall assembly, possibly for cross‐linking between the glucan and chitosan layers; expressed during sporulation ‐1.1315 0.0171 RIM9 YMR063W Protein of unknown function, involved in the proteolytic activation of Rim101p in response to alkaline pH; has similarity to A. nidulans PalI; putative membrane protein ‐1.1261 0.0253 NA YCR101C ‐1.1227 0.0171 PCL1 YNL289W Putative protein of unknown function; localizes to the membrane fraction; YCR101C is not an essential gene Cyclin, interacts with cyclin‐dependent kinase Pho85p; member of the Pcl1,2‐like subfamily, involved in the regulation of polarized growth and morphogenesis and progression through the cell cycle; localizes to sites of polarized cell growth ‐1.1030 0.0109 NA YHL018W Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to mitochondria and is induced in response to the DNA‐damaging agent MMS ‐1.0830 0.0191 SVS1 YPL163C ‐1.0814 0.0113 SPS1 YDR523C Cell wall and vacuolar protein, required for wild‐type resistance to vanadate Putative protein serine/threonine kinase expressed at the end of meiosis and localized to the prospore membrane, required for correct localization of enzymes involved in spore wall synthesis ‐1.0764 0.0392 CLN1 YMR199W G1 cyclin involved in regulation of the cell cycle; activates Cdc28p kinase to promote the G1 to S phase transition; late G1 specific expression depends on transcription factor complexes, MBF (Swi6p‐ Mbp1p) and SBF (Swi6p‐Swi4p) ‐1.0744 0.0017 CLB2 YPR119W B‐type cyclin involved in cell cycle progression; activates Cdc28p to promote the transition from G2 to M phase; accumulates during G2 and M, then targeted via a destruction box motif for ubiquitin‐ mediated degradation by the proteasome ‐1.0713 0.0109 RPI1 YIL119C Putative transcriptional regulator; overexpression suppresses the heat shock sensitivity of wild‐type RAS2 overexpression and also suppresses the cell lysis defect of an mpk1 mutation ‐1.0693 0.0489 SPC24 YMR117C Component of the evolutionarily conserved kinetochore‐associated Ndc80 complex (Ndc80p‐Nuf2p‐Spc24p‐Spc25p); involved in chromosome segregation, spindle checkpoint activity and kinetochore clustering ‐1.0451 0.0139 YHP1 YDR451C One of two homeobox transcriptional repressors (see also Yox1p), that bind to Mcm1p and to early cell cycle box (ECB) elements of cell cycle regulated genes, thereby restricting ECB‐mediated transcription to the M/G1 interval ‐1.0427 0.0077 FAR1 YJL157C Cyclin‐dependent kinase inhibitor that mediates cell cycle arrest in response to pheromone; also forms a complex with Cdc24p, Ste4p, and Ste18p that may specify the direction of polarized growth during mating; potential Cdc28p substrate ‐1.0374 0.0013 Symbols ORF Description IMD2 YHR216W SAG1 258 log2(msn5 .Tf/WT.Tf) adj.p. val Protein of unknown function; green fluorescent protein (GFP)‐ fusion protein localizes to the mother side of the bud neck and the vacuole; YOL007C is not an essential gene ‐1.0200 0.0026 YOR237W Protein implicated in the regulation of ergosterol biosynthesis; one of a seven member gene family with a common essential function and non‐essential unique functions; similar to human oxysterol binding protein (OSBP) ‐1.0063 0.0054 GPI19 YDR437W Subunit of GPI‐GlcNAc transferase involved in synthesis of N‐ acetylglucosaminyl phosphatidylinositol (GlcNAc‐PI), which is the first intermediate in glycosylphosphatidylinositol (GPI) anchor synthesis, shares similarity with mammalian PIG‐P ‐0.9955 0.0034 PDR15 YDR406W Plasma membrane ATP binding cassette (ABC) transporter, multidrug transporter and general stress response factor implicated in cellular detoxification; regulated by Pdr1p, Pdr3p and Pdr8p; promoter contains a PDR responsive element ‐0.9730 0.0026 NRM1 YNR009W Transcriptional co‐repressor of MBF (MCB binding factor)‐regulated gene expression; Nrm1p associates stably with promoters via MBF to repress transcription upon exit from G1 phase ‐0.9644 0.0032 RTC4 YNL254C Protein of unknown function; null mutation suppresses cdc13‐1 temperature sensitivity; (GFP)‐fusion protein localizes to both the cytoplasm and the nucleus ‐0.9440 0.0029 STE3 YKL178C Receptor for a factor pheromone, couples to MAP kinase cascade to mediate pheromone response; transcribed in alpha cells and required for mating by alpha cells, ligand bound receptors endocytosed and recycled to the plasma membrane; GPCR ‐0.9394 0.0013 AQY1 YPR192W Spore‐specific water channel that mediates the transport of water across cell membranes, developmentally controlled; may play a role in spore maturation, probably by allowing water outflow, may be involved in freeze tolerance ‐0.9382 0.0116 NA YGR035C ‐0.9278 0.0130 FRE8 YLR047C Putative protein of unknown function, potential Cdc28p substrate; transcription is activated by paralogous transcription factors Yrm1p and Yrr1p along with genes involved in multidrug resistance Protein with sequence similarity to iron/copper reductases, involved in iron homeostasis; deletion mutant has iron deficiency/accumulation growth defects; expression increased in the absence of copper‐ responsive transcription factor Mac1p ‐0.9271 0.0091 NA YGR109W‐B Retrotransposon TYA Gag and TYB Pol genes; transcribed/translated as one unit; polyprotein is processed to make a nucleocapsid‐like protein (Gag), reverse transcriptase (RT), protease (PR), and integrase (IN); similar to retroviral genes ‐0.9258 0.0034 PET122 YER153C Mitochondrial translational activator specific for the COX3 mRNA, acts together with Pet54p and Pet494p; located in the mitochondrial inner membrane ‐0.9248 0.0159 NA YAR068W ‐0.9239 0.0221 LRG1 YDL240W ‐0.9226 0.0023 FMP45 YDL222C Fungal‐specific protein of unknown function; induced in respiratory‐ deficient cells Putative GTPase‐activating protein (GAP) involved in the Pkc1p‐ mediated signaling pathway that controls cell wall integrity; appears to specifically regulate 1,3‐beta‐glucan synthesis Integral membrane protein localized to mitochondria (untagged protein); required for sporulation and maintaining sphingolipid content; has sequence similarity to SUR7 and YNL194C ‐0.9193 0.0109 Symbols ORF Description CSI2 YOL007C HES1 259 log2(msn5 .Tf/WT.Tf) adj.p. val v‐SNARE binding protein that facilitates specific protein retrieval from a late endosome to the Golgi; modulates arginine uptake, possible role in mediating pH homeostasis between the vacuole and plasma membrane H(+)‐ATPase ‐0.9012 0.0050 YGL175C Endonuclease that processes hairpin DNA structures with the MRX complex; involved in meiotic and mitotic double‐strand break repair; phosphorylated in response to DNA damage and required for normal resistance to DNA‐damaging agents ‐0.8847 0.0096 YOX1 YML027W Homeodomain‐containing transcriptional repressor, binds to Mcm1p and to early cell cycle boxes (ECBs) in the promoters of cell cycle‐ regulated genes expressed in M/G1 phase; expression is cell cycle‐ regulated; potential Cdc28p substrate ‐0.8839 0.0355 CLN2 YPL256C G1 cyclin involved in regulation of the cell cycle; activates Cdc28p kinase to promote the G1 to S phase transition; late G1 specific expression depends on transcription factor complexes, MBF (Swi6p‐ Mbp1p) and SBF (Swi6p‐Swi4p) ‐0.8757 0.0068 NA NA PTH1 NA NA YHR189W NA NA One of two (see also PTH2) mitochondrially‐localized peptidyl‐tRNA hydrolases; dispensable for respiratory growth on rich medium, but required for respiratory growth on minimal medium ‐0.8703 ‐0.8682 ‐0.8578 0.0148 0.0170 0.0495 CLB6 YGR109C B‐type cyclin involved in DNA replication during S phase; activates Cdc28p to promote initiation of DNA synthesis; functions in formation of mitotic spindles along with Clb3p and Clb4p; most abundant during late G1 ‐0.8495 0.0144 NA YDR124W ‐0.8480 0.0187 GIC1 YHR061C Putative protein of unknown function; non‐essential gene; expression is strongly induced by alpha factor Protein of unknown function involved in initiation of budding and cellular polarization, interacts with Cdc42p via the Cdc42/Rac‐ interactive binding (CRIB) domain ‐0.8437 0.0183 OPT1 YJL212C Proton‐coupled oligopeptide transporter of the plasma membrane; also transports glutathione and phytochelatin; member of the OPT family ‐0.8396 0.0456 SFK1 YKL051W Plasma membrane protein that may act together with or upstream of Stt4p to generate normal levels of the essential phospholipid PI4P, at least partially mediates proper localization of Stt4p to the plasma membrane ‐0.8301 0.0308 PLM2 YDR501W Forkhead Associated domain containing protein and putative transcription factor found associated with chromatin; target of SBF transcription factor; induced in response to DNA damaging agents and deletion of telomerase; similar to TOS4 ‐0.8251 0.0057 DCI1 YOR180C ‐0.8201 0.0242 CIN8 YEL061C ‐0.8094 0.0143 YJU2 YKL095W ‐0.7977 0.0162 PMA2 YPL036W Peroxisomal protein; identification as a delta(3,5)‐delta(2,4)‐dienoyl‐ CoA isomerase involved in fatty acid metabolism is disputed Kinesin motor protein involved in mitotic spindle assembly and chromosome segregation Essential protein required for pre‐mRNA splicing; associates transiently with the spliceosomal NTC ("nineteen complex") and acts after Prp2p to promote the first catalytic reaction of splicing Plasma membrane H+‐ATPase, isoform of Pma1p, involved in pumping protons out of the cell; regulator of cytoplasmic pH and plasma membrane potential ‐0.7944 0.0221 DED1 YOR204W ‐0.7753 0.0126 Symbols ORF Description BTN2 YGR142W SAE2 ATP‐dependent DEAD (Asp‐Glu‐Ala‐Asp)‐box RNA helicase, required for translation initiation of all yeast mRNAs; mutations in human DEAD‐box DBY are a frequent cause of male infertility 260 log2(msn5 .Tf/WT.Tf) adj.p. val Putative lipase; involved in lipid metabolism; YDL109C is not an essential gene Putative protein of unknown function Subunit of the conserved chromosomal passenger complex (CPC; Ipl1p‐Sli15p‐Bir1p‐Nbl1p), which regulates mitotic chromosome segregation; not required for the kinase activity of the complex; mediates the interaction of Sli15p and Bir1p ‐0.7744 0.0057 ‐0.7739 ‐0.7731 0.0141 0.0206 YHR084W Transcription factor that is activated by a MAP kinase signaling cascade, activates genes involved in mating or pseudohyphal/invasive growth pathways; cooperates with Tec1p transcription factor to regulate genes specific for invasive growth ‐0.7682 0.0054 SFL1 YOR140W Transcriptional repressor and activator; involved in repression of flocculation‐related genes, and activation of stress responsive genes; negatively regulated by cAMP‐dependent protein kinase A subunit Tpk2p ‐0.7648 0.0144 NRT1 YOR071C ‐0.7616 0.0191 RTG1 YOL067C ‐0.7588 0.0122 GEM1 YAL048C High‐affinity nicotinamide riboside transporter; also transports thiamine with low affinity; shares sequence similarity with Thi7p and Thi72p; proposed to be involved in 5‐fluorocytosine sensitivity Transcription factor (bHLH) involved in interorganelle communication between mitochondria, peroxisomes, and nucleus Evolutionarily‐conserved tail‐anchored outer mitochondrial membrane GTPase which regulates mitochondrial morphology; cells lacking Gem1p contain collapsed, globular, or grape‐like mitochondria; not required for pheromone‐induced cell death ‐0.7553 0.0175 CTA1 YDR256C ‐0.7546 0.0186 FHL1 YPR104C Catalase A, breaks down hydrogen peroxide in the peroxisomal matrix formed by acyl‐CoA oxidase (Pox1p) during fatty acid beta‐oxidation Regulator of ribosomal protein transcription; has forkhead associated domain that binds phosphorylated proteins; also has forkhead DNA‐ binding domain but does not bind DNA in vitro; suppresses RNA pol III and splicing factor prp4 mutants ‐0.7505 0.0054 WHI5 YOR083W Repressor of G1 transcription that binds to SCB binding factor (SBF) at SCB target promoters in early G1; phosphorylation of Whi5p by the CDK, Cln3p/Cdc28p relieves repression and promoter binding by Whi5; periodically expressed in G1 ‐0.7453 0.0276 NA TOS2 YCR100C YGR221C Putative protein of unknown function Protein involved in localization of Cdc24p to the site of bud growth; may act as a membrane anchor; localizes to the bud neck and bud tip; potentially phosphorylated by Cdc28p ‐0.7411 ‐0.7389 0.0269 0.0033 PRM3 YPL192C ‐0.7386 0.0116 YHK8 YHR048W ‐0.7354 0.0168 DFG10 NA INO2 YIL049W YNR063W YDR123C Pheromone‐regulated protein required for nuclear envelope fusion during karyogamy; localizes to the outer face of the nuclear membrane; interacts with Kar5p at the spindle pole body Presumed antiporter of the DHA1 family of multidrug resistance transporters; contains 12 predicted transmembrane spans; expression of gene is up‐regulated in cells exhibiting reduced susceptibility to azoles Protein of unknown function, involved in filamentous growth Putative zinc‐cluster protein of unknown function Component of the heteromeric Ino2p/Ino4p basic helix‐loop‐helix transcription activator that binds inositol/choline‐responsive elements (ICREs), required for derepression of phospholipid biosynthetic genes in response to inositol depletion ‐0.7329 ‐0.7263 ‐0.7210 0.0029 0.0316 0.0308 NA YOL014W ‐0.7177 0.0328 Symbols ORF Description NA YDL109C NA NBL1 YCL021W‐A YHR199C‐A STE12 Putative protein of unknown function 261 log2(msn5 .Tf/WT.Tf) adj.p. val ATP‐binding cassette (ABC) transporter, multidrug transporter involved in multiple drug resistance; mediates sterol uptake when sterol biosynthesis is compromisedregulated by Pdr1p; required for anaerobic growth ‐0.7172 0.0105 YOL028C YGR055W Putative basic leucine zipper (bZIP) transcription factor High affinity methionine permease, integral membrane protein with 13 putative membrane‐spanning regions; also involved in cysteine uptake ‐0.7075 ‐0.7045 0.0483 0.0339 FIG2 YCR089W ‐0.6983 0.0041 PHO4 YFR034C Cell wall adhesin, expressed specifically during mating; may be involved in maintenance of cell wall integrity during mating Basic helix‐loop‐helix (bHLH) transcription factor of the myc‐family; binds cooperatively with Pho2p to the PHO5 promoter; function is regulated by phosphorylation at multiple sites and by phosphate availability ‐0.6959 0.0171 RIM4 YHL024W ‐0.6924 0.0452 TKL2 YBR117C Putative RNA‐binding protein required for the expression of early and middle sporulation genes Transketolase, similar to Tkl1p; catalyzes conversion of xylulose‐5‐ phosphate and ribose‐5‐phosphate to sedoheptulose‐7‐phosphate and glyceraldehyde‐3‐phosphate in the pentose phosphate pathway; needed for synthesis of aromatic amino acids ‐0.6871 0.0044 SRB8 YCR081W Subunit of the RNA polymerase II mediator complex; associates with core polymerase subunits to form the RNA polymerase II holoenzyme; essential for transcriptional regulation; involved in glucose repression ‐0.6858 0.0072 IZH1 YDR492W Membrane protein involved in zinc ion homeostasis, member of the four‐protein IZH family; transcription is regulated directly by Zap1p, expression induced by zinc deficiency and fatty acids; deletion increases sensitivity to elevated zinc ‐0.6721 0.0241 SCW11 YGL028C ‐0.6677 0.0241 AIM20 YIL158W Cell wall protein with similarity to glucanases; may play a role in conjugation during mating based on its regulation by Ste12p Putative protein of unknown function; overexpression causes a cell cycle delay or arrest; green fluorescent protein (GFP)‐fusion protein localizes to the vacuole; null mutant displays elevated frequency of mitochondrial genome loss ‐0.6643 0.0066 WHI3 YNL197C RNA binding protein that sequesters CLN3 mRNA in cytoplasmic foci; cytoplasmic retention factor for Cdc28p and associated cyclins; regulates cell fate and dose‐dependently regulates the critical cell size required for passage through Start ‐0.6550 0.0356 TOS6 YNL300W ‐0.6550 0.0223 TEC1 YBR083W Glycosylphosphatidylinositol‐dependent cell wall protein, expression is periodic and decreases in respone to ergosterol perturbation or upon entry into stationary phase; depletion increases resistance to lactic acid Transcription factor required for full Ty1 expression, Ty1‐mediated gene activation, and haploid invasive and diploid pseudohyphal growth; TEA/ATTS DNA‐binding domain family member ‐0.6480 0.0051 JJJ2 YJL162C ‐0.6452 0.0200 MSB2 YGR014W ‐0.6385 0.0123 ATF2 YGR177C ‐0.6337 0.0266 Symbols ORF Description PDR11 YIL013C YAP7 MUP1 Protein of unknown function, contains a J‐domain, which is a region with homology to the E. coli DnaJ protein Mucin family member involved in the Cdc42p‐ and MAP kinase‐ dependent filamentous growth signaling pathway; also functions as an osmosensor in parallel to the Sho1p‐mediated pathway; potential Cdc28p substrate Alcohol acetyltransferase, may play a role in steroid detoxification; forms volatile esters during fermentation, which is important for brewing and winemaking 262 log2(msn5 .Tf/WT.Tf) adj.p. val Transcriptional activator essential for nuclear division; localized to the nucleus; essential component of the mechanism that activates the expression of a set of late‐S‐phase‐specific genes ‐0.6298 0.0242 YDR138W Subunit of THO/TREX complexes that couple transcription elongation with mitotic recombination and with mRNA metabolism and export, subunit of an RNA Pol II complex; regulates lifespan; involved in telomere maintenance; similar to Top1p ‐0.6239 0.0036 FRK1 YPL141C Putative protein kinase; similar to Kin4p; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm; interacts with rRNA transcription and ribosome biogenesis factors and the long chain fatty acyl‐CoA synthetase Faa3 ‐0.6223 0.0209 NA YLL066W‐B ‐0.6210 0.0065 SFP1 YLR403W Putative protein of unknown function; overexpression causes a cell cycle delay or arrest Transcription factor that controls expression of ribosome biogenesis genes in response to nutrients and stress, regulates G2/M transitions during mitotic cell cycle and DNA‐damage response, modulates cell size; regulated by TORC1 and Mrs6p ‐0.6205 0.0483 ADH4 YGL256W Alcohol dehydrogenase isoenzyme type IV, dimeric enzyme demonstrated to be zinc‐dependent despite sequence similarity to iron‐activated alcohol dehydrogenases; transcription is induced in response to zinc deficiency ‐0.6147 0.0108 RAX2 YLR084C N‐glycosylated protein involved in the maintenance of bud site selection during bipolar budding; localization requires Rax1p; RAX2 mRNA stability is regulated by Mpt5p ‐0.6133 0.0188 STB5 YHR178W Transcription factor, involved in regulating multidrug resistance and oxidative stress response; forms a heterodimer with Pdr1p; contains a Zn(II)2Cys6 zinc finger domain that interacts with a pleiotropic drug resistance element in vitro ‐0.6112 0.0089 KAR5 YMR065W Protein required for nuclear membrane fusion during karyogamy, localizes to the membrane with a soluble portion in the endoplasmic reticulum lumen, may form a complex with Jem1p and Kar2p; expression of the gene is regulated by pheromone ‐0.6068 0.0060 VBA2 NA YBR293W YJL107C Permease of basic amino acids in the vacuolar membrane Putative protein of unknown function; expression is induced by activation of the HOG1 mitogen‐activated signaling pathway and this induction is Hog1p/Pbs2p dependent; YJL107C and adjacent ORF, YJL108C are merged in related fungi ‐0.6052 ‐0.6049 0.0231 0.0171 Symbols ORF Description NDD1 YOR372C HPR1 263 APPENDIX M: Up-regulated genes in msn5Δ cells in fed condition This section includes the list of transcriptionally differentially expressed genes (cutoff conditions: log2 FC 0.6 and adjusted p-value < 0.05) in msn5Δ cells in fed condition. Up-regulated genes in msn5Δ cells in fed condition log2(msn5 .Tf/WT.Tf) adj.p.v al High‐affinity inorganic phosphate (Pi) transporter and low‐affinity manganese transporter; regulated by Pho4p and Spt7p; mutation confers resistance to arsenate; exit from the ER during maturation requires Pho86p 6.5678 0.0000 YHR136C Protein with similarity to cyclin‐dependent kinase inhibitors; downregulates low‐affinity phosphate transport during phosphate limitation; overproduction suppresses a plc1 null mutation; GFP‐ fusion protein localizes to the cytoplasm 5.1812 0.0000 PHO89 YBR296C 2.7683 0.0065 PHO11 YAR071W Na+/Pi cotransporter, active in early growth phase; similar to phosphate transporters of Neurospora crassa; transcription regulated by inorganic phosphate concentrations and Pho4p One of three repressible acid phosphatases, a glycoprotein that is transported to the cell surface by the secretory pathway; induced by phosphate starvation and coordinately regulated by PHO4 and PHO2 2.5966 0.0026 PHM6 YDR281C 2.3875 0.0109 PHO5 YBR093C Protein of unknown function, expression is regulated by phosphate levels Repressible acid phosphatase (1 of 3) that also mediates extracellular nucleotide‐derived phosphate hydrolysis; secretory pathway derived cell surface glycoprotein; induced by phosphate starvation and coordinately regulated by PHO4 and PHO2 2.3445 0.0014 VTC3 YPL019C Subunit of the vacuolar transporter chaperone (VTC) complex involved in membrane trafficking, vacuolar polyphosphate accumulation, microautophagy and non‐autophagic vacuolar fusion 2.3172 0.0005 CYC7 YEL039C Cytochrome c isoform 2, expressed under hypoxic conditions; electron carrier of the mitochondrial intermembrane space that transfers electrons from ubiquinone‐cytochrome c oxidoreductase to cytochrome c oxidase during cellular respiration 1.8015 0.0071 NA MTH1 YLR312C YDR277C Putative protein of unknown function Negative regulator of the glucose‐sensing signal transduction pathway, required for repression of transcription by Rgt1p; interacts with Rgt1p and the Snf3p and Rgt2p glucose sensors; phosphorylated by Yck1p, triggering Mth1p degradation 1.7179 1.6946 0.0028 0.0015 Symbols ORF Description PHO84 YML123C SPL2 264 log2(msn5 .Tf/WT.Tf) adj.p.v al Cytochrome b2 (L‐lactate cytochrome‐c oxidoreductase), component of the mitochondrial intermembrane space, required for lactate utilization; expression is repressed by glucose and anaerobic conditions 1.5974 0.0004 YFR017C Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm and is induced in response to the DNA‐damaging agent MMS; YFR017C is not an essential gene 1.5519 0.0005 NA YGL262W Putative protein of unknown function; null mutant displays elevated sensitivity to expression of a mutant huntingtin fragment or of alpha‐ synuclein; YGL262W is not an essential gene 1.5464 0.0219 SOL4 NA YGR248W YHR214C‐E 6‐phosphogluconolactonase with similarity to Sol3p Putative protein of unknown function; identified by gene‐trapping, microarray‐based expression analysis, and genome‐wide homology searching 1.5294 1.4988 0.0170 0.0110 CRC1 YOR100C Mitochondrial inner membrane carnitine transporter, required for carnitine‐dependent transport of acetyl‐CoA from peroxisomes to mitochondria during fatty acid beta‐oxidation 1.4600 0.0016 DCS2 YOR173W Non‐essential, stress induced regulatory protein; modulates m7G‐ oligoribonucleotide metabolism; inhibits Dcs1p; regulated by Msn2p, Msn4p, and the Ras‐cAMP‐cAPK signaling pathway; mutant has increased aneuploidy tolerance 1.4014 0.0026 GPM2 YDL021W Homolog of Gpm1p phosphoglycerate mutase, which converts 3‐ phosphoglycerate to 2‐phosphoglycerate in glycolysis; may be non‐ functional derivative of a gene duplication event 1.4000 0.0002 NA YMR090W Putative protein of unknown function with similarity to DTDP‐glucose 4,6‐dehydratases; GFP‐fusion protein localizes to the cytoplasm; up‐ regulated in response to the fungicide mancozeb; not essential for viability 1.3741 0.0022 NA YDR018C Probable membrane protein with three predicted transmembrane domains; homologous to Ybr042cp, similar to C. elegans F55A11.5 and maize 1‐acyl‐glycerol‐3‐phosphate acyltransferase 1.3658 0.0013 AIM17 YHL021C Putative protein of unknown function; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐ throughput studies; null mutant displays reduced frequency of mitochondrial genome loss 1.3625 0.0073 GDH3 YAL062W NADP(+)‐dependent glutamate dehydrogenase, synthesizes glutamate from ammonia and alpha‐ketoglutarate; rate of alpha‐ ketoglutarate utilization differs from Gdh1p; expression regulated by nitrogen and carbon sources 1.3458 0.0091 STF2 YGR008C Protein involved in regulation of the mitochondrial F1F0‐ATP synthase; Stf1p and Stf2p may act as stabilizing factors that enhance inhibitory action of the Inh1p protein 1.3182 0.0109 YPT53 YNL093W 1.3130 0.0042 HXT4 YHR092C Rab family GTPase, similar to Ypt51p and Ypt52p and to mammalian rab5; required for vacuolar protein sorting and endocytosis High‐affinity glucose transporter of the major facilitator superfamily, expression is induced by low levels of glucose and repressed by high levels of glucose 1.2365 0.0308 COS12 YGL263W 1.2307 0.0269 Symbols ORF Description CYB2 YML054C NA Protein of unknown function, member of the DUP380 subfamily of conserved, often subtelomerically‐encoded proteins 265 log2(msn5 .Tf/WT.Tf) adj.p.v al Alkaline ceramidase that also has reverse (CoA‐independent) ceramide synthase activity, catalyzes both breakdown and synthesis of phytoceramide; overexpression confers fumonisin B1 resistance Putative NADP(+) coupled glycerol dehydrogenase, proposed to be involved in an alternative pathway for glycerol catabolism; also has mRNA binding activity; member of the aldo‐keto reductase (AKR) family 1.1755 0.0109 1.1738 0.0015 YCL026C‐B Putative protein of unknown function; similar to bacterial nitroreductases; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm and nucleus; protein becomes insoluble upon intracellular iron depletion 1.1731 0.0065 PHO8 YDR481C Repressible alkaline phosphatase, a glycoprotein localized to the vacuole; regulated by levels of inorganic phosphate and by a system consisting of Pho4p, Pho9p, Pho80p, Pho81p and Pho85p; dephosphorylates phosphotyrosyl peptides 1.1708 0.0005 ARO9 YHR137W 1.1616 0.0464 BOP2 DUR1,2 YLR267W YBR208C Aromatic aminotransferase II, catalyzes the first step of tryptophan, phenylalanine, and tyrosine catabolism Protein of unknown function Urea amidolyase, contains both urea carboxylase and allophanate hydrolase activities, degrades urea to CO2 and NH3; expression sensitive to nitrogen catabolite repression and induced by allophanate, an intermediate in allantoin degradation 1.1607 1.1596 0.0398 0.0070 PRX1 YBL064C Mitochondrial peroxiredoxin (1‐Cys Prx) with thioredoxin peroxidase activity, has a role in reduction of hydroperoxides; reactivation requires Trr2p and glutathione; induced during respiratory growth and oxidative stress; phosphorylated 1.1478 0.0014 NA YKL107W 1.1472 0.0096 MAL31 YBR298C Putative protein of unknown function; proposed to be a palmitoylated membrane protein Maltose permease, high‐affinity maltose transporter (alpha‐glucoside transporter); encoded in the MAL3 complex locus; member of the 12 transmembrane domain superfamily of sugar transporters; functional in genomic reference strain S288C 1.1389 0.0016 USV1 YPL230W Putative transcription factor containing a C2H2 zinc finger; mutation affects transcriptional regulation of genes involved in growth on non‐ fermentable carbon sources, response to salt stress and cell wall biosynthesis 1.1211 0.0088 MAL32 YBR299W Maltase (alpha‐D‐glucosidase), inducible protein involved in maltose catabolism; encoded in the MAL3 complex locus; functional in genomic reference strain S288C; hydrolyzes the disaccharides maltose, turanose, maltotriose, and sucrose 1.1189 0.0147 DAL1 YIR027C Allantoinase, converts allantoin to allantoate in the first step of allantoin degradation; expression sensitive to nitrogen catabolite repression 1.1186 0.0032 REG2 YBR050C Regulatory subunit of the Glc7p type‐1 protein phosphatase; involved with Reg1p, Glc7p, and Snf1p in regulation of glucose‐ repressible genes, also involved in glucose‐induced proteolysis of maltose permease 1.0843 0.0047 NA YHL048C‐A 1.0822 0.0022 MOH1 YBL049W Putative protein of unknown function; identified by expression profiling and mass spectrometry Protein of unknown function, has homology to kinase Snf7p; not required for growth on nonfermentable carbon sources; essential for survival in stationary phase 1.0763 0.0080 Symbols ORF Description YPC1 YBR183W GCY1 YOR120W HBN1 266 log2(msn5 .Tf/WT.Tf) adj.p.v al Putative transmembrane protein involved in export of ammonia, a starvation signal that promotes cell death in aging colonies; phosphorylated in mitochondria; member of the TC 9.B.33 YaaH family; homolog of Ady2p and Y. lipolytica Gpr1p 1.0748 0.0186 Putative channel‐like protein; similar to Fps1p; mediates passive diffusion of glycerol in the presence of ethanol Vacuolar membrane polyphosphate polymerase; subunit of the vacuolar transporter chaperone (VTC) complex involved in synthesis and transfer of polyP to the vacuole; regulates membrane trafficking; role in non‐autophagic vacuolar fusion 1.0654 0.0014 1.0636 0.0023 Pyruvate kinase that appears to be modulated by phosphorylation; PYK2 transcription is repressed by glucose, and Pyk2p may be active under low glycolytic flux 1.0456 0.0060 YKL221W Protein with similarity to mammalian monocarboxylate permeases, which are involved in transport of monocarboxylic acids across the plasma membrane; mutant is not deficient in monocarboxylate transport 1.0449 0.0066 NA YOR289W Putative protein of unknown function; transcription induced by the unfolded protein response; green fluorescent protein (GFP)‐fusion protein localizes to both the cytoplasm and the nucleus 1.0422 0.0036 NA YDR391C Putative protein of unknown function, possibly involved in zinc homeostasis; Bdf1p‐dependent transcription induced by salt stress; green fluorescent protein (GFP)‐fusion protein localizes to both the cytoplasm and the nucleus 1.0397 0.0026 YPS6 YIR039C 1.0321 0.0029 PHO81 YGR233C Putative GPI‐anchored aspartic protease, member of the yapsin family of proteases involved in cell wall growth and maintenance Cyclin‐dependent kinase (CDK) inhibitor, regulates Pho80p‐Pho85p and Pcl7p‐Pho85p cyclin‐CDK complexes in response to phosphate levels; inhibitory activity for Pho80p‐Pho85p requires myo‐D‐inositol heptakisphosphate (IP7) generated by Vip1p 1.0245 0.0045 FMP33 YJL161W Putative protein of unknown function; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐ throughput studies 1.0212 0.0047 PET10 YKR046C Protein of unknown function that co‐purifies with lipid particles; expression pattern suggests a role in respiratory growth; computational analysis of large‐scale protein‐protein interaction data suggests a role in ATP/ADP exchange 1.0203 0.0052 NA YBL086C 1.0196 0.0026 CLD1 YGR110W Protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cell periphery Mitochondrial cardiolipin‐specific phospholipase; functions upstream of Taz1p to generate monolyso‐cardiolipin; transcription increases upon genotoxic stress; involved in restricting Ty1 transposition; has homology to mammalian CGI‐58 1.0169 0.0186 GAP1 YKR039W 1.0123 0.0109 NA YEL073C 1.0117 0.0070 KIN82 YCR091W 1.0036 0.0148 Symbols ORF Description ATO2 YNR002C NA YFL054C VTC4 YJL012C PYK2 YOR347C MCH2 General amino acid permease; localization to the plasma membrane is regulated by nitrogen source Putative protein of unknown function; located adjacent to ARS503 and the telomere on the left arm of chromosome V; regulated by inositol/choline Putative serine/threonine protein kinase implicated in the regulation of phospholipid asymmetry through the activation of phospholipid translocases (flippases) Lem3p‐Dnf1p/Dnf2p; similar to Fpk1p 267 log2(msn5 .Tf/WT.Tf) adj.p.v al Glycogen branching enzyme, involved in glycogen accumulation; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm in a punctate pattern 0.9895 0.0047 Mitochondrial phosphate carrier, imports inorganic phosphate into mitochondria; functionally redundant with Mir1p but less abundant than Mir1p under normal conditions; expression is induced at high temperature Putative protein of unknown function; YKL151C promoter contains STREs (stress response elements) and expression is induced by heat shock or methyl methanesulfonate; green fluorescent protein (GFP)‐ fusion protein localizes to the cytoplasm 0.9859 0.0142 0.9750 0.0028 0.9655 0.0089 0.9614 0.9590 0.0411 0.0036 0.9533 0.0204 Neutral trehalase, degrades trehalose; required for thermotolerance and may mediate resistance to other cellular stresses; may be phosphorylated by Cdc28p 0.9406 0.0022 YLR270W Non‐essential hydrolase involved in mRNA decapping, may function in a feedback mechanism to regulate deadenylation, contains pyrophosphatase activity and a HIT (histidine triad) motif; interacts with neutral trehalase Nth1p 0.9212 0.0077 STF1 YDL130W‐A Protein involved in regulation of the mitochondrial F1F0‐ATP synthase; Stf1p and Stf2p may act as stabilizing factors that enhance inhibitory action of the Inh1p protein 0.9205 0.0109 DAL3 YIR032C Ureidoglycolate hydrolase, converts ureidoglycolate to glyoxylate and urea in the third step of allantoin degradation; expression sensitive to nitrogen catabolite repression 0.9195 0.0266 NA YMR206W 0.9149 0.0098 BNA2 YJR078W Putative protein of unknown function; YMR206W is not an essential gene Putative tryptophan 2,3‐dioxygenase or indoleamine 2,3‐ dioxygenase, required for de novo biosynthesis of NAD from tryptophan via kynurenine; interacts genetically with telomere capping gene CDC13; regulated by Hst1p and Aftp 0.9143 0.0091 GSY1 YFR015C 0.9140 0.0091 NA NA NA NA YER079W YMR262W 0.9093 0.9050 0.9024 0.0102 0.0132 0.0046 NA YHL044W 0.8932 0.0014 HXT9 YJL219W 0.8838 0.0153 Symbols ORF Description GLC3 YEL011W PIC2 YER053C NA YKL151C SUE1 YPR151C NA LAP4 NA YKL103C TSL1 YML100W NTH1 YDR001C DCS1 Mitochondrial protein required for degradation of unstable forms of cytochrome c NA Vacuolar aminopeptidase yscI; zinc metalloproteinase that belongs to the peptidase family M18; often used as a marker protein in studies of autophagy and cytosol to vacuole targeting (CVT) pathway Large subunit of trehalose 6‐phosphate synthase (Tps1p)/phosphatase (Tps2p) complex, which converts uridine‐5'‐ diphosphoglucose and glucose 6‐phosphate to trehalose, similar to Tps3p and may share function; mutant has aneuploidy tolerance Glycogen synthase with similarity to Gsy2p, the more highly expressed yeast homolog; expression induced by glucose limitation, nitrogen starvation, environmental stress, and entry into stationary phase NA Putative protein of unknown function Protein of unknown function; interacts weakly with Knr4p; YMR262W is not an essential gene Putative integral membrane protein, member of DUP240 gene family; green fluorescent protein (GFP)‐fusion protein localizes to the plasma membrane in a punctate pattern Putative hexose transporter that is nearly identical to Hxt11p, has similarity to major facilitator superfamily (MFS) transporters, expression of HXT9 is regulated by transcription factors Pdr1p and Pdr3p 268 log2(msn5 .Tf/WT.Tf) adj.p.v al Meiosis‐specific component of sister chromatid cohesion complex; maintains cohesion between sister chromatids during meiosis I; maintains cohesion between centromeres of sister chromatids until meiosis II; homolog of S. pombe Rec8p 0.8821 0.0135 YLR281C Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to mitochondria; YLR281C is not an essential gene 0.8820 0.0021 UBP11 YKR098C 0.8811 0.0092 NA YNR034W‐A 0.8799 0.0185 HXK1 YFR053C Ubiquitin‐specific protease that cleaves ubiquitin from ubiquitinated proteins Putative protein of unknown function; expression is regulated by Msn2p/Msn4p Hexokinase isoenzyme 1, a cytosolic protein that catalyzes phosphorylation of glucose during glucose metabolism; expression is highest during growth on non‐glucose carbon sources; glucose‐ induced repression involves the hexokinase Hxk2p 0.8792 0.0206 EMI2 YDR516C Non‐essential protein of unknown function required for transcriptional induction of the early meiotic‐specific transcription factor IME1; required for sporulation; expression is regulated by glucose‐repression transcription factors Mig1/2p 0.8772 0.0114 ROM1 YGR070W GDP/GTP exchange protein (GEP) for Rho1p; mutations are synthetically lethal with mutations in rom2, which also encodes a GEP 0.8720 0.0141 NA YNL200C Putative protein of unknown function; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐ throughput studies 0.8719 0.0025 FMP46 YKR049C Putative redox protein containing a thioredoxin fold; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐throughput studies 0.8706 0.0031 NA YHR138C Putative protein of unknown function; has similarity to Pbi2p; double null mutant lacking Pbi2p and Yhr138p exhibits highly fragmented vacuoles 0.8657 0.0047 CAR1 YPL111W 0.8615 0.0081 AMS1 YGL156W Arginase, responsible for arginine degradation, expression responds to both induction by arginine and nitrogen catabolite repression; disruption enhances freeze tolerance Vacuolar alpha mannosidase, involved in free oligosaccharide (fOS) degradation; delivered to the vacuole in a novel pathway separate from the secretory pathway 0.8532 0.0206 JID1 YPR061C Probable Hsp40p co‐chaperone, has a DnaJ‐like domain and appears to be involved in ER‐associated degradation of misfolded proteins containing a tightly folded cytoplasmic domain; inhibits replication of Brome mosaic virus in S. cerevisiae 0.8513 0.0015 THI13 YDL244W Protein involved in synthesis of the thiamine precursor hydroxymethylpyrimidine (HMP); member of a subtelomeric gene family including THI5, THI11, THI12, and THI13 0.8493 0.0041 GDB1 YPR184W 0.8449 0.0189 DGR2 YKL121W 0.8397 0.0091 ISF1 YMR081C 0.8299 0.0487 COX20 YDR231C Glycogen debranching enzyme containing glucanotranferase and alpha‐1,6‐amyloglucosidase activities, required for glycogen degradation; phosphorylated in mitochondria Protein of unknown function; null mutant is resistant to 2‐deoxy‐D‐ glucose and displays abnormally elongated buds Serine‐rich, hydrophilic protein with similarity to Mbr1p; overexpression suppresses growth defects of hap2, hap3, and hap4 mutants; expression is under glucose control; cotranscribed with NAM7 in a cyp1 mutant Mitochondrial inner membrane protein, required for proteolytic processing of Cox2p and its assembly into cytochrome c oxidase 0.8294 0.0024 Symbols ORF Description REC8 YPR007C NA 269 log2(msn5 .Tf/WT.Tf) adj.p.v al Non‐essential protein of unknown function Glyoxylate reductase; null mutation results in increased biomass after diauxic shift; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐throughput studies Xylulokinase, converts D‐xylulose and ATP to xylulose 5‐phosphate and ADP; rate limiting step in fermentation of xylulose; required for xylose fermentation by recombinant S. cerevisiae strains cAMP‐dependent protein kinase catalytic subunit; promotes vegetative growth in response to nutrients via the Ras‐cAMP signaling pathway; inhibited by regulatory subunit Bcy1p in the absence of cAMP; partially redundant with Tpk2p and Tpk3p 0.8267 0.8265 0.0071 0.0171 0.8165 0.0131 0.8118 0.0142 YCL040W Glucokinase, catalyzes the phosphorylation of glucose at C6 in the first irreversible step of glucose metabolism; one of three glucose phosphorylating enzymes; expression regulated by non‐fermentable carbon sources 0.8064 0.0054 PIG2 YIL045W 0.7987 0.0210 YAK1 YJL141C Putative type‐1 protein phosphatase targeting subunit that tethers Glc7p type‐1 protein phosphatase to Gsy2p glycogen synthase Serine‐threonine protein kinase that is part of a glucose‐sensing system involved in growth control in response to glucose availability; translocates from the cytoplasm to the nucleus and phosphorylates Pop2p in response to a glucose signal 0.7940 0.0070 GSP2 YOR185C GTP binding protein (mammalian Ranp homolog) involved in the maintenance of nuclear organization, RNA processing and transport; interacts with Kap121p, Kap123p and Pdr6p (karyophilin betas); Gsp1p homolog that is not required for viability 0.7909 0.0144 RTC2 YBR147W Protein of unknown function; identified in a screen for mutants with decreased levels of rDNA transcription; detected in highly purified mitochondria; null mutant suppresses cdc13‐1; similar to a G‐protein coupled receptor from S. pombe 0.7881 0.0105 NA YLR345W Similar to 6‐phosphofructo‐2‐kinase/fructose‐2,6‐bisphosphatase enzymes responsible for the metabolism of fructoso‐2,6‐ bisphosphate; mRNA expression is repressed by the Rfx1p‐Tup1p‐ Ssn6p repressor complex; YLR345W is not an essential gene 0.7815 0.0065 GSY2 YLR258W Glycogen synthase, similar to Gsy1p; expression induced by glucose limitation, nitrogen starvation, heat shock, and stationary phase; activity regulated by cAMP‐dependent, Snf1p and Pho85p kinases as well as by the Gac1p‐Glc7p phosphatase 0.7814 0.0337 NA NA YBR056W YGR174W‐A 0.7795 0.7790 0.0091 0.0419 CMC4 YMR194C‐B Putative cytoplasmic protein of unknown function Putative protein of unknown function; predicted to have a role in cell budding based on computational "guilt by association" analysis Protein that localizes to the mitochondrial intermembrane space via the Mia40p‐Erv1p system; contains twin cysteine‐x(9)‐cysteine motifs 0.7790 0.0090 NA YJR008W 0.7728 0.0227 NA YAL016C‐B 0.7654 0.0224 VID24 YBR105C Putative protein of unknown function; expression repressed by inosine and choline in an Opi1p‐dependent manner; expression induced by mild heat‐stress on a non‐fermentable carbon source. Dubious open reading frame unlikely to encode a protein, based on available experimental and comparative sequence data Peripheral membrane protein located at Vid (vacuole import and degradation) vesicles; regulates fructose‐1,6‐bisphosphatase (FBPase) targeting to the vacuole; promotes proteasome‐dependent catabolite degradation of FBPase 0.7647 0.0065 PTK2 YJR059W 0.7631 0.0096 Symbols ORF Description ECM8 GOR1 YBR076W YNL274C XKS1 YGR194C TPK1 YJL164C GLK1 Putative serine/threonine protein kinase involved in regulation of ion transport across plasma membrane; enhances spermine uptake 270 log2(msn5 .Tf/WT.Tf) adj.p.v al 0.7585 0.0467 0.7576 0.0115 0.7569 0.0416 0.7496 0.0110 Subunit of the mitochondrial inner membrane peptidase, which is required for maturation of mitochondrial proteins of the intermembrane space; Som1p facilitates cleavage of a subset of substrates; contains twin cysteine‐x9‐cysteine motifs 0.7431 0.0097 Ubiquitin‐like protein modifier, may function in modification of Sph1p and Hbt1p, functionally complemented by the human or S. pombe ortholog; mechanism of Hub1p adduct formation not yet clear Putative protein of unknown function; proposed to be involved in responding to environmental stresses; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐ throughput studies Putative protein of unknown function; identified by homology 0.7430 0.0068 0.7410 0.0051 0.7370 0.0054 YCR083W Mitochondrial thioredoxin, highly conserved oxidoreductase required to maintain the redox homeostasis of the cell, forms the mitochondrial thioredoxin system with Trr2p, redox state is maintained by both Trr2p and Glr1p 0.7367 0.0034 ALD3 YMR169C 0.7357 0.0400 BNA1 YJR025C Cytoplasmic aldehyde dehydrogenase, involved in beta‐alanine synthesis; uses NAD+ as the preferred coenzyme; very similar to Ald2p; expression is induced by stress and repressed by glucose 3‐hydroxyanthranilic acid dioxygenase, required for the de novo biosynthesis of NAD from tryptophan via kynurenine; expression regulated by Hst1p 0.7334 0.0034 NA YNL217W Putative protein of unknown function; weak sequence similarity to bis (5'‐nucleotidyl)‐tetraphosphatases; (GFP)‐fusion protein localizes to the vacuole; null mutant is highly sensitive to azaserine and resistant to sodium‐O‐vandate 0.7263 0.0091 TPS3 YMR261C 0.7217 0.0060 EMP46 YLR080W 0.7217 0.0292 NA YGL185C Regulatory subunit of trehalose‐6‐phosphate synthase/phosphatase complex, which synthesizes the storage carbohydrate trehalose; expression is induced by stress conditions and repressed by the Ras‐ cAMP pathway Integral membrane component of endoplasmic reticulum‐derived COPII‐coated vesicles, which function in ER to Golgi transport Putative protein with sequence similarity to hydroxyacid dehydrogenases; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm 0.7171 0.0093 DOG2 YHR043C 2‐deoxyglucose‐6‐phosphate phosphatase, member of a family of low molecular weight phosphatases, similar to Dog1p, induced by oxidative and osmotic stress, confers 2‐deoxyglucose resistance when overexpressed 0.7164 0.0134 AGP1 YCL025C Low‐affinity amino acid permease with broad substrate range, involved in uptake of asparagine, glutamine, and other amino acids; expression is regulated by the SPS plasma membrane amino acid sensor system (Ssy1p‐Ptr3p‐Ssy5p) 0.7129 0.0085 Symbols ORF Description NA YDR379C‐A MAL11 YGR289C YTP1 YNL237W ECM4 YKR076W Protein involved in the assembly of the mitochondrial succinate dehydrogenase complex; mutations in human ortholog SDHAF1 are associated with infantile leukoencephalopathy Inducible high‐affinity maltose transporter (alpha‐glucoside transporter); encoded in the MAL1 complex locus; broad substrate specificity that includes maltotriose; required for isomaltose utilization Probable type‐III integral membrane protein of unknown function, has regions of similarity to mitochondrial electron transport proteins Omega class glutathione transferase; not essential; similar to Ygr154cp; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm SOM1 YEL059C‐A HUB1 YNR032C‐A FMP40 YPL222W NA YFR012W‐A TRX3 271 log2(msn5 .Tf/WT.Tf) adj.p.v al L‐ornithine transaminase (OTAse), catalyzes the second step of arginine degradation, expression is dually‐regulated by allophanate induction and a specific arginine induction process; not nitrogen catabolite repression sensitive 0.7105 0.0246 YDR185C Mitochondrial protein of unknown function; similar to Ups1p and Ups2p which are involved in regulation of mitochondrial cardiolipin and phosphatidylethanolamine levels; null is viable but interacts synthetically with ups1 and ups2 mutations 0.7060 0.0111 CMK2 YOL016C Calmodulin‐dependent protein kinase; may play a role in stress response, many CA++/calmodulan dependent phosphorylation substrates demonstrated in vitro, amino acid sequence similar to Cmk1p and mammalian Cam Kinase II 0.7045 0.0138 OYE3 YPL171C Conserved NADPH oxidoreductase containing flavin mononucleotide (FMN), homologous to Oye2p with different ligand binding and catalytic properties; has potential roles in oxidative stress response and programmed cell death 0.7035 0.0308 VTC1 YER072W Subunit of the vacuolar transporter chaperone (VTC) complex involved in membrane trafficking, vacuolar polyphosphate accumulation, microautophagy and non‐autophagic vacuolar fusion; also has mRNA binding activity 0.7030 0.0060 UGX2 YDL169C 0.6974 0.0285 PNC1 YGL037C Protein of unknown function, transcript accumulates in response to any combination of stress conditions Nicotinamidase that converts nicotinamide to nicotinic acid as part of the NAD(+) salvage pathway, required for life span extension by calorie restriction; PNC1 expression responds to all known stimuli that extend replicative life span 0.6966 0.0181 NGL3 YML118W Putative endonuclease, has a domain similar to a magnesium‐ dependent endonuclease motif in mRNA deadenylase Ccr4p; similar to Ngl1p and Ngl2p 0.6952 0.0328 NA NA YBR230W‐A YPL247C 0.6950 0.6893 0.0242 0.0170 GAL3 YDR009W Putative protein of unknown function Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm and nucleus; similar to the petunia WD repeat protein an11; overexpression causes a cell cycle delay or arrest Transcriptional regulator involved in activation of the GAL genes in response to galactose; forms a complex with Gal80p to relieve Gal80p inhibition of Gal4p; binds galactose and ATP but does not have galactokinase activity 0.6848 0.0225 MIC14 YDR031W 0.6802 0.0104 UBP9 YER098W 0.6722 0.0109 TMA17 YDL110C Mitochondrial intermembrane space protein, required for normal oxygen consumption; contains twin cysteine‐x9‐cysteine motifs Ubiquitin carboxyl‐terminal hydrolase, ubiquitin‐specific protease that cleaves ubiquitin‐protein fusions Protein of unknown function that associates with ribosomes; heterozygous deletion demonstrated increases in chromosome instability in a rad9 deletion background; protein abundance is decreased upon intracellular iron depletion 0.6718 0.0292 VPS73 YGL104C 0.6713 0.0190 VTC2 YFL004W Mitochondrial protein; mutation affects vacuolar protein sorting; putative transporter; member of the sugar porter family Subunit of the vacuolar transporter chaperone (VTC) complex involved in membrane trafficking, vacuolar polyphosphate accumulation, microautophagy and non‐autophagic vacuolar fusion 0.6706 0.0292 NA YMR105W‐ A Putative protein of unknown function 0.6697 0.0476 Symbols ORF Description CAR2 YLR438W UPS3 272 log2(msn5 .Tf/WT.Tf) adj.p.v al Putative protein of unknown function; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐ throughput studies 0.6671 0.0091 YER062C One of two redundant DL‐glycerol‐3‐phosphatases (RHR2/GPP1 encodes the other) involved in glycerol biosynthesis; induced in response to hyperosmotic stress and oxidative stress, and during the diauxic transition 0.6665 0.0041 GPX1 YKL026C 0.6663 0.0079 NTH2 YBR001C 0.6662 0.0054 NA YGR226C Phospholipid hydroperoxide glutathione peroxidase induced by glucose starvation that protects cells from phospholipid hydroperoxides and nonphospholipid peroxides during oxidative stress Putative neutral trehalase, required for thermotolerance and may mediate resistance to other cellular stresses Dubious open reading frame, unlikely to encode a protein; not conserved in closely related Saccharomyces species; overlaps significantly with a verified ORF, AMA1/YGR225W 0.6654 0.0104 UBC12 YLR306W Enzyme that mediates the conjugation of Rub1p, a ubiquitin‐like protein, to other proteins; related to E2 ubiquitin‐conjugating enzymes 0.6566 0.0086 SDH4 YDR178W Membrane anchor subunit of succinate dehydrogenase (Sdh1p, Sdh2p, Sdh3p, Sdh4p), which couples the oxidation of succinate to the transfer of electrons to ubiquinone as part of the TCA cycle and the mitochondrial respiratory chain 0.6563 0.0090 GRX1 YCL035C Hydroperoxide and superoxide‐radical responsive heat‐stable glutathione‐dependent disulfide oxidoreductase with active site cysteine pair; protects cells from oxidative damage 0.6534 0.0171 DAN4 YJR151C Cell wall mannoprotein with similarity to Tir1p, Tir2p, Tir3p, and Tir4p; expressed under anaerobic conditions, completely repressed during aerobic growth 0.6460 0.0091 YSC84 YHR016C 0.6443 0.0109 NA YBR085C‐A 0.6425 0.0165 PUT1 YLR142W Actin‐binding protein involved in bundling of actin filaments and endocytosis of actin cortical patches; activity stimulated by Las17p; contains SH3 domain similar to Rvs167p Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm and to the nucleus Proline oxidase, nuclear‐encoded mitochondrial protein involved in utilization of proline as sole nitrogen source; PUT1 transcription is induced by Put3p in the presence of proline and the absence of a preferred nitrogen source 0.6412 0.0090 ATG34 YOL083W Receptor protein involved in selective autophagy during starvation; specifically involved in the transport of cargo protein alpha‐ mannosidase (Ams1p); Atg19p paralog 0.6396 0.0253 NA YNR014W 0.6388 0.0292 PMC1 YGL006W Putative protein of unknown function; expression is cell‐cycle regulated, Azf1p‐dependent, and heat‐inducible Vacuolar Ca2+ ATPase involved in depleting cytosol of Ca2+ ions; prevents growth inhibition by activation of calcineurin in the presence of elevated concentrations of calcium; similar to mammalian PMCA1a 0.6367 0.0170 ICY1 YMR195W 0.6366 0.0073 SFC1 YJR095W Protein of unknown function, required for viability in rich media of cells lacking mitochondrial DNA; mutants have an invasive growth defect with elongated morphology; induced by amino acid starvation Mitochondrial succinate‐fumarate transporter, transports succinate into and fumarate out of the mitochondrion; required for ethanol and acetate utilization 0.6324 0.0403 Symbols ORF Description NA YJL133C‐A HOR2 273 log2(msn5 .Tf/WT.Tf) adj.p.v al Protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the ER; YBR287W is not an essential gene Subunit 8 of ubiquinol cytochrome‐c reductase complex, which is a component of the mitochondrial inner membrane electron transport chain; oriented facing the intermembrane space; expression is regulated by Abf1p and Cpf1p 0.6307 0.0091 0.6294 0.0081 YMR291W Putative kinase of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm and nucleus; YMR291W is not an essential gene 0.6287 0.0045 VID30 YGL227W Protein involved in proteasome‐dependent catabolite degradation of fructose‐1,6‐bisphosphatase (FBPase); binds FBPase; shifts the balance of nitrogen metabolism toward glutamate production; localizes to the nucleus and the cytoplasm 0.6285 0.0150 ZTA1 YBR046C NADPH‐dependent quinone reductase, GFP‐tagged protein localizes to the cytoplasm and nucleus; has similarity to E. coli quinone oxidoreductase and to human zeta‐crystallin 0.6240 0.0104 REE1 YJL217W Cytoplasmic protein involved in the regulation of enolase (ENO1); mRNA expression is induced by calcium shortage, copper deficiency (via Mac1p) and the presence of galactose (via Gal4p); mRNA expression is also regulated by the cell cycle 0.6214 0.0400 OSH2 YDL019C Member of an oxysterol‐binding protein family with seven members in S. cerevisiae; family members have overlapping, redundant functions in sterol metabolism and collectively perform a function essential for viability 0.6211 0.0113 COX5B YIL111W Subunit Vb of cytochrome c oxidase, which is the terminal member of the mitochondrial inner membrane electron transport chain; predominantly expressed during anaerobic growth while its isoform Va (Cox5Ap) is expressed during aerobic growth 0.6204 0.0337 DCG1 YIR030C Protein of unknown function, expression is sensitive to nitrogen catabolite repression and regulated by Dal80p; contains transmembrane domain 0.6182 0.0166 NA YLR177W Putative protein of unknown function; phosphorylated by Dbf2p‐ Mob1p in vitro; some strains contain microsatellite polymophisms at this locus; YLR177W is not an essential gene 0.6074 0.0127 ODC1 YPL134C 0.6048 0.0144 NA YJL213W Mitochondrial inner membrane transporter, exports 2‐oxoadipate and 2‐oxoglutarate from the mitochondrial matrix to the cytosol for lysine and glutamate biosynthesis and lysine catabolism; suppresses, in multicopy, an fmc1 null mutation Protein of unknown function that may interact with ribosomes; periodically expressed during the yeast metabolic cycle; phosphorylated in vitro by the mitotic exit network (MEN) kinase complex, Dbf2p/Mob1p 0.6020 0.0253 ALG14 YBR070C Component of UDP‐GlcNAc transferase required for the second step of dolichyl‐linked oligosaccharide synthesis; anchors the catalytic subunit Alg13p to the ER membrane; similar to bacterial and human glycosyltransferases 0.6006 0.0080 Symbols ORF Description NA YBR287W QCR8 YJL166W NA 274 APPENDIX N: Down-regulated genes in msn5Δ cells in amino acid starvation condition This section includes the list of transcriptionally differentially expressed genes (cutoff conditions: log2 FC 0.6 and adjusted p-value < 0.05) in msn5Δ cells in amino acid starvation condition. Down-regulated genes in msn5Δ cells in amino acid starvation condition Symbols ORF Description MSN5 YDR335W MF(ALPHA) 2 YGL089C PCL1 YNL289W PRM1 YNL279W YOX1 YML027W BAR1 YIL015W FUS1 YCL027W IMD1 YAR073W Karyopherin involved in nuclear import and export of proteins, including import of replication protein A and export of Swi6p, Far1p, and Pho4p; required for re‐export of mature tRNAs after their retrograde import from the cytoplasm Mating pheromone alpha‐factor, made by alpha cells; interacts with mating type a cells to induce cell cycle arrest and other responses leading to mating; also encoded by MF(ALPHA)1, which is more highly expressed than MF(ALPHA)2 Cyclin, interacts with cyclin‐dependent kinase Pho85p; member of the Pcl1,2‐like subfamily, involved in the regulation of polarized growth and morphogenesis and progression through the cell cycle; localizes to sites of polarized cell growth Pheromone‐regulated multispanning membrane protein involved in membrane fusion during mating; predicted to have 5 transmembrane segments and a coiled coil domain; localizes to the shmoo tip; regulated by Ste12p Homeodomain‐containing transcriptional repressor, binds to Mcm1p and to early cell cycle boxes (ECBs) in the promoters of cell cycle‐ regulated genes expressed in M/G1 phase; expression is cell cycle‐ regulated; potential Cdc28p substrate Aspartyl protease secreted into the periplasmic space of mating type a cells, helps cells find mating partners, cleaves and inactivates alpha factor allowing cells to recover from alpha‐factor‐induced cell cycle arrest Membrane protein localized to the shmoo tip, required for cell fusion; expression regulated by mating pheromone; proposed to coordinate signaling, fusion, and polarization events required for fusion; potential Cdc28p substrate Nonfunctional protein with homology to IMP dehydrogenase; probable pseudogene, located close to the telomere; is not 275 log2(msn5 .Ts/WT.Ts) adj.p.v al ‐7.9130 0.0000 ‐4.6487 0.0000 ‐3.2756 0.0002 ‐2.9190 0.0000 ‐2.7175 0.0007 ‐2.3923 0.0004 ‐2.1300 0.0004 ‐2.1086 0.0025 Symbols ORF FIG1 YBR040W YHP1 YDR451C SAG1 YJR004C ANS1 YHR126C AGA1 YNR044W WSC4 YHL028W AGA2 YGL032C ASG7 YJL170C NA YER085C IMD2 YHR216W CSI2 YOL007C TOS6 YNL300W PRM6 YML047C PHM7 YOL084W HO YDL227C DAN1 YJR150C NA YAR066W RIM9 YMR063W SFG1 YOR315W Description expressed at detectable levels; YAR073W and YAR075W comprise a continuous reading frame in some strains of S. cerevisiae Integral membrane protein required for efficient mating; may participate in or regulate the low affinity Ca2+ influx system, which affects intracellular signaling and cell‐cell fusion during mating One of two homeobox transcriptional repressors (see also Yox1p), that bind to Mcm1p and to early cell cycle box (ECB) elements of cell cycle regulated genes, thereby restricting ECB‐mediated transcription to the M/G1 interval Alpha‐agglutinin of alpha‐cells, binds to Aga1p during agglutination, N‐terminal half is homologous to the immunoglobulin superfamily and contains binding site for a‐agglutinin, C‐terminal half is highly glycosylated and contains GPI anchor Putative protein of unknown function; transcription dependent upon Azf1p Anchorage subunit of a‐agglutinin of a‐cells, highly O‐glycosylated protein with N‐terminal secretion signal and C‐terminal signal for addition of GPI anchor to cell wall, linked to adhesion subunit Aga2p via two disulfide bonds ER membrane protein involved in the translocation of soluble secretory proteins and insertion of membrane proteins into the ER membrane; may also have a role in the stress response but has only partial functional overlap with WSC1‐3 Adhesion subunit of a‐agglutinin of a‐cells, C‐terminal sequence acts as a ligand for alpha‐agglutinin (Sag1p) during agglutination, modified with O‐linked oligomannosyl chains, linked to anchorage subunit Aga1p via two disulfide bonds Protein that regulates signaling from a G protein beta subunit Ste4p and its relocalization within the cell; specific to a‐cells and induced by alpha‐factor Putative protein of unknown function Inosine monophosphate dehydrogenase, catalyzes the rate‐limiting step in GTP biosynthesis, expression is induced by mycophenolic acid resulting in resistance to the drug, expression is repressed by nutrient limitatio Protein of unknown function; green fluorescent protein (GFP)‐ fusion protein localizes to the mother side of the bud neck and the vacuole; YOL007C is not an essential gene Glycosylphosphatidylinositol‐dependent cell wall protein, expression is periodic and decreases in respone to ergosterol perturbation or upon entry into stationary phase; depletion increases resistance to lactic acid Pheromone‐regulated protein, predicted to have 2 transmembrane segments; regulated by Ste12p during mating Protein of unknown function, expression is regulated by phosphate levels; green fluorescent protein (GFP)‐fusion protein localizes to the cell periphery and vacuole Site‐specific endonuclease required for gene conversion at the MAT locus (homothallic switching) through the generation of a ds DNA break; expression restricted to mother cells in late G1 as controlled by Swi4p‐Swi6p, Swi5p and Ash1p Cell wall mannoprotein with similarity to Tir1p, Tir2p, Tir3p, and Tir4p; expressed under anaerobic conditions, completely repressed during aerobic growth Putative GPI protein Protein of unknown function, involved in the proteolytic activation of Rim101p in response to alkaline pH; has similarity to A. nidulans PalI; putative membrane protein Nuclear protein, putative transcription factor required for growth of 276 log2(msn5 .Ts/WT.Ts) adj.p.v al ‐1.9293 0.0075 ‐1.9256 0.0007 ‐1.9071 0.0001 ‐1.8395 0.0039 ‐1.8349 0.0004 ‐1.7878 0.0072 ‐1.7575 0.0034 ‐1.7416 0.0021 ‐1.7129 0.0013 ‐1.7000 0.0007 ‐1.6657 0.0003 ‐1.6583 0.0009 ‐1.6562 0.0183 ‐1.6327 0.0200 ‐1.6270 0.0019 ‐1.6217 0.0061 ‐1.6055 0.0009 ‐1.5709 0.0062 ‐1.5696 0.0042 Symbols ORF SCW10 YMR305C PLM2 YDR501W HUG1 YML058W‐A RPI1 YIL119C MFA1 YDR461W STE3 YKL178C CWP1 YKL096W ADH2 YMR303C SST2 YLR452C FAR1 YJL157C NA YIL082W‐A DTR1 YBR180W NA YAL018C NA YOR214C SPC24 YMR117C CRR1 YLR213C ALP1 YNL270C CLB2 YPR119W Description superficial pseudohyphae (which do not invade the agar substrate) but not for invasive pseudohyphal growth; may act together with Phd1p; potential Cdc28p substrate Cell wall protein with similarity to glucanases; may play a role in conjugation during mating based on mutant phenotype and its regulation by Ste12p Forkhead Associated domain containing protein and putative transcription factor found associated with chromatin; target of SBF transcription factor; induced in response to DNA damaging agents and deletion of telomerase; similar to TOS4 Protein involved in the Mec1p‐mediated checkpoint pathway that responds to DNA damage or replication arrest, transcription is induced by DNA damage Putative transcriptional regulator; overexpression suppresses the heat shock sensitivity of wild‐type RAS2 overexpression and also suppresses the cell lysis defect of an mpk1 mutation Mating pheromone a‐factor, made by a cells; interacts with alpha cells to induce cell cycle arrest and other responses leading to mating; biogenesis involves C‐terminal modification, N‐terminal proteolysis, and export; also encoded by MFA2 Receptor for a factor pheromone, couples to MAP kinase cascade to mediate pheromone response; transcribed in alpha cells and required for mating by alpha cells, ligand bound receptors endocytosed and recycled to the plasma membrane; GPCR Cell wall mannoprotein that localizes specifically to birth scars of daughter cells, linked to a beta‐1,3‐ and beta‐1,6‐glucan heteropolymer through a phosphodiester bond; required for propionic acid resistance Glucose‐repressible alcohol dehydrogenase II, catalyzes the conversion of ethanol to acetaldehyde; involved in the production of certain carboxylate esters; regulated by ADR1 GTPase‐activating protein for Gpa1p, regulates desensitization to alpha factor pheromone; also required to prevent receptor‐ independent signaling of the mating pathway; member of the RGS (regulator of G‐protein signaling) family Cyclin‐dependent kinase inhibitor that mediates cell cycle arrest in response to pheromone; also forms a complex with Cdc24p, Ste4p, and Ste18p that may specify the direction of polarized growth during mating; potential Cdc28p substrate Retrotransposon TYA Gag and TYB Pol genes; transcribed/translated as one unit; polyprotein is processed to make a nucleocapsid‐like protein (Gag), reverse transcriptase (RT), protease (PR), and integrase (IN); similar to retroviral genes Putative dityrosine transporter, required for spore wall synthesis; expressed during sporulation; member of the major facilitator superfamily (DHA1 family) of multidrug resistance transporters Putative protein of unknown function Putative protein of unknown function; YOR214C is not an essential gene Component of the evolutionarily conserved kinetochore‐associated Ndc80 complex (Ndc80p‐Nuf2p‐Spc24p‐Spc25p); involved in chromosome segregation, spindle checkpoint activity and kinetochore clustering Putative glycoside hydrolase of the spore wall envelope; required for normal spore wall assembly, possibly for cross‐linking between the glucan and chitosan layers; expressed during sporulation Arginine transporter; expression is normally very low and it is unclear what conditions would induce significant expression B‐type cyclin involved in cell cycle progression; activates Cdc28p to 277 log2(msn5 .Ts/WT.Ts) adj.p.v al ‐1.5512 0.0004 ‐1.5402 0.0004 ‐1.5216 0.0030 ‐1.5166 0.0111 ‐1.5020 0.0019 ‐1.4891 0.0001 ‐1.4830 0.0210 ‐1.4705 0.0020 ‐1.4700 0.0004 ‐1.4390 0.0002 ‐1.4350 0.0003 ‐1.4312 0.0001 ‐1.4045 0.0031 ‐1.3955 0.0182 ‐1.3870 0.0035 ‐1.3782 0.0062 ‐1.3741 0.0062 ‐1.3465 0.0033 Symbols ORF ECM11 YDR446W SVS1 YPL163C PTH1 YHR189W PMA2 YPL036W STE12 YHR084W NA YHL018W KAR4 YCL055W HES1 YOR237W RSB1 YOR049C NA YMR084W CDC6 YJL194W MSB2 YGR014W FRE8 YLR047C SMA1 YPL027W NA YLR042C PCL2 YDL127W NRM1 YNR009W NA YCL021W‐A Description promote the transition from G2 to M phase; accumulates during G2 and M, then targeted via a destruction box motif for ubiquitin‐ mediated degradation by the proteasome Non‐essential protein apparently involved in meiosis, GFP fusion protein is present in discrete clusters in the nucleus throughout mitosis; may be involved in maintaining chromatin structure Cell wall and vacuolar protein, required for wild‐type resistance to vanadate One of two (see also PTH2) mitochondrially‐localized peptidyl‐tRNA hydrolases; dispensable for respiratory growth on rich medium, but required for respiratory growth on minimal medium Plasma membrane H+‐ATPase, isoform of Pma1p, involved in pumping protons out of the cell; regulator of cytoplasmic pH and plasma membrane potential Transcription factor that is activated by a MAP kinase signaling cascade, activates genes involved in mating or pseudohyphal/invasive growth pathways; cooperates with Tec1p transcription factor to regulate genes specific for invasive growth Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to mitochondria and is induced in response to the DNA‐damaging agent MMS Transcription factor required for gene regulation in response to pheromones; also required during meiosis; exists in two forms, a slower‐migrating form more abundant during vegetative growth and a faster‐migrating form induced by pheromone Protein implicated in the regulation of ergosterol biosynthesis; one of a seven member gene family with a common essential function and non‐essential unique functions; similar to human oxysterol binding protein (OSBP) Suppressor of sphingoid long chain base (LCB) sensitivity of an LCB‐ lyase mutation; putative integral membrane transporter or flippase that may transport LCBs from the cytoplasmic side toward the extracytoplasmic side of the membrane Putative protein of unknown function; YMR084W and adjacent ORF YMR085W are merged in related strains Essential ATP‐binding protein required for DNA replication, component of the pre‐replicative complex (pre‐RC) which requires ORC to associate with chromatin and is in turn required for Mcm2‐7p DNA association; homologous to S. pombe Cdc18p Mucin family member involved in the Cdc42p‐ and MAP kinase‐ dependent filamentous growth signaling pathway; also functions as an osmosensor in parallel to the Sho1p‐mediated pathway; potential Cdc28p substrate Protein with sequence similarity to iron/copper reductases, involved in iron homeostasis; deletion mutant has iron deficiency/accumulation growth defects; expression increased in the absence of copper‐responsive transcription factor Mac1p Protein of unknown function involved in the assembly of the prospore membrane during sporulation; interacts with Spo14p Protein of unknown function; localizes to the cytoplasm; YLL042C is not an essential gene Cyclin, interacts with cyclin‐dependent kinase Pho85p; member of the Pcl1,2‐like subfamily, involved in the regulation of polarized growth and morphogenesis and progression through the cell cycle; localizes to sites of polarized cell growth Transcriptional co‐repressor of MBF (MCB binding factor)‐regulated gene expression; Nrm1p associates stably with promoters via MBF to repress transcription upon exit from G1 phase Putative protein of unknown function 278 log2(msn5 .Ts/WT.Ts) adj.p.v al ‐1.3128 0.0051 ‐1.3010 0.0040 ‐1.2941 0.0095 ‐1.2830 0.0032 ‐1.2510 0.0007 ‐1.2395 0.0083 ‐1.2261 0.0007 ‐1.2197 0.0019 ‐1.1971 0.0182 ‐1.1706 0.0117 ‐1.1641 0.0062 ‐1.1203 0.0015 ‐1.1190 0.0030 ‐1.1130 0.0102 ‐1.1123 0.0016 ‐1.0947 0.0016 ‐1.0944 0.0015 ‐1.0911 0.0030 Symbols ORF Description AQY1 YPR192W NA YOR316C‐A RTG1 YOL067C NFT1 YKR103W CLN1 YMR199W TIR1 YER011W Spore‐specific water channel that mediates the transport of water across cell membranes, developmentally controlled; may play a role in spore maturation, probably by allowing water outflow, may be involved in freeze tolerance Putative protein of unknown function; identified by fungal homology and RT‐PCR Transcription factor (bHLH) involved in interorganelle communication between mitochondria, peroxisomes, and nucleus Putative transporter of the multidrug resistance‐associated protein (MRP) subfamily; adjacent ORFs YKR103W and YKR104W are merged in different strain backgrounds. G1 cyclin involved in regulation of the cell cycle; activates Cdc28p kinase to promote the G1 to S phase transition; late G1 specific expression depends on transcription factor complexes, MBF (Swi6p‐ Mbp1p) and SBF (Swi6p‐Swi4p) Cell wall mannoprotein of the Srp1p/Tip1p family of serine‐alanine‐ rich proteins; expression is downregulated at acidic pH and induced by cold shock and anaerobiosis; abundance is increased in cells cultured without shaking Protein of unknown function, involved in filamentous growth DFG10 YIL049W FRM2 YCL026C‐A BSC5 YNR069C PPM2 YOL141W PLB2 YMR006C GPI19 YDR437W NA YMR279C NA YOL014W FUS2 YMR232W PRM2 YIL037C CLN2 YPL256C WHI5 YOR083W SLI15 YBR156C log2(msn5 .Ts/WT.Ts) adj.p.v al ‐1.0892 0.0048 ‐1.0829 0.0109 ‐1.0808 0.0025 ‐1.0804 0.0130 ‐1.0786 0.0012 ‐1.0773 0.0025 ‐1.0770 0.0005 Protein of unknown function, involved in the integration of lipid signaling pathways with cellular homeostasis; expression induced in cells treated with the mycotoxin patulin; has similarity to bacterial nitroreductases Protein of unknown function, ORF exhibits genomic organization compatible with a translational readthrough‐dependent mode of expression AdoMet‐dependent tRNA methyltransferase also involved in methoxycarbonylation; required for the synthesis of wybutosine (yW), a modified guanosine found at the 3'‐position adjacent to the anticodon of phe‐tRNA; similarity to Ppm1p Phospholipase B (lysophospholipase) involved in phospholipid metabolism; displays transacylase activity in vitro; overproduction confers resistance to lysophosphatidylcholine Subunit of GPI‐GlcNAc transferase involved in synthesis of N‐ acetylglucosaminyl phosphatidylinositol (GlcNAc‐PI), which is the first intermediate in glycosylphosphatidylinositol (GPI) anchor synthesis, shares similarity with mammalian PIG‐P Putative paralog of ATR1, but not required for boron tolerance; identified as a heat‐induced gene in a high‐throughout screen; YMR279C is not an essential gene Putative protein of unknown function ‐1.0733 0.0062 ‐1.0651 0.0110 ‐1.0581 0.0034 ‐1.0523 0.0034 ‐1.0492 0.0019 ‐1.0426 0.0015 ‐1.0419 0.0070 Cytoplasmic protein localized to the shmoo tip; required for the alignment of parental nuclei before nuclear fusion during mating Pheromone‐regulated protein, predicted to have 4 transmembrane segments and a coiled coil domain; regulated by Ste12p; required for efficient nuclear fusion G1 cyclin involved in regulation of the cell cycle; activates Cdc28p kinase to promote the G1 to S phase transition; late G1 specific expression depends on transcription factor complexes, MBF (Swi6p‐ Mbp1p) and SBF (Swi6p‐Swi4p) Repressor of G1 transcription that binds to SCB binding factor (SBF) at SCB target promoters in early G1; phosphorylation of Whi5p by the CDK, Cln3p/Cdc28p relieves repression and promoter binding by Whi5; periodically expressed in G1 Subunit of the conserved chromosomal passenger complex (CPC; Ipl1p‐Sli15p‐Bir1p‐Nbl1p), which regulates kinetochore‐microtubule attachments, activation of the spindle tension checkpoint, and mitotic spindle disassembly ‐1.0404 0.0033 ‐1.0365 0.0287 ‐1.0334 0.0025 ‐1.0283 0.0069 ‐1.0253 0.0009 279 log2(msn5 .Ts/WT.Ts) adj.p.v al Protein of unknown function; associates with meiosis‐specific protein Spo1p NA ‐1.0248 0.0070 ‐1.0171 0.0069 Exo‐1,3‐beta‐glucanase, involved in cell wall beta‐glucan assembly; may be anchored to the plasma membrane via a glycosylphosphatidylinositol (GPI) anchor Receptor for alpha‐factor pheromone; seven transmembrane‐ domain GPCR that interacts with both pheromone and a heterotrimeric G protein to initiate the signaling response that leads to mating between haploid a and alpha cells Protein of unknown function ‐1.0140 0.0035 ‐1.0118 0.0016 ‐1.0117 0.0014 Putative GTPase‐activating protein (GAP) involved in the Pkc1p‐ mediated signaling pathway that controls cell wall integrity; appears to specifically regulate 1,3‐beta‐glucan synthesis Evolutionarily‐conserved tail‐anchored outer mitochondrial membrane GTPase which regulates mitochondrial morphology; cells lacking Gem1p contain collapsed, globular, or grape‐like mitochondria; not required for pheromone‐induced cell death Endonuclease that processes hairpin DNA structures with the MRX complex; involved in meiotic and mitotic double‐strand break repair; phosphorylated in response to DNA damage and required for normal resistance to DNA‐damaging agents Putative protein of unknown function with similarity to acyl‐carrier‐ protein reductases; YDL114W is not an essential gene Middle sporulation‐specific mitogen‐activated protein kinase (MAPK) required for production of the outer spore wall layers; negatively regulates activity of the glucan synthase subunit Gsc2p Endosomal protein of unknown function that contains a phox (PX) homology domain and binds to both phosphatidylinositol‐3‐ phosphate (PtdIns(3)P) and proteins involved in ER‐Golgi or vesicular transport Protein of unknown function, contains an N‐terminal epsin‐like domain; proposed to be involved in the trafficking of Arn1p in the absence of ferrichrome Transporter of the ATP‐binding cassette family, involved in uptake of sterols and anaerobic growth Protein required for nuclear membrane fusion during karyogamy, localizes to the membrane with a soluble portion in the endoplasmic reticulum lumen, may form a complex with Jem1p and Kar2p; expression of the gene is regulated by pheromone Chitin transglycosylase that functions in the transfer of chitin to beta(1‐6) and beta(1‐3) glucans in the cell wall; similar to and functionally redundant with Crh1; glycosylphosphatidylinositol (GPI)‐ anchored protein localized to bud neck Membrane protein involved in zinc ion homeostasis, member of the four‐protein IZH family, expression induced by fatty acids and altered zinc levels; deletion reduces sensitivity to excess zinc; possible role in sterol metabolism Putative RNA‐binding protein required for the expression of early and middle sporulation genes Putative protein of unknown function ‐1.0074 0.0012 ‐1.0037 0.0049 ‐1.0015 0.0040 ‐0.9974 0.0080 ‐0.9917 0.0040 ‐0.9895 0.0090 ‐0.9841 0.0152 ‐0.9791 0.0055 ‐0.9719 0.0009 ‐0.9691 0.0019 ‐0.9617 0.0241 ‐0.9562 0.0109 Symbols ORF Description SPO23 YBR250W NA NA EXG2 YDR261C STE2 YFL026W PRY2 YKR013W LRG1 YDL240W GEM1 YAL048C SAE2 YGL175C NA YDL114W SMK1 YPR054W YPT35 YHR105W ENT4 YLL038C AUS1 YOR011W KAR5 YMR065W UTR2 YEL040W IZH4 YOL101C RIM4 YHL024W NA YMR147W GIC1 YHR061C DCI1 YOR180C NA YJL118W Protein of unknown function involved in initiation of budding and cellular polarization, interacts with Cdc42p via the Cdc42/Rac‐ interactive binding (CRIB) domain Peroxisomal protein; identification as a delta(3,5)‐delta(2,4)‐dienoyl‐ CoA isomerase involved in fatty acid metabolism is disputed Putative protein of unknown function; may interact with ribosomes, based on co‐purification experiments; YJL18W is a non‐essential gene; deletion enhances the toxicity of heterologously expressed 280 ‐0.9502 0.0040 ‐0.9414 0.0086 ‐0.9355 0.0108 ‐0.9331 0.0207 Symbols ORF HEM13 YDR044W YHB1 YGR234W PET122 YER153C NA YCR101C NA YGR109W‐B TPO1 YLL028W SPS1 YDR523C CLB4 YLR210W HMLALPHA 1 YCL066W GAL7 YBR018C SRL1 YOR247W MAM1 YER106W MFA2 YNL145W PDR18 YNR070W FCY22 YER060W‐A HMS1 YOR032C KAR9 YPL269W PRM8 YGL053W Description log2(msn5 .Ts/WT.Ts) adj.p.v al ‐0.9268 0.0048 ‐0.9247 0.0103 ‐0.9217 0.0102 ‐0.9206 0.0209 ‐0.9171 0.0023 ‐0.9144 0.0071 ‐0.9102 0.0439 ‐0.9096 0.0020 ‐0.9033 0.0030 ‐0.9031 0.0035 ‐0.8986 0.0012 ‐0.8973 0.0197 ‐0.8942 0.0056 ‐0.8933 0.0065 ‐0.8926 0.0215 ‐0.8865 0.0025 ‐0.8691 0.0020 ‐0.8682 0.0012 human alpha‐synuclein Coproporphyrinogen III oxidase, an oxygen requiring enzyme that catalyzes the sixth step in the heme biosynthetic pathway; transcription is repressed by oxygen and heme (via Rox1p and Hap1p) Nitric oxide oxidoreductase, flavohemoglobin involved in nitric oxide detoxification; plays a role in the oxidative and nitrosative stress responses Mitochondrial translational activator specific for the COX3 mRNA, acts together with Pet54p and Pet494p; located in the mitochondrial inner membrane Putative protein of unknown function; localizes to the membrane fraction; YCR101C is not an essential gene Retrotransposon TYA Gag and TYB Pol genes; transcribed/translated as one unit; polyprotein is processed to make a nucleocapsid‐like protein (Gag), reverse transcriptase (RT), protease (PR), and integrase (IN); similar to retroviral genes Polyamine transporter that recognizes spermine, putrescine, and spermidine; catalyzes uptake of polyamines at alkaline pH and excretion at acidic pH; phosphorylation enhances activity and sorting to the plasma membrane Putative protein serine/threonine kinase expressed at the end of meiosis and localized to the prospore membrane, required for correct localization of enzymes involved in spore wall synthesis B‐type cyclin involved in cell cycle progression; activates Cdc28p to promote the G2/M transition; may be involved in DNA replication and spindle assembly; accumulates during S phase and G2, then targeted for ubiquitin‐mediated degradation Silenced copy of ALPHA1 at HML, encoding a transcriptional coactivator involved in the regulation of mating‐type alpha‐specific gene expression Galactose‐1‐phosphate uridyl transferase, synthesizes glucose‐1‐ phosphate and UDP‐galactose from UDP‐D‐glucose and alpha‐D‐ galactose‐1‐phosphate in the second step of galactose catabolism Mannoprotein that exhibits a tight association with the cell wall, required for cell wall stability in the absence of GPI‐anchored mannoproteins; has a high serine‐threonine content; expression is induced in cell wall mutants Monopolin, kinetochore associated protein involved in chromosome attachment to meiotic spindle Mating pheromone a‐factor, made by a cells; interacts with alpha cells to induce cell cycle arrest and other responses leading to mating; biogenesis involves C‐terminal modification, N‐terminal proteolysis, and export; also encoded by MFA1 Putative transporter of the ATP‐binding cassette (ABC) family, implicated in pleiotropic drug resistance; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐ throughput studies Putative purine‐cytosine permease, very similar to Fcy2p but cannot substitute for its function Basic helix‐loop‐helix (bHLH) protein with similarity to myc‐family transcription factors; overexpression confers hyperfilamentous growth and suppresses the pseudohyphal filamentation defect of a diploid mep1 mep2 homozygous null mutant Karyogamy protein required for correct positioning of the mitotic spindle and for orienting cytoplasmic microtubules, localizes at the shmoo tip in mating cells and at the tip of the growing bud in small‐ budded cells through anaphase Pheromone‐regulated protein with 2 predicted transmembrane 281 Symbols ORF AAR2 YBL074C NRT1 YOR071C MMS21 YEL019C GAL10 YBR019C FDH1 YOR388C STE14 YDR410C PDR15 YDR406W CDA1 YLR307W NA YCR100C RAX2 YLR084C RPL13A YDL082W MSN4 YKL062W YJU2 YKL095W RTC4 YNL254C NA NA NA YDL109C ATP10 YLR393W FMP45 YDL222C NA YJR154W NIS1 YNL078W Description segments and an FF sequence, a motif involved in COPII binding; forms a complex with Prp9p in the ER; member of DUP240 gene family Component of the U5 snRNP, required for splicing of U3 precursors; originally described as a splicing factor specifically required for splicing pre‐mRNA of the MATa1 cistron High‐affinity nicotinamide riboside transporter; also transports thiamine with low affinity; shares sequence similarity with Thi7p and Thi72p; proposed to be involved in 5‐fluorocytosine sensitivity SUMO ligase involved in chromosomal organization and DNA repair; essential subunit of the Mms21‐Smc5‐Smc6 complex; mutants are sensitive to methyl methanesulfonate and show increased spontaneous mutation and mitotic recombination UDP‐glucose‐4‐epimerase, catalyzes the interconversion of UDP‐ galactose and UDP‐D‐glucose in galactose metabolism; also catalyzes the conversion of alpha‐D‐glucose or alpha‐D‐galactose to their beta‐ anomers NAD(+)‐dependent formate dehydrogenase, may protect cells from exogenous formate Farnesyl cysteine‐carboxyl methyltransferase, mediates the carboxyl methylation step during C‐terminal CAAX motif processing of a‐factor and RAS proteins in the endoplasmic reticulum, localizes to the ER membrane Plasma membrane ATP binding cassette (ABC) transporter, multidrug transporter and general stress response factor implicated in cellular detoxification; regulated by Pdr1p, Pdr3p and Pdr8p; promoter contains a PDR responsive element Chitin deacetylase, together with Cda2p involved in the biosynthesis ascospore wall component, chitosan; required for proper rigidity of the ascospore wall Putative protein of unknown function log2(msn5 .Ts/WT.Ts) adj.p.v al ‐0.8681 0.0062 ‐0.8665 0.0086 ‐0.8660 0.0133 ‐0.8647 0.0048 ‐0.8565 0.0076 ‐0.8546 0.0020 ‐0.8530 0.0025 ‐0.8521 0.0103 ‐0.8420 0.0122 N‐glycosylated protein involved in the maintenance of bud site selection during bipolar budding; localization requires Rax1p; RAX2 mRNA stability is regulated by Mpt5p Protein component of the large (60S) ribosomal subunit, nearly identical to Rpl13Bp; not essential for viability; has similarity to rat L13 ribosomal protein Transcriptional activator related to Msn2p; activated in stress conditions, which results in translocation from the cytoplasm to the nucleus; binds DNA at stress response elements of responsive genes, inducing gene expression Essential protein required for pre‐mRNA splicing; associates transiently with the spliceosomal NTC ("nineteen complex") and acts after Prp2p to promote the first catalytic reaction of splicing Protein of unknown function; null mutation suppresses cdc13‐1 temperature sensitivity; (GFP)‐fusion protein localizes to both the cytoplasm and the nucleus NA ‐0.8415 0.0048 ‐0.8407 0.0371 ‐0.8400 0.0208 ‐0.8361 0.0088 ‐0.8320 0.0028 ‐0.8311 0.0029 Putative lipase; involved in lipid metabolism; YDL109C is not an essential gene Mitochondrial inner membrane protein required for assembly of the F0 sector of mitochondrial F1F0 ATP synthase, interacts genetically with ATP6 Integral membrane protein localized to mitochondria (untagged protein); required for sporulation and maintaining sphingolipid content; has sequence similarity to SUR7 and YNL194C Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm Protein localized in the bud neck at G2/M phase; physically interacts ‐0.8304 0.0028 ‐0.8282 0.0062 ‐0.8273 0.0095 ‐0.8261 0.0282 ‐0.8216 0.0034 282 Symbols ORF TEC1 YBR083W SIM1 YIL123W SCW11 YGL028C SRB8 YCR081W MEI4 YER044C‐A IRC15 YPL017C NA YJL107C BNA4 YBL098W NA YDR042C MUC1 YIR019C NUT2 YPR168W NA YKR104W NA YGL101W IRC7 YFR055W DED1 YOR204W BBP1 YPL255W SCC2 YDR180W NDJ1 YOL104C Description log2(msn5 .Ts/WT.Ts) adj.p.v al ‐0.8197 0.0016 ‐0.8182 0.0056 ‐0.8108 0.0088 ‐0.8099 0.0025 ‐0.8094 0.0077 ‐0.8088 0.0055 ‐0.8087 0.0048 ‐0.8074 0.0017 ‐0.8053 0.0091 ‐0.8024 0.0200 ‐0.7998 0.0324 ‐0.7994 0.0187 ‐0.7988 0.0075 ‐0.7980 0.0043 ‐0.7979 0.0070 ‐0.7966 0.0028 ‐0.7923 0.0176 ‐0.7823 0.0043 with septins; possibly involved in a mitotic signaling network Transcription factor required for full Ty1 expression, Ty1‐mediated gene activation, and haploid invasive and diploid pseudohyphal growth; TEA/ATTS DNA‐binding domain family member Protein of the SUN family (Sim1p, Uth1p, Nca3p, Sun4p) that may participate in DNA replication, promoter contains SCB regulation box at ‐300 bp indicating that expression may be cell cycle‐regulated Cell wall protein with similarity to glucanases; may play a role in conjugation during mating based on its regulation by Ste12p Subunit of the RNA polymerase II mediator complex; associates with core polymerase subunits to form the RNA polymerase II holoenzyme; essential for transcriptional regulation; involved in glucose repression Meiosis‐specific protein involved in double‐strand break formation during meiotic recombination; required for chromosome synapsis and production of viable spores Microtubule associated protein; regulates microtubule dynamics; required for accurate meiotic chromosome segregation; null mutant displays large budded cells due to delayed mitotic progression, increased levels of spontaneous Rad52 foci Putative protein of unknown function; expression is induced by activation of the HOG1 mitogen‐activated signaling pathway and this induction is Hog1p/Pbs2p dependent; YJL107C and adjacent ORF, YJL108C are merged in related fungi Kynurenine 3‐mono oxygenase, required for the de novo biosynthesis of NAD from tryptophan via kynurenine; expression regulated by Hst1p; putative therapeutic target for Huntington disease Putative protein of unknown function; expression is increased in ssu72‐ts69 mutant GPI‐anchored cell surface glycoprotein (flocculin) required for pseudohyphal formation, invasive growth, flocculation, and biofilms; transcriptionally regulated by the MAPK pathway (via Ste12p and Tec1p) and the cAMP pathway (via Flo8p) Subunit of the RNA polymerase II mediator complex; associates with core polymerase subunits to form the RNA polymerase II holoenzyme; required for transcriptional activation and has a role in basal transcription Putative transporter of the multidrug resistance‐associated protein (MRP) subfamily; contains a stop codon in S288C; adjacent ORFs YKR103W and YKR104W are merged in different strain backgrounds Putative protein of unknown function; non‐essential gene with similarity to YBR242W; interacts with the DNA helicase Hpr5p Putative cystathionine beta‐lyase; involved in copper ion homeostasis and sulfur metabolism; null mutant displays increased levels of spontaneous Rad52p foci; expression induced by nitrogen limitation in a GLN3, GAT1‐dependent manner ATP‐dependent DEAD (Asp‐Glu‐Ala‐Asp)‐box RNA helicase, required for translation initiation of all yeast mRNAs; mutations in human DEAD‐box DBY are a frequent cause of male infertility Protein required for the spindle pole body (SPB) duplication, localized at the central plaque periphery; forms a complex with a nuclear envelope protein Mps2p and SPB components Spc29p and Kar1p; required for mitotic functions of Cdc5p Subunit of cohesin loading factor (Scc2p‐Scc4p), a complex required for loading of cohesin complexes onto chromosomes; involved in establishing sister chromatid cohesion during DSB repair via histone H2AX; evolutionarily‐conserved adherin Meiosis‐specific telomere protein, required for bouquet formation, 283 Symbols ORF Description effective homolog pairing, ordered cross‐over distribution, sister chromatid cohesion at meiotic telomeres, chromosomal segregation and telomere‐led rapid prophase movement Amino acid transport protein for valine, leucine, isoleucine, and tyrosine, low‐affinity tryptophan and histidine transporter; overexpression confers FK506 and FTY720 resistance Plasma membrane protein that may act together with or upstream of Stt4p to generate normal levels of the essential phospholipid PI4P, at least partially mediates proper localization of Stt4p to the plasma membrane Putative protein of unknown function, potential Cdc28p substrate; transcription is activated by paralogous transcription factors Yrm1p and Yrr1p along with genes involved in multidrug resistance Permease of basic amino acids in the vacuolar membrane log2(msn5 .Ts/WT.Ts) adj.p.v al ‐0.7816 0.0016 ‐0.7796 0.0249 ‐0.7784 0.0136 ‐0.7779 0.0070 ‐0.7771 0.0021 ‐0.7767 0.0034 ‐0.7758 0.0102 ‐0.7749 0.0056 ‐0.7748 0.0232 ‐0.7731 0.0022 TAT1 YBR069C SFK1 YKL051W NA YGR035C VBA2 YBR293W THI7 YLR237W SPC42 YKL042W HBT1 YDL223C KSS1 YGR040W PHD1 YKL043W NA YPR071W Plasma membrane transporter responsible for the uptake of thiamine, member of the major facilitator superfamily of transporters; mutation of human ortholog causes thiamine‐ responsive megaloblastic anemia Central plaque component of spindle pole body (SPB); involved in SPB duplication, may facilitate attachment of the SPB to the nuclear membrane Substrate of the Hub1p ubiquitin‐like protein that localizes to the shmoo tip (mating projection); mutants are defective for mating projection formation, thereby implicating Hbt1p in polarized cell morphogenesis Mitogen‐activated protein kinase (MAPK) involved in signal transduction pathways that control filamentous growth and pheromone response; the KSS1 gene is nonfunctional in S288C strains and functional in W303 strains Transcriptional activator that enhances pseudohyphal growth; regulates expression of FLO11, an adhesin required for pseudohyphal filament formation; similar to StuA, an A. nidulans developmental regulator; potential Cdc28p substrate Putative membrane protein; YPR071W is not an essential gene NA YNR062C Putative membrane protein of unknown function ‐0.7713 0.0039 ERD1 YDR414C ‐0.7569 0.0095 NA YLR012C ‐0.7569 0.0234 POP3 YNL282W ‐0.7522 0.0198 ATG15 YCR068W ‐0.7504 0.0072 UME6 YDR207C Predicted membrane protein required for the retention of lumenal endoplasmic reticulum proteins; mutants secrete the endogenous ER protein, BiP (Kar2p) Putative protein of unknown function; YLR012C is not an essential gene Subunit of both RNase MRP, which cleaves pre‐rRNA, and nuclear RNase P, which cleaves tRNA precursors to generate mature 5' ends Lipase required for intravacuolar lysis of autophagic bodies and Cvt bodies; targeted to intravacuolar vesicles during autophagy via the multivesicular body (MVB) pathway Key transcriptional regulator of early meiotic genes, binds URS1 upstream regulatory sequence, couples metabolic responses to nutritional cues with initiation and progression of meiosis, forms complex with Ime1p, and also with Sin3p‐Rpd3p Putative integral membrane protein of unknown function ‐0.7464 0.0070 ‐0.7434 0.0210 Transcription factor, involved in regulating multidrug resistance and oxidative stress response; forms a heterodimer with Pdr1p; contains a Zn(II)2Cys6 zinc finger domain that interacts with a pleiotropic drug resistance element in vitro Rab family GTPase that interacts with the C‐terminal tail domain of Myo2p; mediates distribution of mitochondria and endoplasmic reticuli to daughter cells Protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cell periphery and vacuole ‐0.7386 0.0028 ‐0.7380 0.0059 ‐0.7335 0.0070 NA YHR140W STB5 YHR178W YPT11 YNL304W NA YOL019W 284 Symbols ORF log2(msn5 .Ts/WT.Ts) Description adj.p.v al NA NA NA ‐0.7333 0.0158 RPL18B YNL301C ‐0.7313 0.0187 NA YOL163W ‐0.7266 0.0479 MPS2 YGL075C ‐0.7252 0.0044 GAS3 YMR215W ‐0.7239 0.0031 NA YJR124C ‐0.7204 0.0282 PDR12 YPL058C ‐0.7192 0.0051 FIG2 YCR089W ‐0.7180 0.0024 IRC4 YDR540C ‐0.7167 0.0230 HXT5 YHR096C ‐0.7130 0.0334 BUB3 YOR026W ‐0.7124 0.0165 MF(ALPHA) 1 YPL187W ‐0.7113 0.0016 MCD4 YKL165C ‐0.7080 0.0049 CIS3 YJL158C ‐0.7077 0.0016 NA YOR186W ‐0.7077 0.0154 SMP1 YBR182C ‐0.7069 0.0277 FHN1 YGR131W ‐0.7053 0.0145 HUA2 YOR284W ‐0.7041 0.0152 PUN1 YLR414C ‐0.7038 0.0073 NA YAR068W Protein component of the large (60S) ribosomal subunit, identical to Rpl18Ap and has similarity to rat L18 ribosomal protein Putative protein of unknown function; member of the Dal5p subfamily of the major facilitator family Essential membrane protein localized at the nuclear envelope and spindle pole body (SPB), required for insertion of the newly duplicated SPB into the nuclear envelope; potentially phosphorylated by Cdc28p Low abundance, possibly inactive member of the GAS family of GPI‐ containing proteins; putative 1,3‐beta‐glucanosyltransferase with similarity to other GAS family members; localizes to the cell wall; mRNA induced during sporulation Putative protein of unknown function; expression induced under calcium shortage Plasma membrane ATP‐binding cassette (ABC) transporter, weak‐ acid‐inducible multidrug transporter required for weak organic acid resistance; induced by sorbate and benzoate and regulated by War1p; mutants exhibit sorbate hypersensitivity Cell wall adhesin, expressed specifically during mating; may be involved in maintenance of cell wall integrity during mating Putative protein of unknown function; null mutant displays increased levels of spontaneous Rad52p foci; green fluorescent protein (GFP)‐ fusion protein localizes to the cytoplasm and nucleus Hexose transporter with moderate affinity for glucose, induced in the presence of non‐fermentable carbon sources, induced by a decrease in growth rate, contains an extended N‐terminal domain relative to other HXTs Kinetochore checkpoint WD40 repeat protein that localizes to kinetochores during prophase and metaphase, delays anaphase in the presence of unattached kinetochores; forms complexes with Mad1p‐Bub1p and with Cdc20p, binds Mad2p and Mad3p Mating pheromone alpha‐factor, made by alpha cells; interacts with mating type a cells to induce cell cycle arrest and other responses leading to mating; also encoded by MF(ALPHA)2, although MF(ALPHA)1 produces most alpha‐factor Protein involved in glycosylphosphatidylinositol (GPI) anchor synthesis; multimembrane‐spanning protein that localizes to the endoplasmic reticulum; highly conserved among eukaryotes Mannose‐containing glycoprotein constituent of the cell wall; member of the PIR (proteins with internal repeats) family Putative protein of unknown function; proper regulation of expression during heat stress is sphingolipid‐dependent Putative transcription factor involved in regulating the response to osmotic stress; member of the MADS‐box family of transcription factors Protein of unknown function; induced by ketoconazole; promoter region contains sterol regulatory element motif, which has been identified as a Upc2p‐binding site; overexpression complements function of Nce102p in NCE102 deletion strain Cytoplasmic protein of unknown function; computational analysis of large‐scale protein‐protein interaction data suggests a possible role in actin patch assembly Putative protein of unknown function; localizes to bud and cytoplasm; co‐localizes with Sur7p in punctate patches in the plasma membrane; null mutant displays decreased thermotolerance; transcription induced on cell wall damage Fungal‐specific protein of unknown function; induced in respiratory‐ deficient cells ‐0.7036 0.0335 285 Symbols log2(msn5 .Ts/WT.Ts) adj.p.v al ORF Description NA YNR061C Putative protein of unknown function ‐0.7011 0.0025 NA YJL147C ‐0.6997 0.0043 NA YNL034W ‐0.6963 0.0080 CAR2 YLR438W ‐0.6951 0.0180 WHI3 YNL197C ‐0.6947 0.0201 NA YLR040C ‐0.6903 0.0019 PRP39 YML046W ‐0.6869 0.0017 TOS2 YGR221C ‐0.6843 0.0027 NA YDR124W ‐0.6827 0.0241 AAD6 YFL056C ‐0.6815 0.0292 PDR11 YIL013C ‐0.6740 0.0077 NA YJL160C ‐0.6737 0.0182 TCA17 YEL048C ‐0.6699 0.0411 NA YPL152W‐A ‐0.6694 0.0048 NA YLR415C ‐0.6694 0.0051 PST1 YDR055W ‐0.6671 0.0152 NA YIL166C ‐0.6659 0.0155 SDS3 YIL084C ‐0.6653 0.0243 MCM3 YEL032W Mitochondrial protein of unknown function; homozygous diploid deletion strain has a sporulation defect characterized by elevated dityrosine in the soluble fraction; expression induced by calcium shortage; YJL147W is a non‐essential gene Putative protein of unknown function; YNL034W is not an essential gene L‐ornithine transaminase (OTAse), catalyzes the second step of arginine degradation, expression is dually‐regulated by allophanate induction and a specific arginine induction process; not nitrogen catabolite repression sensitive RNA binding protein that sequesters CLN3 mRNA in cytoplasmic foci; cytoplasmic retention factor for Cdc28p and associated cyclins; regulates cell fate and dose‐dependently regulates the critical cell size required for passage through Start Putative protein of unknown function; localizes to the cell wall; predicted to be a GPI‐attached protein; upregulated by Mcm1p‐ Alpha1p transcription factor; partially overlaps the dubious ORF YLR041W; YLR040C is not essential U1 snRNP protein involved in splicing, contains multiple tetriatricopeptide repeats Protein involved in localization of Cdc24p to the site of bud growth; may act as a membrane anchor; localizes to the bud neck and bud tip; potentially phosphorylated by Cdc28p Putative protein of unknown function; non‐essential gene; expression is strongly induced by alpha factor Putative aryl‐alcohol dehydrogenase with similarity to P. chrysosporium aryl‐alcohol dehydrogenase, involved in the oxidative stress response; expression induced in cells treated with the mycotoxin patulin ATP‐binding cassette (ABC) transporter, multidrug transporter involved in multiple drug resistance; mediates sterol uptake when sterol biosynthesis is compromisedregulated by Pdr1p; required for anaerobic growth Putative protein of unknown function; member of the PIR (proteins with internal repeats) family of cell wall proteins; non‐essential gene that is required for sporulation; mRNA is weakly cell cycle regulated, peaking in mitosis Protein that interacts with subunits of the TRAPP complex and may play a role its assembly or stability; mutation is synthetically lethal with gcs1 deletion; Sedlin_N family member; human Sedlin mutations cause the skeletal disorder SEDT Identified by gene‐trapping, microarray‐based expression analysis, and genome‐wide homology searching Putative protein of unknown function; YLR415C is not an essential gene Cell wall protein that contains a putative GPI‐attachment site; secreted by regenerating protoplasts; up‐regulated by activation of the cell integrity pathway, as mediated by Rlm1p; upregulated by cell wall damage via disruption of FKS1 Putative protein with similarity to the allantoate permease (Dal5p) subfamily of the major facilitator superfamily; mRNA expression is elevated by sulfur limitation; YIL166C is a non‐essential gene Component of the Rpd3p/Sin3p deacetylase complex required for its structural integrity and catalytic activity, involved in transcriptional silencing and required for sporulation; cells defective in SDS3 display pleiotropic phenotypes Protein involved in DNA replication; component of the Mcm2‐7 hexameric complex that binds chromatin as a part of the pre‐ ‐0.6633 0.0019 286 Symbols ORF NA YPL272C PIP2 YOR363C COG1 YGL223C RNR1 YER070W NA YMR144W YHK8 YHR048W NA YJR149W FEN2 YCR028C HST3 YOR025W SPR1 YOR190W ASE1 YOR058C SPC29 YPL124W SMA2 YML066C RNR3 YIL066C CDC5 YMR001C AXL2 YIL140W ECM25 YJL201W ECI1 YLR284C ATG29 YPL166W Description log2(msn5 .Ts/WT.Ts) adj.p.v al ‐0.6627 0.0131 ‐0.6605 0.0055 ‐0.6579 0.0056 ‐0.6579 0.0384 ‐0.6562 0.0086 ‐0.6523 0.0153 ‐0.6496 0.0117 ‐0.6495 0.0065 ‐0.6470 0.0345 ‐0.6467 0.0283 ‐0.6467 0.0144 ‐0.6440 0.0043 ‐0.6404 0.0239 ‐0.6382 0.0157 ‐0.6375 0.0104 ‐0.6372 0.0043 ‐0.6367 0.0056 ‐0.6365 0.0025 ‐0.6347 0.0208 replicative complex Putative protein of unknown function; gene expression induced in response to ketoconazole; YPL272C is not an essential gene Autoregulatory oleate‐specific transcriptional activator of peroxisome proliferation, contains Zn(2)‐Cys(6) cluster domain, forms heterodimer with Oaf1p, binds oleate response elements (OREs), activates beta‐oxidation genes Essential component of the conserved oligomeric Golgi complex (Cog1p through Cog8p), a cytosolic tethering complex that functions in protein trafficking to mediate fusion of transport vesicles to Golgi compartments Major isoform of the large subunit of ribonucleotide‐diphosphate reductase; the RNR complex catalyzes rate‐limiting step in dNTP synthesis, regulated by DNA replication and DNA damage checkpoint pathways via localization of small subunits Putative protein of unknown function; localized to the nucleus; YMR144W is not an essential gene Presumed antiporter of the DHA1 family of multidrug resistance transporters; contains 12 predicted transmembrane spans; expression of gene is up‐regulated in cells exhibiting reduced susceptibility to azoles Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm Plasma membrane H+‐pantothenate symporter; confers sensitivity to the antifungal agent fenpropimorph Member of the Sir2 family of NAD(+)‐dependent protein deacetylases; involved along with Hst4p in telomeric silencing, cell cycle progression, radiation resistance, genomic stability and short‐ chain fatty acid metabolism Sporulation‐specific exo‐1,3‐beta‐glucanase; contributes to ascospore thermoresistance Mitotic spindle midzone localized microtubule‐associated protein (MAP) family member; required for spindle elongation and stabilization; undergoes cell cycle‐regulated degradation by anaphase promoting complex; potential Cdc28p substrate Inner plaque spindle pole body (SPB) component, links the central plaque component Spc42p to the inner plaque component Spc110p; required for SPB duplication Meiosis‐specific prospore membrane protein; required to produce bending force necessary for proper assembly of the prospore membrane during sporulation Minor isoform of the large subunit of ribonucleotide‐diphosphate reductase; the RNR complex catalyzes rate‐limiting step in dNTP synthesis, regulated by DNA replication and DNA damage checkpoint pathways via localization of small subunits Polo‐like kinase with multiple functions in mitosis and cytokinesis through substrate phosphorylation, also functions in adaptation to DNA damage during meiosis; has similarity to Xenopus Plx1 and S. pombe Plo1p; possible Cdc28p substrate Integral plasma membrane protein required for axial budding in haploid cells, localizes to the incipient bud site and bud neck; glycosylated by Pmt4p; potential Cdc28p substrate Non‐essential protein of unknown function; promoter contains a consensus binding sequence for factor Abf1p Peroxisomal delta3,delta2‐enoyl‐CoA isomerase, hexameric protein that converts 3‐hexenoyl‐CoA to trans‐2‐hexenoyl‐CoA, essential for the beta‐oxidation of unsaturated fatty acids, oleate‐induced Autophagy‐specific protein that is required for recruitment of other ATG proteins to the pre‐autophagosomal structure (PAS); interacts 287 Symbols ORF PCL9 YDL179W THI72 YOR192C MSG5 YNL053W HOF1 YMR032W MSN2 YMR037C CAF120 YNL278W FUS3 YBL016W HPR1 YDR138W HCM1 YCR065W KIP1 YBL063W NA YPL067C MUM3 YOR298W MPD2 YOL088C SFL1 YOR140W RAX1 YOR301W NA YPL041C NA YGL015C Description with Atg17p and localizas to the PAS in a manner interdependent with Atg17p and Cis1p; not conserved Cyclin, forms a functional kinase complex with Pho85p cyclin‐ dependent kinase (Cdk), expressed in late M/early G1 phase, activated by Swi5p Transporter of thiamine or related compound; shares sequence similarity with Thi7p Dual‐specificity protein phosphatase; exists in 2 isoforms; required for maintenance of a low level of signaling through the cell integrity pathway, adaptive response to pheromone; regulates and is regulated by Slt2p; dephosphorylates Fus3p Bud neck‐localized, SH3 domain‐containing protein required for cytokinesis; regulates actomyosin ring dynamics and septin localization; interacts with the formins, Bni1p and Bnr1p, and with Cyk3p, Vrp1p, and Bni5p Transcriptional activator related to Msn4p; activated in stress conditions, which results in translocation from the cytoplasm to the nucleus; binds DNA at stress response elements of responsive genes, inducing gene expression Part of the evolutionarily‐conserved CCR4‐NOT transcriptional regulatory complex involved in controlling mRNA initiation, elongation, and degradation Mitogen‐activated serine/threonine protein kinase involved in mating; phosphoactivated by Ste7p; substrates include Ste12p, Far1p, Bni1p, Sst2p; inhibits invasive growth during mating by phosphorylating Tec1p, promoting its degradation Subunit of THO/TREX complexes that couple transcription elongation with mitotic recombination and with mRNA metabolism and export, subunit of an RNA Pol II complex; regulates lifespan; involved in telomere maintenance; similar to Top1p Forkhead transcription factor that drives S‐phase specific expression of genes involved in chromosome segregation, spindle dynamics, and budding; suppressor of calmodulin mutants with specific SPB assembly defects; telomere maintenance role Kinesin‐related motor protein required for mitotic spindle assembly, chromosome segregation, and 2 micron plasmid partitioning; functionally redundant with Cin8p for chromosomal but not plasmid functions Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm; YPL067C is not an essential gene Protein of unknown function involved in the organization of the outer spore wall layers; has similarity to the tafazzins superfamily of acyltransferases Member of the protein disulfide isomerase (PDI) family, exhibits chaperone activity; overexpression suppresses the lethality of a pdi1 deletion but does not complement all Pdi1p functions; undergoes oxidation by Ero1p Transcriptional repressor and activator; involved in repression of flocculation‐related genes, and activation of stress responsive genes; negatively regulated by cAMP‐dependent protein kinase A subunit Tpk2p Protein involved in bud site selection during bipolar budding; localization requires Rax2p; has similarity to members of the insulin‐ related peptide superfamily Protein of unknown function involved in maintenance of proper telomere length Putative protein of unknown function; null mutants accumulate cargo in the Golgi 288 log2(msn5 .Ts/WT.Ts) adj.p.v al ‐0.6312 0.0064 ‐0.6296 0.0345 ‐0.6290 0.0141 ‐0.6275 0.0096 ‐0.6271 0.0069 ‐0.6264 0.0362 ‐0.6248 0.0062 ‐0.6167 0.0025 ‐0.6155 0.0206 ‐0.6143 0.0208 ‐0.6141 0.0023 ‐0.6135 0.0366 ‐0.6114 0.0034 ‐0.6109 0.0182 ‐0.6097 0.0061 ‐0.6090 0.0104 ‐0.6085 0.0101 Symbols ORF Description GAS1 YMR307W log2(msn5 .Ts/WT.Ts) adj.p.v al ‐0.6079 0.0049 RGD2 YFL047W Beta‐1,3‐glucanosyltransferase, required for cell wall assembly and also has a role in transcriptional silencing; localizes to the cell surface via a glycosylphosphatidylinositol (GPI) anchor; also found at the nuclear periphery GTPase‐activating protein (RhoGAP) for Cdc42p and Rho5p ‐0.6060 0.0039 NA YOR390W Putative protein of unknown function ‐0.6034 0.0034 PRM3 YPL192C Pheromone‐regulated protein required for nuclear envelope fusion during karyogamy; localizes to the outer face of the nuclear membrane; interacts with Kar5p at the spindle pole body ‐0.6013 0.0138 289 APPENDIX O: Up-regulated genes in msn5Δ cells in amino acid starvation condition This section includes the list of transcriptionally differentially expressed genes (cutoff conditions: log2 FC 0.6 and adjusted p-value < 0.05) in msn5Δ cells in fed condition. Up-regulated genes in msn5Δ cells in amino acid starvation condition log2(msn5 .Ts/WT.Ts) adj.p.val High‐affinity inorganic phosphate (Pi) transporter and low‐affinity manganese transporter; regulated by Pho4p and Spt7p; mutation confers resistance to arsenate; exit from the ER during maturation requires Pho86p 6.0173 0.0000 YHR136C Protein with similarity to cyclin‐dependent kinase inhibitors; downregulates low‐affinity phosphate transport during phosphate limitation; overproduction suppresses a plc1 null mutation; GFP‐ fusion protein localizes to the cytoplasm 4.4287 0.0001 PHO89 YBR296C Na+/Pi cotransporter, active in early growth phase; similar to phosphate transporters of Neurospora crassa; transcription regulated by inorganic phosphate concentrations and Pho4p 4.0506 0.0013 PHM6 YDR281C Protein of unknown function, expression is regulated by phosphate levels 2.4826 0.0062 PHO11 YAR071W One of three repressible acid phosphatases, a glycoprotein that is transported to the cell surface by the secretory pathway; induced by phosphate starvation and coordinately regulated by PHO4 and PHO2 2.2526 0.0026 NA YHR214C‐E Putative protein of unknown function; identified by gene‐trapping, microarray‐based expression analysis, and genome‐wide homology searching 2.1899 0.0021 NA YGL262W Putative protein of unknown function; null mutant displays elevated sensitivity to expression of a mutant huntingtin fragment or of alpha‐ synuclein; YGL262W is not an essential gene 1.9801 0.0066 VTC3 YPL019C Subunit of the vacuolar transporter chaperone (VTC) complex involved in membrane trafficking, vacuolar polyphosphate accumulation, microautophagy and non‐autophagic vacuolar fusion 1.8455 0.0009 MTH1 YDR277C Negative regulator of the glucose‐sensing signal transduction pathway, required for repression of transcription by Rgt1p; interacts with Rgt1p and the Snf3p and Rgt2p glucose sensors; phosphorylated by Yck1p, triggering Mth1p degradation 1.6631 0.0012 Symbols ORF Description PHO84 YML123C SPL2 290 COS12 YGL263W Protein of unknown function, member of the DUP380 subfamily of conserved, often subtelomerically‐encoded proteins 1.5196 0.0095 NA YLR053C Putative protein of unknown function 1.4665 0.0069 NA YHR214C‐ D Putative protein of unknown function; identified by gene‐trapping, microarray‐based expression analysis, and genome‐wide homology searching 1.4197 0.0173 SOL4 YGR248W 6‐phosphogluconolactonase with similarity to Sol3p 1.3921 0.0147 REG2 YBR050C Regulatory subunit of the Glc7p type‐1 protein phosphatase; involved with Reg1p, Glc7p, and Snf1p in regulation of glucose‐repressible genes, also involved in glucose‐induced proteolysis of maltose permease 1.3834 0.0015 GDH3 YAL062W NADP(+)‐dependent glutamate dehydrogenase, synthesizes glutamate from ammonia and alpha‐ketoglutarate; rate of alpha‐ ketoglutarate utilization differs from Gdh1p; expression regulated by nitrogen and carbon sources 1.3764 0.0053 MOH1 YBL049W Protein of unknown function, has homology to kinase Snf7p; not required for growth on nonfermentable carbon sources; essential for survival in stationary phase 1.3625 0.0023 RGI1 YER067W Protein of unknown function involved in energy metabolism under respiratory conditions; protein abundance is increased upon intracellular iron depletion 1.3593 0.0093 SSA1 YAL005C ATPase involved in protein folding and nuclear localization signal (NLS)‐directed nuclear transport; member of heat shock protein 70 (HSP70) family; forms a chaperone complex with Ydj1p; localized to the nucleus, cytoplasm, and cell wall 1.3518 0.0026 DGR2 YKL121W Protein of unknown function; null mutant is resistant to 2‐deoxy‐D‐ glucose and displays abnormally elongated buds 1.3377 0.0015 NA YNL144C Putative protein of unknown function; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐throughput studies; YNL144C is not an essential gene 1.3179 0.0210 NA YBL055C 3'‐‐>5' exonuclease and endonuclease with a possible role in apoptosis; has similarity to mammalian and C. elegans apoptotic nucleases 1.2901 0.0016 PHO81 YGR233C Cyclin‐dependent kinase (CDK) inhibitor, regulates Pho80p‐Pho85p and Pcl7p‐Pho85p cyclin‐CDK complexes in response to phosphate levels; inhibitory activity for Pho80p‐Pho85p requires myo‐D‐inositol heptakisphosphate (IP7) generated by Vip1p 1.2787 0.0015 CYB2 YML054C Cytochrome b2 (L‐lactate cytochrome‐c oxidoreductase), component of the mitochondrial intermembrane space, required for lactate utilization; expression is repressed by glucose and anaerobic conditions 1.2371 0.0007 PHO5 YBR093C Repressible acid phosphatase (1 of 3) that also mediates extracellular nucleotide‐derived phosphate hydrolysis; secretory pathway derived cell surface glycoprotein; induced by phosphate starvation and coordinately regulated by PHO4 and PHO2 1.2277 0.0066 YPT53 YNL093W Rab family GTPase, similar to Ypt51p and Ypt52p and to mammalian rab5; required for vacuolar protein sorting and endocytosis 1.2200 0.0033 TRX3 YCR083W Mitochondrial thioredoxin, highly conserved oxidoreductase required to maintain the redox homeostasis of the cell, forms the mitochondrial thioredoxin system with Trr2p, redox state is maintained by both Trr2p and Glr1p 1.2171 0.0004 291 NA YOR289W Putative protein of unknown function; transcription induced by the unfolded protein response; green fluorescent protein (GFP)‐fusion protein localizes to both the cytoplasm and the nucleus 1.1988 0.0016 HUB1 YNR032C‐ A Ubiquitin‐like protein modifier, may function in modification of Sph1p and Hbt1p, functionally complemented by the human or S. pombe ortholog; mechanism of Hub1p adduct formation not yet clear 1.1861 0.0010 NA YKL151C Putative protein of unknown function; YKL151C promoter contains STREs (stress response elements) and expression is induced by heat shock or methyl methanesulfonate; green fluorescent protein (GFP)‐ fusion protein localizes to the cytoplasm 1.1577 0.0012 CSR2 YPR030W Nuclear protein proposed to regulate utilization of nonfermentable carbon sources and endocytosis of plasma membrane proteins; overproduction suppresses chs5 spa2 lethality at high temp; ubiquitinated by Rsp5p, deubiquitinated by Ubp2p 1.1536 0.0022 HXT4 YHR092C High‐affinity glucose transporter of the major facilitator superfamily, expression is induced by low levels of glucose and repressed by high levels of glucose 1.1515 0.0259 NA YLR312C Putative protein of unknown function 1.1370 0.0065 DCS2 YOR173W Non‐essential, stress induced regulatory protein; modulates m7G‐ oligoribonucleotide metabolism; inhibits Dcs1p; regulated by Msn2p, Msn4p, and the Ras‐cAMP‐cAPK signaling pathway; mutant has increased aneuploidy tolerance 1.1227 0.0034 NA NA NA 1.1215 0.0032 ISF1 YMR081C Serine‐rich, hydrophilic protein with similarity to Mbr1p; overexpression suppresses growth defects of hap2, hap3, and hap4 mutants; expression is under glucose control; cotranscribed with NAM7 in a cyp1 mutant 1.1158 0.0131 NA Putative protein of unknown function; predicted to have a role in cell budding based on computational "guilt by association" analysis Outer kinetochore protein, required for accurate mitotic chromosome segregation; component of the kinetochore sub‐ complex COMA (Ctf19p, Okp1p, Mcm21p, Ame1p) that functions as a platform for kinetochore assembly 1.0900 0.0101 CTF19 YGR174W‐ A YPL018W 1.0857 0.0195 RAD28 YDR030C Protein involved in DNA repair, related to the human CSA protein that is involved in transcription‐coupled repair nucleotide excision repair 1.0854 0.0009 NA YNR034W‐ A Putative protein of unknown function; expression is regulated by Msn2p/Msn4p 1.0818 0.0065 TMA23 YMR269W Nucleolar protein of unknown function implicated in ribosome biogenesis; TMA23 may be a fungal‐specific gene as no homologs have been yet identified in higher eukaryotes 1.0680 0.0154 VTC4 YJL012C Vacuolar membrane polyphosphate polymerase; subunit of the vacuolar transporter chaperone (VTC) complex involved in synthesis and transfer of polyP to the vacuole; regulates membrane trafficking; role in non‐autophagic vacuolar fusion 1.0654 0.0016 HXK1 YFR053C Hexokinase isoenzyme 1, a cytosolic protein that catalyzes phosphorylation of glucose during glucose metabolism; expression is highest during growth on non‐glucose carbon sources; glucose‐ induced repression involves the hexokinase Hxk2p 1.0552 0.0076 RTC3 YHR087W Protein of unknown function involved in RNA metabolism; has structural similarity to SBDS, the human protein mutated in Shwachman‐Diamond Syndrome (the yeast SBDS ortholog = SDO1); null mutation suppresses cdc13‐1 temperature sensitivity 1.0531 0.0485 292 NA YDR018C Probable membrane protein with three predicted transmembrane domains; homologous to Ybr042cp, similar to C. elegans F55A11.5 and maize 1‐acyl‐glycerol‐3‐phosphate acyltransferase 1.0382 0.0018 BCD1 YHR040W Essential protein required for the accumulation of box C/D snoRNA 1.0303 0.0152 VTC1 YER072W Subunit of the vacuolar transporter chaperone (VTC) complex involved in membrane trafficking, vacuolar polyphosphate accumulation, microautophagy and non‐autophagic vacuolar fusion; also has mRNA binding activity 1.0297 0.0012 NA YBL086C Protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cell periphery 1.0109 0.0018 CYC7 YEL039C Cytochrome c isoform 2, expressed under hypoxic conditions; electron carrier of the mitochondrial intermembrane space that transfers electrons from ubiquinone‐cytochrome c oxidoreductase to cytochrome c oxidase during cellular respiration 1.0009 0.0260 FUN19 YAL034C Non‐essential protein of unknown function; expression induced in response to heat stress 0.9980 0.0007 TSA2 YDR453C Stress inducible cytoplasmic thioredoxin peroxidase; cooperates with Tsa1p in the removal of reactive oxygen, nitrogen and sulfur species using thioredoxin as hydrogen donor; deletion enhances the mutator phenotype of tsa1 mutants 0.9953 0.0104 NA YGR102C Subunit of the trimeric GatFAB AmidoTransferase(AdT) complex; involved in the formation of Q‐tRNAQ; transposon insertion mutant is salt sensitive and null mutant has growth defects; non‐tagged protein is detected in purified mitochondria 0.9890 0.0025 CHA1 YCL064C Catabolic L‐serine (L‐threonine) deaminase, catalyzes the degradation of both L‐serine and L‐threonine; required to use serine or threonine as the sole nitrogen source, transcriptionally induced by serine and threonine 0.9776 0.0192 UGX2 YDL169C Protein of unknown function, transcript accumulates in response to any combination of stress conditions 0.9618 0.0071 PUT1 YLR142W Proline oxidase, nuclear‐encoded mitochondrial protein involved in utilization of proline as sole nitrogen source; PUT1 transcription is induced by Put3p in the presence of proline and the absence of a preferred nitrogen source 0.9616 0.0016 USV1 YPL230W Putative transcription factor containing a C2H2 zinc finger; mutation affects transcriptional regulation of genes involved in growth on non‐ fermentable carbon sources, response to salt stress and cell wall biosynthesis 0.9583 0.0083 HSP78 YDR258C Oligomeric mitochondrial matrix chaperone that cooperates with Ssc1p in mitochondrial thermotolerance after heat shock; able to prevent the aggregation of misfolded proteins as well as resolubilize protein aggregates 0.9579 0.0012 COX17 YLL009C Copper metallochaperone that transfers copper to Sco1p and Cox11p for eventual delivery to cytochrome c oxidase; contains twin cysteine‐ x9‐cysteine motifs 0.9496 0.0028 STF2 YGR008C Protein involved in regulation of the mitochondrial F1F0‐ATP synthase; Stf1p and Stf2p may act as stabilizing factors that enhance inhibitory action of the Inh1p protein 0.9492 0.0189 FMP46 YKR049C Putative redox protein containing a thioredoxin fold; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐throughput studies 0.9400 0.0016 293 PHO87 YCR037C Low‐affinity inorganic phosphate (Pi) transporter, involved in activation of PHO pathway; expression is independent of Pi concentration and Pho4p activity; contains 12 membrane‐spanning segments 0.9287 0.0025 HSP104 YLL026W Heat shock protein that cooperates with Ydj1p (Hsp40) and Ssa1p (Hsp70) to refold and reactivate previously denatured, aggregated proteins; responsive to stresses including: heat, ethanol, and sodium arsenite; involved in [PSI+] propagation 0.9260 0.0042 YPS6 YIR039C 0.9259 0.0026 NA YCL073C Putative GPI‐anchored aspartic protease, member of the yapsin family of proteases involved in cell wall growth and maintenance Protein of unconfirmed function; displays a topology characteristic of the Major Facilitators Superfamily of membrane proteins; coding sequence 98% identical to that of YKR106W 0.9120 0.0039 NA YGR226C Dubious open reading frame, unlikely to encode a protein; not conserved in closely related Saccharomyces species; overlaps significantly with a verified ORF, AMA1/YGR225W 0.9005 0.0025 NA YER137C Putative protein of unknown function 0.9003 0.0207 NA YMR090W Putative protein of unknown function with similarity to DTDP‐glucose 4,6‐dehydratases; GFP‐fusion protein localizes to the cytoplasm; up‐ regulated in response to the fungicide mancozeb; not essential for viability 0.8945 0.0055 NA YFR017C Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm and is induced in response to the DNA‐damaging agent MMS; YFR017C is not an essential gene 0.8890 0.0020 MAL31 YBR298C Maltose permease, high‐affinity maltose transporter (alpha‐glucoside transporter); encoded in the MAL3 complex locus; member of the 12 transmembrane domain superfamily of sugar transporters; functional in genomic reference strain S288C 0.8863 0.0024 UBC12 YLR306W Enzyme that mediates the conjugation of Rub1p, a ubiquitin‐like protein, to other proteins; related to E2 ubiquitin‐conjugating enzymes 0.8802 0.0020 MRPS16 YPL013C Mitochondrial ribosomal protein of the small subunit 0.8708 0.0044 AMS1 YGL156W Vacuolar alpha mannosidase, involved in free oligosaccharide (fOS) degradation; delivered to the vacuole in a novel pathway separate from the secretory pathway 0.8658 0.0131 NA YJL077W‐B Putative protein of unknown function; identified by gene‐trapping, microarray‐based expression analysis, and genome‐wide homology searching 0.8633 0.0102 MIC14 YDR031W Mitochondrial intermembrane space protein, required for normal oxygen consumption; contains twin cysteine‐x9‐cysteine motifs 0.8615 0.0030 ALG14 YBR070C Component of UDP‐GlcNAc transferase required for the second step of dolichyl‐linked oligosaccharide synthesis; anchors the catalytic subunit Alg13p to the ER membrane; similar to bacterial and human glycosyltransferases 0.8593 0.0016 GIS3 YLR094C Protein of unknown function 0.8541 0.0272 NA YMR105W ‐A YKL221W Putative protein of unknown function 0.8520 0.0149 Protein with similarity to mammalian monocarboxylate permeases, which are involved in transport of monocarboxylic acids across the plasma membrane; mutant is not deficient in monocarboxylate transport 0.8483 0.0075 MCH2 294 SYF2 YGR129W Member of the NineTeen Complex (NTC) that contains Prp19p and stabilizes U6 snRNA in catalytic forms of the spliceosome containing U2, U5, and U6 snRNAs; isy1 syf2 cells have defective spindles activiating cell cycle arrest 0.8396 0.0384 DAL1 YIR027C Allantoinase, converts allantoin to allantoate in the first step of allantoin degradation; expression sensitive to nitrogen catabolite repression 0.8333 0.0054 OM45 YIL136W Protein of unknown function, major constituent of the mitochondrial outer membrane; located on the outer (cytosolic) face of the outer membrane 0.8269 0.0260 GAC1 YOR178C Regulatory subunit for Glc7p type‐1 protein phosphatase (PP1), tethers Glc7p to Gsy2p glycogen synthase, binds Hsf1p heat shock transcription factor, required for induction of some HSF‐regulated genes under heat shock 0.8158 0.0493 COA2 YPL189C‐A Cytochrome oxidase assembly factor; null mutation results in respiratory deficiency with specific loss of cytochrome oxidase activity; functions downstream of assembly factors Mss51p and Coa1p and interacts with assembly factor Shy1p 0.8143 0.0051 SFC1 YJR095W Mitochondrial succinate‐fumarate transporter, transports succinate into and fumarate out of the mitochondrion; required for ethanol and acetate utilization 0.8136 0.0123 VTC2 YFL004W Subunit of the vacuolar transporter chaperone (VTC) complex involved in membrane trafficking, vacuolar polyphosphate accumulation, microautophagy and non‐autophagic vacuolar fusion 0.8104 0.0109 MTC3 YGL226W Protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the mitochondrion; mtc3 is synthetically sick with cdc13‐1 0.8094 0.0061 DAD4 YDR320C‐ A Essential subunit of the Dam1 complex (aka DASH complex), couples kinetochores to the force produced by MT depolymerization thereby aiding in chromosome segregation; is transferred to the kinetochore prior to mitosis 0.8058 0.0020 NA YPR159C‐A Identified by gene‐trapping, microarray‐based expression analysis, and genome‐wide homology searching 0.8050 0.0075 UBP9 YER098W Ubiquitin carboxyl‐terminal hydrolase, ubiquitin‐specific protease that cleaves ubiquitin‐protein fusions 0.8030 0.0040 HSP42 YDR171W Small heat shock protein (sHSP) with chaperone activity; forms barrel‐ shaped oligomers that suppress unfolded protein aggregation; involved in cytoskeleton reorganization after heat shock 0.7986 0.0235 CBP4 YGR174C Mitochondrial protein required for assembly of ubiquinol cytochrome‐c reductase complex (cytochrome bc1 complex); interacts with Cbp3p and function is partially redundant with that of Cbp3p 0.7978 0.0019 DIA3 YDL024C Protein of unknown function, involved in invasive and pseudohyphal growth 0.7901 0.0019 BUD27 YFL023W Unconventional prefoldin protein involved in translation initiation; mutants have inappropriate expression of nutrient sensitive genes due to translational derepression of Gcn4p transcription factor; diploid mutants show random budding 0.7900 0.0313 FIS1 YIL065C Protein involved in mitochondrial membrane fission and peroxisome abundance; required for localization of Dnm1p and Mdv1p during mitochondrial division; mediates ethanol‐induced apoptosis and ethanol‐induced mitochondrial fragmentation 0.7878 0.0025 YAE1 YJR067C Protein of unknown function, essential for growth under standard (aerobic) conditions but not under anaerobic conditions 0.7848 0.0035 295 NA YNL200C Putative protein of unknown function; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐throughput studies Vacuolar protein sorting (VPS) protein required for cytoplasm to vacuole targeting of proteins 0.7817 0.0022 VPS62 YGR141W 0.7755 0.0040 YPC1 YBR183W Alkaline ceramidase that also has reverse (CoA‐independent) ceramide synthase activity, catalyzes both breakdown and synthesis of phytoceramide; overexpression confers fumonisin B1 resistance 0.7734 0.0250 SRX1 YKL086W Sulfiredoxin, contributes to oxidative stress resistance by reducing cysteine‐sulfinic acid groups in the peroxiredoxin Tsa1p, which is formed upon exposure to oxidants; conserved in higher eukaryotes 0.7716 0.0061 NA YAR064W Putative protein of unknown function 0.7701 0.0234 GPX1 YKL026C 0.7660 0.0030 GDB1 YPR184W Phospholipid hydroperoxide glutathione peroxidase induced by glucose starvation that protects cells from phospholipid hydroperoxides and nonphospholipid peroxides during oxidative stress Glycogen debranching enzyme containing glucanotranferase and alpha‐1,6‐amyloglucosidase activities, required for glycogen degradation; phosphorylated in mitochondria 0.7653 0.0174 NA YPL247C Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm and nucleus; similar to the petunia WD repeat protein an11; overexpression causes a cell cycle delay or arrest 0.7629 0.0080 HSP82 YPL240C Hsp90 chaperone required for pheromone signaling and negative regulation of Hsf1p; docks with Tom70p for mitochondrial preprotein delivery; promotes telomerase DNA binding and nucleotide addition; interacts with Cns1p, Cpr6p, Cpr7p, Sti1p 0.7489 0.0109 PGM3 YMR278W Phosphoglucomutase, catalyzes interconversion of glucose‐1‐ phosphate and glucose‐6‐phospate; transcription induced in response to stress; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm and nucleus; non‐essential 0.7485 0.0249 RRP17 YDR412W Component of the pre‐60S pre‐ribosomal particle; required for cell viability under standard (aerobic) conditions but not under anaerobic conditions; exonuclease required for 5′ end processing of pre‐ 60S ribosomal RNA 0.7468 0.0112 IES2 YNL215W Protein that associates with the INO80 chromatin remodeling complex under low‐salt conditions; essential for growth under anaerobic conditions 0.7451 0.0048 RGM1 YMR182C Putative transcriptional repressor with proline‐rich zinc fingers; overproduction impairs cell growth 0.7449 0.0444 GIT1 YCR098C Plasma membrane permease, mediates uptake of glycerophosphoinositol and glycerophosphocholine as sources of the nutrients inositol and phosphate; expression and transport rate are regulated by phosphate and inositol availability 0.7445 0.0029 NA YIL014C‐A Putative protein of unknown function 0.7402 0.0138 BUD20 YLR074C Protein involved in bud‐site selection; diploid mutants display a random budding pattern instead of the wild‐type bipolar pattern 0.7397 0.0466 NA YBR230W‐ A YPL186C Putative protein of unknown function 0.7382 0.0133 Protein that interacts with Ulp1p, a Ubl (ubiquitin‐like protein)‐ specific protease for Smt3p protein conjugates; detected in a phosphorylated state in the mitochondrial outer membrane; also detected in ER and nuclear envelope 0.7308 0.0027 UIP4 296 DAL3 YIR032C Ureidoglycolate hydrolase, converts ureidoglycolate to glyoxylate and urea in the third step of allantoin degradation; expression sensitive to nitrogen catabolite repression 0.7308 0.0362 NA YMR244C‐ A Putative protein of unknown function; green fluorescent protein (GFP)‐fusion protein localizes to both the cytoplasm and nucleus and is induced in response to the DNA‐damaging agent MMS; YMR244C‐A is not an essential gene 0.7252 0.0061 MRPL51 YPR100W Mitochondrial ribosomal protein of the large subunit 0.7213 0.0048 RIO1 YOR119C Essential serine kinase involved in cell cycle progression and processing of the 20S pre‐rRNA into mature 18S rRNA 0.7196 0.0069 HIT1 YJR055W Protein of unknown function, required for growth at high temperature 0.7192 0.0131 TRM13 YOL125W 2'‐O‐methyltransferase responsible for modification of tRNA at position 4; C‐terminal domain has similarity to Rossmann‐fold (RFM) superfamily of RNA methyltransferases 0.7142 0.0388 NA YOR032W‐ A Identified by gene‐trapping, microarray‐based expression analysis, and genome‐wide homology searching 0.7141 0.0449 NA YLR156W Putative protein of unknown function; exhibits a two‐hybrid interaction with Jsn1p in a large‐scale analysis 0.7139 0.0112 NA YOL114C Putative protein of unknown function with similarity to human ICT1 and prokaryotic factors that may function in translation termination; YOL114C is not an essential gene 0.7136 0.0225 NA YFL054C Putative channel‐like protein; similar to Fps1p; mediates passive diffusion of glycerol in the presence of ethanol 0.7120 0.0030 AUA1 YFL010W‐ A Protein required for the negative regulation by ammonia of Gap1p, which is a general amino acid permease 0.7116 0.0020 KIN82 YCR091W Putative serine/threonine protein kinase implicated in the regulation of phospholipid asymmetry through the activation of phospholipid translocases (flippases) Lem3p‐Dnf1p/Dnf2p; similar to Fpk1p 0.7107 0.0268 CUS2 YNL286W Protein that binds to U2 snRNA and Prp11p, may be involved in U2 snRNA folding; contains two RNA recognition motifs (RRMs) 0.7092 0.0019 URM1 YIL008W Ubiquitin‐like protein involved in thiolation of cytoplasmic tRNAs; receives sulfur from the E1‐like enzyme Uba4p and transfers it to tRNA; also functions as a protein tag with roles in nutrient sensing and oxidative stress response 0.7054 0.0234 MBR1 YKL093W Protein involved in mitochondrial functions and stress response; overexpression suppresses growth defects of hap2, hap3, and hap4 mutants 0.7043 0.0056 GLC3 YEL011W Glycogen branching enzyme, involved in glycogen accumulation; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm in a punctate pattern 0.7022 0.0085 NA Putative protein of unknown function 0.6998 0.0071 PBI2 YGR169C‐ A YNL015W Cytosolic inhibitor of vacuolar proteinase B (PRB1), required for efficient vacuole inheritance; with thioredoxin forms protein complex LMA1, which assists in priming SNARE molecules and promotes vacuole fusion 0.6981 0.0215 PYK2 YOR347C Pyruvate kinase that appears to be modulated by phosphorylation; PYK2 transcription is repressed by glucose, and Pyk2p may be active under low glycolytic flux 0.6923 0.0131 EMI1 YDR512C Non‐essential protein required for transcriptional induction of the early meiotic‐specific transcription factor IME1, also required for sporulation; contains twin cysteine‐x9‐cysteine motifs 0.6824 0.0025 297 NGL3 YML118W Putative endonuclease, has a domain similar to a magnesium‐ dependent endonuclease motif in mRNA deadenylase Ccr4p; similar to Ngl1p and Ngl2p 0.6799 0.0235 NA YHR138C Putative protein of unknown function; has similarity to Pbi2p; double null mutant lacking Pbi2p and Yhr138p exhibits highly fragmented vacuoles 0.6765 0.0063 GCY1 YOR120W Putative NADP(+) coupled glycerol dehydrogenase, proposed to be involved in an alternative pathway for glycerol catabolism; also has mRNA binding activity; member of the aldo‐keto reductase (AKR) family 0.6754 0.0056 COX13 YGL191W Subunit VIa of cytochrome c oxidase, which is the terminal member of the mitochondrial inner membrane electron transport chain; not essential for cytochrome c oxidase activity but may modulate activity in response to ATP 0.6743 0.0040 ERR3 YMR323W Protein of unknown function, has similarity to enolases 0.6742 0.0349 NA YAR029W Member of DUP240 gene family but contains no transmembrane domains; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm in a punctate pattern 0.6719 0.0230 PRX1 YBL064C Mitochondrial peroxiredoxin (1‐Cys Prx) with thioredoxin peroxidase activity, has a role in reduction of hydroperoxides; reactivation requires Trr2p and glutathione; induced during respiratory growth and oxidative stress; phosphorylated 0.6701 0.0049 COS4 YFL062W Protein of unknown function, member of the DUP380 subfamily of conserved, often subtelomerically‐encoded proteins 0.6699 0.0131 HBN1 YCL026C‐B Putative protein of unknown function; similar to bacterial nitroreductases; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm and nucleus; protein becomes insoluble upon intracellular iron depletion 0.6678 0.0221 NA YFL041W‐ A Putative protein of unknown function; identified by fungal homology and RT‐PCR 0.6676 0.0050 CRS5 YOR031W Copper‐binding metallothionein, required for wild‐type copper resistance 0.6672 0.0120 CAT8 YMR280C Zinc cluster transcriptional activator necessary for derepression of a variety of genes under non‐fermentative growth conditions, active after diauxic shift, binds carbon source responsive elements 0.6671 0.0092 NA YKL107W Putative protein of unknown function; proposed to be a palmitoylated membrane protein 0.6645 0.0319 CAR1 YPL111W Arginase, responsible for arginine degradation, expression responds to both induction by arginine and nitrogen catabolite repression; disruption enhances freeze tolerance 0.6645 0.0109 NA YKL018C‐A Putative protein of unknown function; identified by homology; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm 0.6631 0.0059 MND2 YIR025W Subunit of the anaphase‐promoting complex (APC); necessary for maintaining sister chromatid cohesion in prophase I of meiosis by inhibiting premature ubiquitination and subsequent degradation of substrates by the APC(Ama1) ubiquitin ligase 0.6627 0.0039 FMP10 YER182W Putative protein of unknown function; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐throughput studies 0.6600 0.0078 DAN4 YJR151C Cell wall mannoprotein with similarity to Tir1p, Tir2p, Tir3p, and Tir4p; expressed under anaerobic conditions, completely repressed during aerobic growth 0.6591 0.0054 298 CGR1 YGL029W Protein involved in nucleolar integrity and processing of the pre‐rRNA for the 60S ribosome subunit; transcript is induced in response to cytotoxic stress but not genotoxic stress 0.6578 0.0430 ATS1 YAL020C Protein required, with Elongator complex, Kti11p, and Kti12p, for modification of wobble nucleosides in tRNA; has a potential role in regulatory interactions between microtubules and the cell cycle 0.6567 0.0078 NA YDL160C‐A Putative protein of unknown function; mutant in a srs2 mutant background displays MMS hypersensitivity; ortholog of human MHF2, a component of the Fanconi anemia (FA) complex that is involved in maintaining genome stability 0.6555 0.0208 SNF3 YDL194W Plasma membrane low glucose sensor that regulates glucose transport; contains 12 predicted transmembrane segments and a long C‐terminal tail required for induction of hexose transporters; also senses fructose and mannose; similar to Rgt2p 0.6539 0.0074 REX3 YLR107W RNA exonuclease; required for maturation of the RNA component of RNase MRP; functions redundantly with Rnh70p and Rex2p in processing of U5 snRNA and RNase P RNA; member of RNase D family of exonucleases 0.6536 0.0069 NA YOL164W‐ A Putative protein of unknown function; identified by fungal homology and RT‐PCR 0.6530 0.0359 COS2 YBR302C Protein of unknown function, member of the DUP380 subfamily of conserved, often subtelomerically‐encoded proteins 0.6524 0.0073 TOA1 YOR194C TFIIA large subunit; involved in transcriptional activation, acts as antirepressor or as coactivator; homologous to largest and second largest subunits of human and Drosophila TFIIA 0.6522 0.0227 ASR1 YPR093C Ubiquitin ligase that modifies and regulates RNA Pol II; involved in a putative alcohol‐responsive signaling pathway; accumulates in the nucleus under alcohol stress; contains a Ring/PHD finger domain similar to the mammalian rA9 protein 0.6490 0.0094 GRX1 YCL035C Hydroperoxide and superoxide‐radical responsive heat‐stable glutathione‐dependent disulfide oxidoreductase with active site cysteine pair; protects cells from oxidative damage 0.6475 0.0115 CRP1 YHR146W Protein that binds to cruciform DNA structures 0.6469 0.0028 CPD1 YGR247W Cyclic nucleotide phosphodiesterase, hydrolyzes ADP‐ribose 1'', 2''‐ cyclic phosphate to ADP‐ribose 1''‐phosphate; may have a role in tRNA splicing; no detectable phenotype is conferred by null mutation or by overexpression 0.6466 0.0032 SSA2 YLL024C ATP binding protein involved in protein folding and vacuolar import of proteins; member of heat shock protein 70 (HSP70) family; associated with the chaperonin‐containing T‐complex; present in the cytoplasm, vacuolar membrane and cell wall 0.6461 0.0107 TOP3 YLR234W DNA Topoisomerase III, conserved protein that functions in a complex with Sgs1p and Rmi1p to relax single‐stranded negatively‐supercoiled DNA preferentially, involved in telomere stability and regulation of mitotic recombination 0.6445 0.0146 CUE4 YML101C Protein of unknown function; has a CUE domain that binds ubiquitin, which may facilitate intramolecular monoubiquitination 0.6434 0.0031 MOG1 YJR074W Conserved nuclear protein that interacts with GTP‐Gsp1p, which is a Ran homolog of the Ras GTPase family, and stimulates nucleotide release, involved in nuclear protein import, nucleotide release is inhibited by Yrb1p 0.6417 0.0048 299 TPC1 YGR096W Mitochondrial membrane transporter that mediates uptake of the essential cofactor thiamine pyrophosphate (ThPP) into mitochondria; expression appears to be regulated by carbon source; member of the mitochondrial carrier family 0.6399 0.0075 STI1 YOR027W Hsp90 cochaperone, interacts with the Ssa group of the cytosolic Hsp70 chaperones and activates Ssa1p ATPase activity; interacts with Hsp90 chaperones and inhibits their ATPase activity; homolog of mammalian Hop 0.6373 0.0052 NA YIR018C‐A Putative protein of unknown function; identified by expression profiling and mass spectrometry 0.6368 0.0481 NA Putative protein of unknown function 0.6356 0.0109 VIP1 YMR182W ‐A YLR410W Inositol hexakisphosphate (IP6) and inositol heptakisphosphate (IP7) kinase; IP7 production is important for phosphate signaling; involved in cortical actin cytoskeleton function, and invasive pseudohyphal growth analogous to S. pombe asp1 0.6332 0.0059 NQM1 YGR043C 0.6321 0.0167 BRE2 YLR015W Transaldolase of unknown function; transcription is repressed by Mot1p and induced by alpha‐factor and during diauxic shift Subunit of COMPASS (Set1C) complex, which methylates Lys4 of histone H3 and functions in silencing at telomeres; has a C‐terminal Sdc1 Dpy‐30 Interaction (SDI) domain that mediates binding to Sdc1p; similar to trithorax‐group protein ASH2L 0.6310 0.0059 MOD5 YOR274W Delta 2‐isopentenyl pyrophosphate:tRNA isopentenyl transferase, required for biosynthesis of the modified base isopentenyladenosine in mitochondrial and cytoplasmic tRNAs; gene is nuclear and encodes two isozymic forms 0.6248 0.0356 MER1 YNL210W Protein with RNA‐binding motifs required for meiosis‐specific mRNA splicing; required for chromosome pairing and meiotic recombination 0.6239 0.0354 ECM4 YKR076W Omega class glutathione transferase; not essential; similar to Ygr154cp; green fluorescent protein (GFP)‐fusion protein localizes to the cytoplasm 0.6174 0.0127 SDH4 YDR178W Membrane anchor subunit of succinate dehydrogenase (Sdh1p, Sdh2p, Sdh3p, Sdh4p), which couples the oxidation of succinate to the transfer of electrons to ubiquinone as part of the TCA cycle and the mitochondrial respiratory chain 0.6155 0.0065 NA YDR391C Putative protein of unknown function, possibly involved in zinc homeostasis; Bdf1p‐dependent transcription induced by salt stress; green fluorescent protein (GFP)‐fusion protein localizes to both the cytoplasm and the nucleus 0.6152 0.0081 TEN1 YLR010C Protein that regulates telomeric length; protects telomeric ends in a complex with Cdc13p and Stn1p 0.6147 0.0051 EMP46 YLR080W 0.6143 0.0323 NA YPL039W Integral membrane component of endoplasmic reticulum‐derived COPII‐coated vesicles, which function in ER to Golgi transport Putative protein of unknown function; YPL039W is not an essential gene 0.6138 0.0062 NCE101 YJL205C Protein of unknown function, involved in secretion of proteins that lack classical secretory signal sequences 0.6120 0.0030 NVJ1 YHR195W Nuclear envelope protein, anchored to the nuclear inner membrane, that interacts with the vacuolar membrane protein Vac8p to promote formation of nucleus‐vacuole junctions during piecemeal microautophagy of the nucleus (PMN) 0.6113 0.0120 RAD2 YGR258C Single‐stranded DNA endonuclease, cleaves single‐stranded DNA during nucleotide excision repair to excise damaged DNA; subunit of Nucleotide Excision Repair Factor 3 (NEF3); homolog of human XPG protein 0.6112 0.0065 300 QCR8 YJL166W Subunit 8 of ubiquinol cytochrome‐c reductase complex, which is a component of the mitochondrial inner membrane electron transport chain; oriented facing the intermembrane space; expression is regulated by Abf1p and Cpf1p 0.6105 0.0055 LAP4 YKL103C Vacuolar aminopeptidase yscI; zinc metalloproteinase that belongs to the peptidase family M18; often used as a marker protein in studies of autophagy and cytosol to vacuole targeting (CVT) pathway 0.6101 0.0097 SEM1 YDR363W‐ A Component of the lid subcomplex of the regulatory subunit of the 26S proteasome; involved in mRNA export mediated by the TREX‐2 complex (Sac3p‐Thp1p); ortholog of human DSS1 0.6080 0.0187 FMP33 YJL161W Putative protein of unknown function; the authentic, non‐tagged protein is detected in highly purified mitochondria in high‐throughput studies 0.6077 0.0144 MXR2 YCL033C Methionine‐R‐sulfoxide reductase, involved in the response to oxidative stress; protects iron‐sulfur clusters from oxidative inactivation along with MXR1; involved in the regulation of lifespan 0.6049 0.0025 PRP2 YNR011C RNA‐dependent ATPase in the DEAH‐box family, required for activation of the spliceosome before the first transesterification step in RNA splicing; orthologous to human protein DHX16 0.6044 0.0062 NA YPR015C Putative protein of unknown function; overexpression causes a cell cycle delay or arrest 0.6044 0.0100 YCT1 YLL055W High‐affinity cysteine‐specific transporter with similarity to the Dal5p family of transporters; green fluorescent protein (GFP)‐fusion protein localizes to the endoplasmic reticulum; YCT1 is not an essential gene 0.6023 0.0209 NHA1 YLR138W Na+/H+ antiporter involved in sodium and potassium efflux through the plasma membrane; required for alkali cation tolerance at acidic pH 0.6001 0.0073 301 APPENDIX P: R codes for microarray data analysis This section includes the R codes for microarray data analysis to generate the differential expressed genes in this project. ## download packages required for microarray analysis >source("http://bioconductor.org/biocLite.R") >biocLite("affyPLM") ## check working environment >sessionInfo() R version 2.13.0 (2011-04-13) Platform: i386-pc-mingw32/i386 (32-bit) locale: [1] LC_COLLATE=English_United States.1252 [2] LC_CTYPE=English_United States.1252 [3] LC_MONETARY=English_United States.1252 [4] LC_NUMERIC=C [5] LC_TIME=English_United States.1252 attached base packages: [1] stats graphics grDevices utils other attached packages: [1] affycoretools_1.24.0 [4] yeast2.db_2.5.0 [7] DBI_0.2-5 [10] yeast2probe_2.8.0 [13] limma_3.8.2 datasets KEGG.db_2.5.0 org.Sc.sgd.db_2.5.0 annotate_1.30.0 yeast2cdf_2.8.0 affy_1.30.0 base GO.db_2.5.0 RSQLite_0.9-4 genefilter_1.34.0 AnnotationDbi_1.14.1 Biobase_2.12.1 loaded via a namespace (and not attached): [1] affyio_1.20.0 annaffy_1.24.0 [4] Biostrings_2.20.4 Category_2.18.0 [7] GOstats_2.18.0 graph_1.30.0 [10] IRanges_1.10.6 preprocessCore_1.14.0 [13] RCurl_1.6-10.1 splines_2.13.0 [16] tools_2.13.0 XML_3.4-2.2 302 methods biomaRt_2.8.1 gcrma_2.24.1 GSEABase_1.14.0 RBGL_1.28.0 survival_2.36-5 xtable_1.5-6 ########### ########### ########### importing cel files follow online manual -- limma ########### ########### ########### setwd("F://2007- Data in Hopper Lab//Microarray data analysis//R wd//wt_mtr10_msn5") library("affy") library("limma") targets <- readTargets("targets_all.txt") aaa <- ReadAffy(filenames = targets $ filename) pData(aaa) setwd("F://2007- Data in Hopper Lab//Microarray data analysis//R_Output//WT_mtr10_msn5_output") pdf("Box plot-Raw data.pdf", width=5, height=5) par(mar=c(10,4,2,1)) boxplot(aaa, las=3, cex.axis=0.7, ylab="log2 intensity", main="Raw data");dev.off() ########### ########### ########### ######### ######### ######### Remove S.p. probes ##### from BioC list--Jenny Jenny Drnevich, Ph.D. ##### Functional Genomics Bioinformatics Specialist ##### W.M. Keck Center for Comparative and Functional Genomics ##### Roy J. Carver Biotechnology Center, University of #####Illinois,Urbana-Champaign library("yeast2probe") ### ### ### ### ### ### The first part is just creating two ojects (ResetEnvir and RemoveProbes) originally written by Ariel Chernomoretz and modified by Jenny Drnevich to remove individual probes and/or entire probesets. Just highlight everything from here until you see STOP and paste it to R all at once ResetEnvir <- function(cleancdf){ cdfpackagename <- paste(cleancdf,"cdf",sep="") probepackagename <- paste(cleancdf,"probe",sep="") ll<-search() cdfpackagepos <- grep(cdfpackagename,ll) if(length(cdfpackagepos)>0) detach(pos=cdfpackagepos) ll<-search() probepackagepos <- grep(probepackagename,ll) if(length(probepackagepos)>0) detach(pos=probepackagepos) require(cdfpackagename,character.only=T) require(probepackagename,character.only=T) 303 require(affy) } RemoveProbes <- function(listOutProbes=NULL, listOutProbeSets=NULL, cleancdf,destructive=TRUE) { #default probe dataset values cdfpackagename <- paste(cleancdf,"cdf",sep="") probepackagename <- paste(cleancdf,"probe",sep="") require(cdfpackagename,character.only = TRUE) require(probepackagename,character.only = TRUE) probe.env.orig <- get(probepackagename) if(!is.null(listOutProbes)){ # taking probes out from CDF env probes<- unlist(lapply(listOutProbes,function(x){ a<-strsplit(x,"at") aux1<-paste(a[[1]][1],"at",sep="") aux2<-as.integer(a[[1]][2]) c(aux1,aux2) })) n1<-as.character(probes[seq(1,(length(probes)/2))*2-1]) n2<-as.integer(probes[seq(1,(length(probes)/2))*2]) probes<-data.frame(I(n1),n2) probes[,1]<-as.character(probes[,1]) probes[,2]<-as.integer(probes[,2]) pset<-unique(probes[,1]) for(i in seq(along=pset)){ ii <-grep(pset[i],probes[,1]) iout<-probes[ii,2] a<-get(pset[i],env=get(cdfpackagename)) a<-a[-iout,] assign(pset[i],a,env=get(cdfpackagename)) } } # taking probesets out from CDF env if(!is.null(listOutProbeSets)){ rm(list=listOutProbeSets,envir=get(cdfpackagename)) } # setting the PROBE env accordingly (idea from gcrma compute.affinities.R) tmp <- get("xy2indices",paste("package:",cdfpackagename,sep="")) newAB <- new("AffyBatch",cdfName=cleancdf) pmIndex <- unlist(indexProbes(newAB,"pm")) subIndex<match(tmp(probe.env.orig$x,probe.env.orig$y,cdf=cdfpackagename),pmIndex ) rm(newAB) 304 iNA } <- which(is.na(subIndex)) if(length(iNA)>0){ ipos<-grep(probepackagename,search()) assign(probepackagename,probe.env.orig[-iNA,],pos=ipos) } ### STOP HERE!!!! PASTE THE ABOVE INTO R AND CHECK TO SEE YOU HAVE ### THE TWO OBJECTS (ResetEnvir and RemoveProbes) WORKSPACE WITH ls() ## All you need now is your affybatch object, and a character vector ## of probe set names ## and/or another vector of individual probes that you want to remove. ## If your affybatch object is called 'rawdata' and the vector of probesets ## is 'maskedprobes', all you need to do is: ## here after: rawdata = aaa, maskedprobes <- scan("sp_IDs.txt", what="") maskedprobes[1] maskedprobes <- as.vector(maskedprobes) ## cleancdf <- cleancdfname(rawdata at cdfName,addcdf=FALSE) cleancdf <- cleancdfname(aaa @cdfName,addcdf=FALSE) # Make sure starting with the original cdf with all the # probes and probesets. ResetEnvir(cleancdf) # Double-check to make sure all probesets are present in your # affybatch by typing in # the name of your affybatch and looking at the output. # rawdata aaa # To remove some probe sets (but not individual probes in this #example), use: RemoveProbes(listOutProbes=NULL, listOutProbeSets=maskedprobes, cleancdf) # # # # # # # # The cdf file will be temporarily modified to mask the indicated probesets & probes, which you can check by typing in the name of your affybatch again and seeing that the number of probesets have decreased. The masking can be undone by using ResetEnvir as above, or by quitting the session. However, any Expression Set objects created when the cdf is modified will have the masked probesets removed permanently because they do not refer to the cdf like an affybatch object does. 305 aaa ## check id numbers to see if it decreases ## normalize with 5900 probesets library("affy") eset <- rma(aaa) dim(exprs(eset)) ### Box plot setwd("F://2007- Data in Hopper Lab//Microarray data analysis//R_Output//WT_mtr10_msn5_output") pdf("Box plot-after removingProbes and RMA normalized.pdf", width=5, height=5) par(mar=c(10,4,2,1)) boxplot(eset, main="After Remove probes and RMA normalized", ylab="log2 intensity", las=2, cex.axis=0.7) ; dev.off() ##### ##### ##### follow online manual(M. Bioconductor exercise) & add annotation info. ##### ##### ##### ### Generate RMA expression data, MAS5 P/M/A calls setwd("F://2007- Data in Hopper Lab//Microarray data analysis//R_Output//WT_mtr10_msn5_output") eset.pm <- mas5calls(aaa) # Generates MAS 5.0 P/M/A calls, not working in one cdf env? frame1 <- data.frame(exprs(eset), exprs(eset.pm)) # Combine RMA intensities, P/M/A calls in one data frame frame2 <- frame1[, sort(names(frame1))] # Sorts columns by cel file name. #write.table(frame1, file="All_expression value and PA calls.txt", sep="\t") #write.table(frame2, file="All_expression value and PA calls_sortbyname.txt", sep="\t") ############# ############# ############# ############# ############# ############# Pre-processing #### #### step 1. filter -- to get rid of low/no singals #### (by log2intensities, no hypothesis tested) #### library("genefilter") 306 f1 <- anyNA f2 <- pOverA(0.50, 6) ff <- filterfun(f1, f2) selected <- genefilter(exprs(eset), ff) esetsub <- eset[selected,] ef <- exprs(esetsub) ; ; sum(selected) ## esetsub is a filtered matrix ef[1:5, 1:3]; dim(ef) write.table(ef, file="All_RMA_expression value_filtered.txt",quote=FALSE, sep='\t') frame2.esetsub <- frame2[selected,]; dim(frame2.esetsub) write.table(frame2.esetsub, file="All_RMA_expression value_filtered plusPAcall.txt",quote=FALSE, sep='\t') ####### ####### ####### add and export annotation info. library("annotate") ; library("yeast2.db") ; ####### ####### ####### for all probesets ls("package:annotate") ls("package:yeast2.db") annotation(eset) <- "yeast2.db" annotation(esetsub) <- "yeast2.db" ## get information for esetsub probeIDs <- featureNames(esetsub) # get affyIDs from esetsub length(probeIDs) ; probeIDs[5] # check # Retrieves gene descriptions symbols <- as.vector(unlist(mget(probeIDs,yeast2GENENAME))) symbols[5] ; length(symbols) # check orf <- as.vector(unlist(mget(probeIDs,yeast2ORF))) ; orf[1:5] ;length(orf) # Retrieves ORF desrp <- as.vector(unlist(mget(probeIDs, yeast2DESCRIPTION))) sym.orf.des <- as.vector(paste(symbols, orf, desrp, sep=".")) sym.orf.des[1:3] sym.des <- as.vector(paste(symbols, desrp, sep=".")) # build data frame df <- as.data.frame(exprs(esetsub)) ; ; dim(df) # Generates data frame with above annotation data. Annot <- data.frame(probeIDs, orf, symbols, desrp, sym.des, df, row.names=NULL) ; dim(Annot) write.table(Annot, file="annot_filtered_dataset_no PA.txt", sep="\t",quote=FALSE) 307 ############# ############# ############# ############# ############# ############# To generate DEG ##### ##### 1. Build dataframe ##### wt.pm.f <- exprs(esetsub[, 1:2])- exprs(esetsub[, 3:4]) ; dim(wt.pm.f) wt.pm.f.avg <- rowMeans(wt.pm.f) wt.pm.s <- exprs(esetsub[, 5:6])- exprs(esetsub[, 7:8]) ; dim(wt.pm.s) wt.pm.s.avg <- rowMeans(wt.pm.s) wt.pm.sf <- wt.pm.s - wt.pm.f ; dim(wt.pm.sf) wt.pm.sf.avg <- rowMeans(wt.pm.sf) msn5.pm.f <- exprs(esetsub[, 9:10]) - exprs(esetsub[, 11:12]) msn5.pm.f.avg <- rowMeans(msn5.pm.f) msn5.pm.s <- exprs(esetsub[, 13:14])- exprs(esetsub[, 15:16]) msn5.pm.s.avg <- rowMeans(msn5.pm.s) msn5.pm.sf <- msn5.pm.s - msn5.pm.f msn5.pm.sf.avg <- rowMeans(msn5.pm.sf) mtr10.pm.f <- exprs(esetsub[, 17:18]) - exprs(esetsub[, 19:20]) mtr10.pm.f.avg <- rowMeans(mtr10.pm.f) mtr10.pm.s <- exprs(esetsub[, 21:22])- exprs(esetsub[, 23:24]) mtr10.pm.s.avg <- rowMeans(mtr10.pm.s) mtr10.pm.sf <- mtr10.pm.s - mtr10.pm.f mtr10.pm.sf.avg <- rowMeans(mtr10.pm.sf) wt.pt.f <- exprs(esetsub[, 1:2])- exprs(esetsub[, 25:26]) dim(wt.pt.f) wt.pt.f.avg <- rowMeans(wt.pt.f) wt.pt.s <- exprs(esetsub[, 5:6])- exprs(esetsub[, 27:28]) dim(wt.pt.s) wt.pt.s.avg <- rowMeans(wt.pt.s) wt.pt.sf <- wt.pt.s - wt.pt.f ; dim(wt.pt.sf) wt.pt.sf.avg <- rowMeans(wt.pt.sf) ; ; msn5.pt.f <- exprs(esetsub[, 9:10])- exprs(esetsub[, 29:30]) ; dim(msn5.pt.f) msn5.pt.f.avg <- rowMeans(msn5.pt.f) msn5.pt.s <- exprs(esetsub[, 13:14])-exprs(esetsub[, 31:32]) msn5.pt.s.avg <- rowMeans(msn5.pt.s) msn5.pt.sf <- msn5.pt.s - msn5.pt.f msn5.pt.sf.avg <- rowMeans(msn5.pt.sf) mtr10.pt.f <- exprs(esetsub[, 17:18])- exprs(esetsub[, 33:34]) ; dim(mtr10.pt.f) mtr10.pt.f.avg <- rowMeans(mtr10.pt.f) mtr10.pt.s <- exprs(esetsub[, 21:22])-exprs(esetsub[, 35:36]) mtr10.pt.s.avg <- rowMeans(mtr10.pt.s) mtr10.pt.sf <- mtr10.pt.s - mtr10.pt.f mtr10.pt.sf.avg <- rowMeans(mtr10.pt.sf) 308 ### TXN wt.t.f wt.t.s wt.t.f.avg wt.t.s.avg <<<<- msn5.t.f <msn5.t.s <msn5.t.f.avg msn5.t.s.avg exprs(esetsub[, 25:26]) ; dim(wt.t.f) exprs(esetsub[, 27:28]) rowMeans (wt.t.f) rowMeans (wt.t.s) exprs(esetsub[, 29:30]) exprs(esetsub[, 31:32]) <- rowMeans (msn5.t.f) <- rowMeans (msn5.t.s) mtr10.t.f <mtr10.t.s <mtr10.t.f.avg mtr10.t.s.avg exprs(esetsub[, 33:34]) exprs(esetsub[, 35:36]) <- rowMeans (mtr10.t.f) <- rowMeans (mtr10.t.s) setwd("F:/2007- Data in Hopper Lab/Microarray data analysis/R_Output/WT_mtr10_msn5_output") pm.mtr10.and.wt <- cbind(wt.pm.f , mtr10.pm.f, wt.pm.s , mtr10.pm.s) dim(pm.mtr10.and.wt) pm.msn5.and.wt <- cbind(wt.pm.f , msn5.pm.f, wt.pm.s , msn5.pm.s) dim(pm.msn5.and.wt) txn.mtr10.and.wt <- cbind(wt.t.f , mtr10.t.f, wt.t.s , mtr10.t.s) dim(txn.mtr10.and.wt) txn.msn5.and.wt <- cbind(wt.t.f , msn5.t.f, wt.t.s , msn5.t.s) ; dim(txn.msn5.and.wt) pt.mtr10.and.wt <- cbind(wt.pt.f , mtr10.pt.f, wt.pt.s , mtr10.pt.s) dim(pt.mtr10.and.wt) ###### ###### Step 2. statistical analysis -- limma ###### ##### Step 2.1 Model-fit ##### ##### compare P/M values library("limma") dim(pm.mtr10.and.wt) dim(pm.msn5.and.wt) ### (Pf/Mf) or (Ps/Ms) ### mtr10 pm.mtr10.and.wt.exprs <- new("ExpressionSet", exprs=as.matrix(pm.mtr10.and.wt)) ; dim(pm.mtr10.and.wt.exprs) mtr10.design <- model.matrix(~0+factor(c(1,1,2,2,3,3,4,4))) colnames(mtr10.design) <- c("wt.f", "mtr10.f", "wt.s", "mtr10.s") ; mtr10.design 309 ; ; ; ; mtr10.fit <- lmFit(exprs(pm.mtr10.and.wt.exprs), mtr10.design) contrast.matrix.mtr10 <- makeContrasts("Pf/Mf-mtr10/wt" = mtr10.f-wt.f, "Ps/Ms-mtr10/wt" = mtr10.s-wt.s, levels=mtr10.design) contrast.matrix.mtr10 fit2.mtr10 <- contrasts.fit(mtr10.fit, contrast.matrix.mtr10) fit3.mtr10 <- eBayes(fit2.mtr10) ### msn5 pm.msn5.and.wt.exprs <- new("ExpressionSet", exprs=as.matrix(pm.msn5.and.wt)) ; dim(pm.msn5.and.wt.exprs) msn5.design <- model.matrix(~0+factor(c(1,1,2,2,3,3,4,4))) colnames(msn5.design) <- c("wt.f", "msn5.f", "wt.s", "msn5.s") ; msn5.design msn5.fit <- lmFit(exprs(pm.msn5.and.wt.exprs), msn5.design) contrast.matrix.msn5 <- makeContrasts("Pf/Mf-msn5/wt" = msn5.f-wt.f, "Ps/Ms-msn5/wt" = msn5.s-wt.s, levels=msn5.design) contrast.matrix.msn5 fit2.msn5 <- contrasts.fit(msn5.fit, contrast.matrix.msn5) fit3.msn5 <- eBayes(fit2.msn5) ##### ##### Model-fit – compare P/T values ##### library("limma") dim(pt.mtr10.and.wt) ### (Pf/Tf) or (Ps/Ts) ### mtr10 pt.mtr10.and.wt.exprs <- new("ExpressionSet", exprs=as.matrix(pt.mtr10.and.wt)) ; dim(pt.mtr10.and.wt.exprs) pt.design <- model.matrix(~0+factor(c(1,1,2,2,3,3,4,4))) colnames(pt.design) <- c("wt.f", "mtr10.f", "wt.s", "mtr10.s") ; pt.design pt.fit <- lmFit(exprs(pt.mtr10.and.wt.exprs), pt.design) pt.contrast.matrix <- makeContrasts("Pf/Tf-mtr10/wt" = mtr10.f-wt.f, "Ps/Ts-mtr10/wt" = mtr10.s-wt.s, levels=pt.design) pt.contrast.matrix pt.fit2 <- contrasts.fit(pt.fit, pt.contrast.matrix) pt.fit3 <- eBayes(pt.fit2) 310 ##### ##### Model-fit – compare Txn (T) values ##### library("limma") dim(txn.mtr10.and.wt) ### Tf or Ts ### mtr10 txn.mtr10.and.wt.exprs <- new("ExpressionSet", exprs=as.matrix(txn.mtr10.and.wt)) ; dim(txn.mtr10.and.wt.exprs) design.txn.mtr10 <- model.matrix(~0+factor(c(1,1,2,2,3,3,4,4))) colnames(design.txn.mtr10) <- c("wt.Tf", "mtr10.Tf", "wt.Ts", "mtr10.Ts") ; design.txn.mtr10 fit.txn.mtr10 <- lmFit(exprs(txn.mtr10.and.wt.exprs), design.txn.mtr10) contrast.matrix.txn.mtr10 <- makeContrasts("Tf-mtr10/wt" = mtr10.Tfwt.Tf, "Ts-mtr10/wt" = mtr10.Ts-wt.Ts, levels=design.txn.mtr10) contrast.matrix.txn.mtr10 fit2.txn.mtr10 <- contrasts.fit(fit.txn.mtr10, contrast.matrix.txn.mtr10) fit3.txn.mtr10 <- eBayes(fit2.txn.mtr10) ### msn5 txn.msn5.and.wt.exprs <- new("ExpressionSet", exprs=as.matrix(txn.msn5.and.wt)) ; dim(txn.msn5.and.wt.exprs) design.txn.msn5 <- model.matrix(~0+factor(c(1,1,2,2,3,3,4,4))) colnames(design.txn.msn5) <- c("wt.Tf", "msn5.Tf", "wt.Ts", "msn5.Ts") ; design.txn.msn5 fit.txn.msn5 <- lmFit(exprs(txn.msn5.and.wt.exprs), design.txn.msn5) contrast.matrix.txn.msn5 <- makeContrasts("Tf-msn5/wt" = msn5.Tf-wt.Tf, "Ts-msn5/wt" = msn5.Ts-wt.Ts, levels=design.txn.msn5) contrast.matrix.txn.msn5 fit2.txn.msn5 <- contrasts.fit(fit.txn.msn5, contrast.matrix.txn.msn5) fit3.txn.msn5 <- eBayes(fit2.txn.msn5) ##### ##### ##### ##### Step 2.2 Select Deferential-expressed genes (DEG) P/M values ### Pf/Mf or Ps/Ms results.mtr10 <- decideTests(fit3.mtr10, method="separate", adjust.method="BH", p.value=0.05) ; summary(results.mtr10) results.msn5 <- decideTests(fit3.msn5, method="separate", adjust.method="BH", p.value=0.05) ; summary(results.msn5) pdf("Venn Diagram- PM values_mtr10 vs wt.pdf") 311 vennDiagram(results.mtr10, include=c("up","down"), cex=1.2, lwd=2, counts.col=c("red", "green4"), circle.col=c("burlywood4"), main= "P/M value_p value cutoff 0.05") ; dev.off() pdf("Venn Diagram- PM values_msn5 vs wt.pdf") vennDiagram(results.msn5, include=c("up","down"), cex=1.2, lwd=2, counts.col=c("red", "green4"), circle.col=c("burlywood4"), main= "P/M value_p value cutoff 0.05") ; dev.off() top.pm.f.mtr10 <- topTable(fit3.mtr10, coef=1, adjust="BH", sort.by="P", number=6000) top.pm.f.mtr10 <- top.pm.f.mtr10[top.pm.f.mtr10$adj.P.Val <0.05,] ; dim(top.pm.f.mtr10) top.pm.f.up.mtr10 <- top.pm.f.mtr10[top.pm.f.mtr10$adj.P.Val <0.05 &(top.pm.f.mtr10 $logFC > 0.6 ),] ; dim(top.pm.f.up.mtr10) top.pm.f.down.mtr10 <- top.pm.f.mtr10[top.pm.f.mtr10$adj.P.Val <0.05 &(top.pm.f.mtr10 $logFC < -0.6 ),] ; dim(top.pm.f.down.mtr10) top.pm.s.mtr10 <- topTable(fit3.mtr10, coef=2, adjust="BH", sort.by="P", number=6000) top.pm.s.mtr10 <- top.pm.s.mtr10[top.pm.s.mtr10$adj.P.Val <0.05,] ; dim(top.pm.s.mtr10) top.pm.s.up.mtr10 <- top.pm.s.mtr10[top.pm.s.mtr10$adj.P.Val <0.05 &(top.pm.s.mtr10 $logFC > 0.6 ),] ; dim(top.pm.s.up.mtr10) top.pm.s.down.mtr10 <- top.pm.s.mtr10[top.pm.s.mtr10$adj.P.Val <0.05 &(top.pm.s.mtr10 $logFC < -0.6 ),] ; dim(top.pm.s.down.mtr10) top.pm.f.msn5 <- topTable(fit3.msn5, coef=1, adjust="BH", sort.by="P", number=6000) top.pm.f.msn5 <- top.pm.f.msn5[top.pm.f.msn5$adj.P.Val <0.05,] ; dim(top.pm.f.msn5) top.pm.f.up.msn5 <- top.pm.f.msn5[top.pm.f.msn5$adj.P.Val <0.05 &(top.pm.f.msn5 $logFC > 0 ),] ; dim(top.pm.f.up.msn5) top.pm.f.down.msn5 <- top.pm.f.msn5[top.pm.f.msn5$adj.P.Val <0.05 &(top.pm.f.msn5 $logFC < 0 ),] ; dim(top.pm.f.down.msn5) top.pm.s.msn5 <- topTable(fit3.msn5, coef=2, adjust="BH", sort.by="P", number=6000) top.pm.s.msn5 <- top.pm.s.msn5[top.pm.s.msn5$adj.P.Val <0.05,] ; dim(top.pm.s.msn5) top.pm.s.up.msn5 <- top.pm.s.msn5[top.pm.s.msn5$adj.P.Val <0.05 &(top.pm.s.msn5 $logFC > 0 ),] ; dim(top.pm.s.up.msn5) top.pm.s.down.msn5 <- top.pm.s.msn5[top.pm.s.msn5$adj.P.Val <0.05 &(top.pm.s.msn5 $logFC < 0 ),] ; dim(top.pm.s.down.msn5) nn <- intersect (top.pm.f.down.mtr10$ID, top.pm.f.down.msn5$ID);length(nn) # number is 7 312 ##### ##### ##### Select DEG -- P/T values ### Pf/Tf or Ps/Ts## use adj.p value 0.01 for DEG selection results.pt.mtr10 <- decideTests(pt.fit3, method="separate", adjust.method="BH", p.value=0.01, lfc=0.6); summary(results.pt.mtr10) pdf("Venn Diagram- PT values_mtr10 vs wt.pdf") vennDiagram(results.pt.mtr10, include=c("up","down"), cex=1.2, lwd=2, counts.col=c("red", "green4"), circle.col=c("burlywood4"), main= "P/T value_p value cutoff 0.01 and lfc=0.6") ; dev.off() top.pt.f.mtr10 <- topTable(pt.fit3, coef=1, adjust="BH", sort.by="P", number=6000) top.pt.f.mtr10 <- top.pt.f.mtr10[top.pt.f.mtr10$adj.P.Val <0.01,] ; dim(top.pt.f.mtr10) top.pt.f.mtr10.up <- top.pt.f.mtr10[top.pt.f.mtr10$adj.P.Val <0.01 &(top.pt.f.mtr10 $logFC > 0.6 ),] ; dim(top.pt.f.mtr10.up) top.pt.f.mtr10.down <- top.pt.f.mtr10[top.pt.f.mtr10$adj.P.Val <0.01 &(top.pt.f.mtr10 $logFC < -0.6 ),] ; dim(top.pt.f.mtr10.down) top.pt.s.mtr10 <- topTable(pt.fit3, coef=2, adjust="BH", sort.by="P", number=6000) top.pt.s.mtr10 <- top.pt.s.mtr10[top.pt.s.mtr10$adj.P.Val <0.01,] ; dim(top.pt.s.mtr10) top.pt.s.mtr10.up <- top.pt.s.mtr10[top.pt.s.mtr10$adj.P.Val <0.01 &(top.pt.s.mtr10 $logFC > 0.6 ),] ; dim(top.pt.s.mtr10.up) top.pt.s.mtr10.down <- top.pt.s.mtr10[top.pt.s.mtr10$adj.P.Val <0.01 &(top.pt.s.mtr10 $logFC < -0.6 ),] ; dim(top.pt.s.mtr10.down) ##### ##### ##### Select DEG -- Txn values #### Tf or Ts results.txn.mtr10 <- decideTests(fit3.txn.mtr10, method="separate", adjust.method="BH", p.value=0.001, lfc=0.6) ; summary(results.txn.mtr10) results.txn.msn5 <- decideTests(fit3.txn.msn5, method="separate", adjust.method="BH", p.value=0.05, lfc=0.6) ; summary(results.txn.msn5) pdf("Venn Diagram- Txn values_mtr10 vs wt.pdf") vennDiagram(results.txn.mtr10, include=c("up","down"), cex=1.2, lwd=3, counts.col=c("red2", "green4"), circle.col=c("burlywood4"), main= "Txn_p value cutoff 0.001 and lfc=0.6") ; dev.off() 313 pdf("Venn Diagram- Txn values_msn5 vs wt.pdf") vennDiagram(results.txn.msn5, include=c("up","down"), cex=1.2, lwd=3, counts.col=c("red2", "green4"), circle.col=c("burlywood4"), main= "Txn_p value cutoff 0.05 and lfc=0.6") ; dev.off() top.Tf.mtr10 <- topTable(fit3.txn.mtr10, coef=1, adjust="BH", sort.by="P", number=6000) top.Tf.mtr10 <- top.Tf.mtr10[top.Tf.mtr10$adj.P.Val <0.001,] ; dim(top.Tf.mtr10) top.Tf.up.mtr10 <- top.Tf.mtr10[top.Tf.mtr10$adj.P.Val <0.001 & (top.Tf.mtr10 $logFC > 0.6 ),]; dim(top.Tf.up.mtr10) top.Tf.down.mtr10 <- top.Tf.mtr10[top.Tf.mtr10$adj.P.Val <0.001 & (top.Tf.mtr10 $logFC < -0.6 ),]; dim(top.Tf.down.mtr10) top.Ts.mtr10 <- topTable(fit3.txn.mtr10, coef=2, adjust="BH", sort.by="P", number=6000) top.Ts.mtr10 <- top.Ts.mtr10 [top.Ts.mtr10 $adj.P.Val <0.001,] ; dim(top.Ts.mtr10 ) top.Ts.up.mtr10 <- top.Ts.mtr10 [top.Ts.mtr10 $adj.P.Val <0.001 & (top.Ts.mtr10 $logFC > 0.6 ),]; dim(top.Ts.up.mtr10 ) top.Ts.down.mtr10 <- top.Ts.mtr10 [top.Ts.mtr10 $adj.P.Val <0.001 & (top.Ts.mtr10 $logFC < -0.6 ),]; dim(top.Ts.down.mtr10 ) top.Tf.msn5 <- topTable(fit3.txn.msn5, coef=1, adjust="BH", sort.by="P", number=6000) top.Tf.msn5 <- top.Tf.msn5[top.Tf.msn5$adj.P.Val <0.05,] ; dim(top.Tf.msn5) top.Tf.up.msn5 <- top.Tf.msn5[top.Tf.msn5$adj.P.Val <0.05 & (top.Tf.msn5 $logFC > 0.6 ),]; dim(top.Tf.up.msn5) top.Tf.down.msn5 <- top.Tf.msn5[top.Tf.msn5$adj.P.Val <0.05 & (top.Tf.msn5 $logFC < -0.6 ),]; dim(top.Tf.down.msn5) top.Ts.msn5 <- topTable(fit3.txn.msn5, coef=2, adjust="BH", sort.by="P", number=6000) top.Ts.msn5 <- top.Ts.msn5 [top.Ts.msn5 $adj.P.Val <0.05,] ; dim(top.Ts.msn5 ) top.Ts.up.msn5 <- top.Ts.msn5 [top.Ts.msn5 $adj.P.Val <0.05 & (top.Ts.msn5 $logFC > 0.6 ),]; dim(top.Ts.up.msn5 ) top.Ts.down.msn5 <- top.Ts.msn5 [top.Ts.msn5 $adj.P.Val <0.05 & (top.Ts.msn5 $logFC < -0.6 ),]; dim(top.Ts.down.msn5 ) rr.up <- intersect(top.Tf.up.msn5$ID , top.Tf.up.mtr10$ID) ;length(rr.up) rr.down <- intersect(top.Tf.down.msn5$ID , top.Tf.down.mtr10$ID) ;length(rr.down) yy.up <- intersect(top.Ts.up.msn5$ID, top.Ts.up.mtr10$ID) ;length(yy.up) yy.down <- intersect(top.Ts.down.msn5$ID, top.Ts.down.mtr10$ID) ;length(yy.down) 314 ##### ##### ##### Step 2.3 Build annotation tables with expression values ##### ##### mtr10 Generate mtr10 annotation table with P/M up-regulated genes P/M values top.pm.f.up.mtr10.id <- as.vector(top.pm.f.up.mtr10$ID) ;length(top.f.up.mtr10.id) # Retrieves gene descriptions symbols <- as.vector(unlist(mget(top.pm.f.up.mtr10.id, yeast2GENENAME))) # Retrieves ORF ORF <- as.vector(unlist(mget(top.pm.f.up.mtr10.id, yeast2ORF))) des <- as.vector(unlist(mget(top.pm.f.up.mtr10.id, yeast2DESCRIPTION))) # Build together sym.orf.des <- as.vector(paste(symbols, ORF, des, sep=".")) top.pm.f.up.df.mtr10 <- data.frame(probeIDs=top.pm.f.up.mtr10.id, symbols, ORF, des, sym.orf.des);dim(top.pm.f.up.df.mtr10) # Retrieves expression value pm.f.up.mtr10.2 <data.frame(exprs(pm.mtr10.and.wt.exprs)[top.pm.f.up.mtr10.id, ]) # Merges everything with above expression data. output.pm.f.up.mtr10 <- merge(top.pm.f.up.df.mtr10 ,pm.f.up.mtr10.2, by.x="probeIDs", by.y=0, all=T);dim(output.pm.f.up.mtr10) output.pm.f.up.mtr10.2 <- cbind(output.pm.f.up.mtr10, top.pm.f.up.mtr10) ;dim(output.pm.f.up.mtr10.2) # Export dataframe write.table(output.pm.f.up.mtr10.2, file="annot_limma_P0.05_pm.f.up.mtr10_vs_WT_usesmall dataset.txt", sep="\t",quote=FALSE) ##### Generate mtr10 annotation table with P/M down-regulated genes top.pm.f.down.mtr10.id <- as.vector(top.pm.f.down.mtr10$ID) ;length(top.pm.f.down.mtr10.id) symbols <- as.vector(unlist(mget(top.pm.f.down.mtr10.id, yeast2GENENAME))) ORF <- as.vector(unlist(mget(top.pm.f.down.mtr10.id, yeast2ORF))) des <- as.vector(unlist(mget(top.pm.f.down.mtr10.id, yeast2DESCRIPTION))) sym.orf.des <- as.vector(paste(symbols, ORF, des, sep=".")) top.pm.f.down.df.mtr10 <- data.frame(probeIDs=top.pm.f.down.mtr10.id, symbols, ORF, des, sym.orf.des);dim(top.pm.f.down.df.mtr10) pm.f.down.mtr10.2 <data.frame(exprs(pm.mtr10.and.wt.exprs)[top.pm.f.down.mtr10.id, ]) 315 output.pm.f.down.mtr10 <- merge(top.pm.f.down.df.mtr10 ,pm.f.down.mtr10.2, by.x="probeIDs", by.y=0, all=T);dim(output.pm.f.down.mtr10) output.pm.f.down.mtr10.2 <- cbind(output.pm.f.down.mtr10, top.pm.f.down.mtr10) ;dim(output.pm.f.down.mtr10.2) write.table(output.pm.f.down.mtr10.2, file="annot_limma_P0.05_pm.f.down.mtr10_vs_WT_usesmall dataset.txt", sep="\t",quote=FALSE) ### Generate mtr10 annotation table with P/M.starved up-regulated ###genes top.pm.s.up.mtr10.id <- as.vector(top.pm.s.up.mtr10$ID) ;length(top.pm.s.up.mtr10.id) symbols <- as.vector(unlist(mget(top.pm.s.up.mtr10.id, yeast2GENENAME))) ORF <- as.vector(unlist(mget(top.pm.s.up.mtr10.id, yeast2ORF))) des <- as.vector(unlist(mget(top.pm.s.up.mtr10.id, yeast2DESCRIPTION))) sym.orf.des <- as.vector(paste(symbols, ORF, des, sep=".")) top.pm.s.up.df.mtr10 <- data.frame(probeIDs=top.pm.s.up.mtr10.id, symbols, ORF, des, sym.orf.des);dim(top.pm.s.up.df.mtr10) pm.s.up.mtr10.2 <data.frame(exprs(pm.mtr10.and.wt.exprs)[top.pm.s.up.mtr10.id, ]) output.pm.s.up.mtr10 <- merge(top.pm.s.up.df.mtr10 ,pm.s.up.mtr10.2, by.x="probeIDs", by.y=0, all=T);dim(output.pm.s.up.mtr10) output.pm.s.up.mtr10.2 <- cbind(output.pm.s.up.mtr10, top.pm.s.up.mtr10) ;dim(output.pm.s.up.mtr10.2) write.table(output.pm.s.up.mtr10.2, file="annot_limma_P0.05_pm.s.up.mtr10_vs_WT_usesmall dataset.txt", sep="\t",quote=FALSE) ### Generate mtr10 annotation table with P/M.starved down-regulated ###genes top.pm.s.down.mtr10.id <- as.vector(top.pm.s.down.mtr10$ID) ;length(top.pm.s.down.mtr10.id) symbols <- as.vector(unlist(mget(top.pm.s.down.mtr10.id, yeast2GENENAME))) ORF <- as.vector(unlist(mget(top.pm.s.down.mtr10.id, yeast2ORF))) des <- as.vector(unlist(mget(top.pm.s.down.mtr10.id, yeast2DESCRIPTION))) sym.orf.des <- as.vector(paste(symbols, ORF, des, sep=".")) top.pm.s.down.df.mtr10 <- data.frame(probeIDs=top.pm.s.down.mtr10.id, symbols, ORF, des, sym.orf.des);dim(top.pm.s.down.df.mtr10) pm.s.down.mtr10.2 <data.frame(exprs(pm.mtr10.and.wt.exprs)[top.pm.s.down.mtr10.id, ]) output.pm.s.down.mtr10 <- merge(top.pm.s.down.df.mtr10 ,pm.s.down.mtr10.2, by.x="probeIDs", by.y=0, all=T);dim(output.pm.s.down.mtr10) output.pm.s.down.mtr10.2 <cbind(output.pm.s.down.mtr10, top.pm.s.down.mtr10) ;dim(output.pm.s.down.mtr10.2) write.table(output.pm.s.down.mtr10.2, file="annot_limma_P0.05_pm.s.down.mtr10_vs_WT_usesmall dataset.txt", sep="\t",quote=FALSE) 316 ### msn5 ### P/M values dim(pm.msn5.and.wt.exprs) ### Generate msn5 annotation table with P/M.fed down-regulated genes top.pm.f.down.msn5.id <- as.vector(top.pm.f.down.msn5$ID) ;length(top.pm.f.down.msn5.id) symbols <- as.vector(unlist(mget(top.pm.f.down.msn5.id, yeast2GENENAME))) ORF <- as.vector(unlist(mget(top.pm.f.down.msn5.id, yeast2ORF))) des <- as.vector(unlist(mget(top.pm.f.down.msn5.id, yeast2DESCRIPTION))) sym.orf.des <- as.vector(paste(symbols, ORF, des, sep=".")) top.pm.f.down.df.msn5 <- data.frame(probeIDs=top.pm.f.down.msn5.id, symbols, ORF, des, sym.orf.des);dim(top.pm.f.down.df.msn5) pm.f.down.msn5.2 <data.frame(exprs(pm.msn5.and.wt.exprs)[top.pm.f.down.msn5.id, ]) output.pm.f.down.msn5 <- merge(top.pm.f.down.df.msn5 ,pm.f.down.msn5.2, by.x="probeIDs", by.y=0, all=T);dim(output.pm.f.down.msn5) output.pm.f.down.msn5.2 <cbind(output.pm.f.down.msn5, top.pm.f.down.msn5) ;dim(output.pm.f.down.msn5.2) write.table(output.pm.f.down.msn5.2, file="annot_limma_P0.05_pm.f.down.msn5_vs_WT_usesmall dataset.txt", sep="\t",quote=FALSE) ## P/T values ## mtr10 only dim(pt.mtr10.and.wt.exprs) ### Generate mtr10 annotation table with P/T.fed up-regulated genes top.pt.f.mtr10.up.id <- as.vector(top.pt.f.mtr10.up$ID) ;length(top.pt.f.mtr10.up.id ) symbols <- as.vector(unlist(mget(top.pt.f.mtr10.up.id, yeast2GENENAME))) ORF <- as.vector(unlist(mget(top.pt.f.mtr10.up.id, yeast2ORF))) des <- as.vector(unlist(mget(top.pt.f.mtr10.up.id, yeast2DESCRIPTION))) sym.orf.des <- as.vector(paste(symbols, ORF, des, sep=".")) top.pt.f.up.df.mtr10 <- data.frame(probeIDs=top.pt.f.mtr10.up.id, symbols, ORF, des, sym.orf.des);dim(top.pt.f.up.df.mtr10) pt.f.up.mtr10.2 <data.frame(exprs(pt.mtr10.and.wt.exprs)[top.pt.f.mtr10.up.id, ]) output.pt.f.up.mtr10 <- merge(top.pt.f.up.df.mtr10 ,pt.f.up.mtr10.2, by.x="probeIDs", by.y=0, all=T);dim(output.pt.f.up.mtr10) output.pt.f.up.mtr10.2 <- cbind(output.pt.f.up.mtr10, top.pt.f.mtr10.up) ;dim(output.pt.f.up.mtr10.2) write.table(output.pt.f.up.mtr10.2, file="annot_limma_P0.01_pt.f.up.mtr10_vs_WT_usesmall dataset.txt", sep="\t",quote=FALSE) 317 ### Generate mtr10 annotation table with P/T.fed down-regulated genes top.pt.f.mtr10.down.id <- as.vector(top.pt.f.mtr10.down$ID) ;length(top.pt.f.mtr10.down.id ) symbols <- as.vector(unlist(mget(top.pt.f.mtr10.down.id, yeast2GENENAME))) ORF <- as.vector(unlist(mget(top.pt.f.mtr10.down.id, yeast2ORF))) des <- as.vector(unlist(mget(top.pt.f.mtr10.down.id, yeast2DESCRIPTION))) sym.orf.des <- as.vector(paste(symbols, ORF, des, sep=".")) top.pt.f.down.df.mtr10 <- data.frame(probeIDs=top.pt.f.mtr10.down.id, symbols, ORF, des, sym.orf.des);dim(top.pt.f.down.df.mtr10) pt.f.down.mtr10.2 <data.frame(exprs(pt.mtr10.and.wt.exprs)[top.pt.f.mtr10.down.id, ]) output.pt.f.down.mtr10 <- merge(top.pt.f.down.df.mtr10 ,pt.f.down.mtr10.2, by.x="probeIDs", by.y=0, all=T);dim(output.pt.f.down.mtr10) output.pt.f.down.mtr10.2 <cbind(output.pt.f.down.mtr10, top.pt.f.mtr10.down) ;dim(output.pt.f.down.mtr10.2) write.table(output.pt.f.down.mtr10.2, file="annot_limma_P0.01_pt.f.down.mtr10_vs_WT_usesmall dataset.txt", sep="\t",quote=FALSE) ### Generate mtr10 annotation table with P/T.starved up-regulated genes top.pt.s.mtr10.up.id <- as.vector(top.pt.s.mtr10.up$ID) ;length(top.pt.s.mtr10.up.id ) symbols <- as.vector(unlist(mget(top.pt.s.mtr10.up.id, yeast2GENENAME))) ORF <- as.vector(unlist(mget(top.pt.s.mtr10.up.id, yeast2ORF))) des <- as.vector(unlist(mget(top.pt.s.mtr10.up.id, yeast2DESCRIPTION))) sym.orf.des <- as.vector(paste(symbols, ORF, des, sep=".")) top.pt.s.up.df.mtr10 <- data.frame(probeIDs=top.pt.s.mtr10.up.id, symbols, ORF, des, sym.orf.des);dim(top.pt.s.up.df.mtr10) pt.s.up.mtr10.2 <data.frame(exprs(pt.mtr10.and.wt.exprs)[top.pt.s.mtr10.up.id, ]) output.pt.s.up.mtr10 <- merge(top.pt.s.up.df.mtr10 ,pt.s.up.mtr10.2, by.x="probeIDs", by.y=0, all=T);dim(output.pt.s.up.mtr10) output.pt.s.up.mtr10.2 <- cbind(output.pt.s.up.mtr10, top.pt.s.mtr10.up) ;dim(output.pt.s.up.mtr10.2) write.table(output.pt.s.up.mtr10.2, file="annot_limma_P0.01_pt.s.up.mtr10_vs_WT_usesmall dataset.txt", sep="\t",quote=FALSE) ### Generate mtr10 annotation table with P/T.starved down-regulated ###genes top.pt.s.mtr10.down.id <- as.vector(top.pt.s.mtr10.down$ID) ;length(top.pt.s.mtr10.down.id ) symbols <- as.vector(unlist(mget(top.pt.s.mtr10.down.id, yeast2GENENAME))) ORF <- as.vector(unlist(mget(top.pt.s.mtr10.down.id, yeast2ORF))) des <- as.vector(unlist(mget(top.pt.s.mtr10.down.id, yeast2DESCRIPTION))) sym.orf.des <- as.vector(paste(symbols, ORF, des, sep=".")) 318 top.pt.s.down.df.mtr10 <- data.frame(probeIDs=top.pt.s.mtr10.down.id, symbols, ORF, des, sym.orf.des);dim(top.pt.s.down.df.mtr10) pt.s.down.mtr10.2 <data.frame(exprs(pt.mtr10.and.wt.exprs)[top.pt.s.mtr10.down.id, ]) output.pt.s.down.mtr10 <- merge(top.pt.s.down.df.mtr10 ,pt.s.down.mtr10.2, by.x="probeIDs", by.y=0, all=T);dim(output.pt.s.down.mtr10) output.pt.s.down.mtr10.2 <- cbind(output.pt.s.down.mtr10, top.pt.s.mtr10.down) ;dim(output.pt.s.down.mtr10.2) write.table(output.pt.s.down.mtr10.2, file="annot_limma_P0.01_pt.s.down.mtr10_vs_WT_usesmall dataset.txt", sep="\t",quote=FALSE) ## Txn values ## mtr10 dim(txn.mtr10.and.wt.exprs) ### Generate mtr10 annotation table with Total.fed up-regulated ###genes top.Tf.up.mtr10.id <- as.vector(top.Tf.up.mtr10$ID) ;length(top.Tf.up.mtr10.id) symbols <- as.vector(unlist(mget(top.Tf.up.mtr10.id, yeast2GENENAME))) ORF <- as.vector(unlist(mget(top.Tf.up.mtr10.id, yeast2ORF))) des <- as.vector(unlist(mget(top.Tf.up.mtr10.id, yeast2DESCRIPTION))) sym.orf.des <- as.vector(paste(symbols, ORF, des, sep=".")) top.Tf.up.df.mtr10 <- data.frame(probeIDs=top.Tf.up.mtr10.id, symbols, ORF, des, sym.orf.des);dim(top.Tf.up.df.mtr10) Tf.up.mtr10.2 <data.frame(exprs(pm.mtr10.and.wt.exprs)[top.Tf.up.mtr10.id, ]) output.Tf.up.mtr10 <- merge(top.Tf.up.df.mtr10 ,Tf.up.mtr10.2, by.x="probeIDs", by.y=0, all=T);dim(output.Tf.up.mtr10) output.Tf.up.mtr10.2 <- cbind(output.Tf.up.mtr10, top.Tf.up.mtr10) ;dim(output.Tf.up.mtr10.2) write.table(output.Tf.up.mtr10.2, file="annot_limma_P0.05_Tf.up.mtr10_vs_WT_usesmall dataset.txt", sep="\t",quote=FALSE) ### Generate mtr10 annotation table with Total.fed down-regulated ###genes top.Tf.down.mtr10.id <- as.vector(top.Tf.down.mtr10$ID) ;length(top.Tf.down.mtr10.id) symbols <- as.vector(unlist(mget(top.Tf.down.mtr10.id, yeast2GENENAME))) ORF <- as.vector(unlist(mget(top.Tf.down.mtr10.id, yeast2ORF))) des <- as.vector(unlist(mget(top.Tf.down.mtr10.id, yeast2DESCRIPTION))) sym.orf.des <- as.vector(paste(symbols, ORF, des, sep=".")) top.Tf.down.df.mtr10 <- data.frame(probeIDs=top.Tf.down.mtr10.id, symbols, ORF, des, sym.orf.des);dim(top.Tf.down.df.mtr10) 319 Tf.down.mtr10.2 <data.frame(exprs(pm.mtr10.and.wt.exprs)[top.Tf.down.mtr10.id, ]) output.Tf.down.mtr10 <- merge(top.Tf.down.df.mtr10 ,Tf.down.mtr10.2, by.x="probeIDs", by.y=0, all=T);dim(output.Tf.down.mtr10) output.Tf.down.mtr10.2 <- cbind(output.Tf.down.mtr10, top.Tf.down.mtr10) ;dim(output.Tf.down.mtr10.2) write.table(output.Tf.down.mtr10.2, file="annot_limma_P0.05_Tf.down.mtr10_vs_WT_usesmall dataset.txt", sep="\t",quote=FALSE) ### Generate mtr10 annotation table with Total.starved up-regulated ###genes top.Ts.up.mtr10.id <- as.vector(top.Ts.up.mtr10$ID) ;length(top.Ts.up.mtr10.id) symbols <- as.vector(unlist(mget(top.Ts.up.mtr10.id, yeast2GENENAME))) ORF <- as.vector(unlist(mget(top.Ts.up.mtr10.id, yeast2ORF))) des <- as.vector(unlist(mget(top.Ts.up.mtr10.id, yeast2DESCRIPTION))) sym.orf.des <- as.vector(paste(symbols, ORF, des, sep=".")) top.Ts.up.df.mtr10 <- data.frame(probeIDs=top.Ts.up.mtr10.id, symbols, ORF, des, sym.orf.des);dim(top.Ts.up.df.mtr10) Ts.up.mtr10.2 <data.frame(exprs(pm.mtr10.and.wt.exprs)[top.Ts.up.mtr10.id, ]) output.Ts.up.mtr10 <- merge(top.Ts.up.df.mtr10 ,Ts.up.mtr10.2, by.x="probeIDs", by.y=0, all=T);dim(output.Ts.up.mtr10) output.Ts.up.mtr10.2 <- cbind(output.Ts.up.mtr10, top.Ts.up.mtr10) ;dim(output.Ts.up.mtr10.2) write.table(output.Ts.up.mtr10.2, file="annot_limma_P0.05_Ts.up.mtr10_vs_WT_usesmall dataset.txt", sep="\t",quote=FALSE) ### Generate mtr10 annotation table with Total.starved down-regulated ###genes top.Ts.down.mtr10.id <- as.vector(top.Ts.down.mtr10$ID) ;length(top.Ts.down.mtr10.id) symbols <- as.vector(unlist(mget(top.Ts.down.mtr10.id, yeast2GENENAME))) ORF <- as.vector(unlist(mget(top.Ts.down.mtr10.id, yeast2ORF))) des <- as.vector(unlist(mget(top.Ts.down.mtr10.id, yeast2DESCRIPTION))) sym.orf.des <- as.vector(paste(symbols, ORF, des, sep=".")) top.Ts.down.df.mtr10 <- data.frame(probeIDs=top.Ts.down.mtr10.id, symbols, ORF, des, sym.orf.des);dim(top.Ts.down.df.mtr10) Ts.down.mtr10.2 <data.frame(exprs(pm.mtr10.and.wt.exprs)[top.Ts.down.mtr10.id, ]) output.Ts.down.mtr10 <- merge(top.Ts.down.df.mtr10 ,Ts.down.mtr10.2, by.x="probeIDs", by.y=0, all=T);dim(output.Ts.down.mtr10) output.Ts.down.mtr10.2 <- cbind(output.Ts.down.mtr10, top.Ts.down.mtr10) ;dim(output.Ts.down.mtr10.2) write.table(output.Ts.down.mtr10.2, file="annot_limma_P0.05_Ts.down.mtr10_vs_WT_usesmall dataset.txt", sep="\t",quote=FALSE) 320 ## Txn values ## msn5 dim(txn.msn5.and.wt.exprs) ### Generate msn5 annotation table with Total.fed up-regulated genes top.Tf.up.msn5.id <- as.vector(top.Tf.up.msn5$ID) ;length(top.Tf.up.msn5.id) symbols <- as.vector(unlist(mget(top.Tf.up.msn5.id, yeast2GENENAME))) ORF <- as.vector(unlist(mget(top.Tf.up.msn5.id, yeast2ORF))) des <- as.vector(unlist(mget(top.Tf.up.msn5.id, yeast2DESCRIPTION))) sym.orf.des <- as.vector(paste(symbols, ORF, des, sep=".")) top.Tf.up.df.msn5 <- data.frame(probeIDs=top.Tf.up.msn5.id, symbols, ORF, des, sym.orf.des);dim(top.Tf.up.df.msn5) Tf.up.msn5.2 <data.frame(exprs(pm.msn5.and.wt.exprs)[top.Tf.up.msn5.id, ]) output.Tf.up.msn5 <- merge(top.Tf.up.df.msn5 ,Tf.up.msn5.2, by.x="probeIDs", by.y=0, all=T);dim(output.Tf.up.msn5) output.Tf.up.msn5.2 <- cbind(output.Tf.up.msn5, top.Tf.up.msn5) ;dim(output.Tf.up.msn5.2) write.table(output.Tf.up.msn5.2, file="annot_limma_P0.05_Tf.up.msn5_vs_WT_usesmall dataset.txt", sep="\t",quote=FALSE) ### Generate msn5 annotation table with Total.fed down-regulated genes top.Tf.down.msn5.id <- as.vector(top.Tf.down.msn5$ID) ;length(top.Tf.down.msn5.id) symbols <- as.vector(unlist(mget(top.Tf.down.msn5.id, yeast2GENENAME))) ORF <- as.vector(unlist(mget(top.Tf.down.msn5.id, yeast2ORF))) des <- as.vector(unlist(mget(top.Tf.down.msn5.id, yeast2DESCRIPTION))) sym.orf.des <- as.vector(paste(symbols, ORF, des, sep=".")) top.Tf.down.df.msn5 <- data.frame(probeIDs=top.Tf.down.msn5.id, symbols, ORF, des, sym.orf.des);dim(top.Tf.down.df.msn5) Tf.down.msn5.2 <data.frame(exprs(pm.msn5.and.wt.exprs)[top.Tf.down.msn5.id, ]) output.Tf.down.msn5 <- merge(top.Tf.down.df.msn5 ,Tf.down.msn5.2, by.x="probeIDs", by.y=0, all=T);dim(output.Tf.down.msn5) output.Tf.down.msn5.2 <- cbind(output.Tf.down.msn5, top.Tf.down.msn5) ;dim(output.Tf.down.msn5.2) write.table(output.Tf.down.msn5.2, file="annot_limma_P0.05_Tf.down.msn5_vs_WT_usesmall dataset.txt", sep="\t",quote=FALSE) ### Generate msn5 annotation table with Total.starved up-regulated ###genes top.Ts.up.msn5.id <- as.vector(top.Ts.up.msn5$ID) ;length(top.Ts.up.msn5.id) symbols <- as.vector(unlist(mget(top.Ts.up.msn5.id, yeast2GENENAME))) ORF <- as.vector(unlist(mget(top.Ts.up.msn5.id, yeast2ORF))) 321 des <- as.vector(unlist(mget(top.Ts.up.msn5.id, yeast2DESCRIPTION))) sym.orf.des <- as.vector(paste(symbols, ORF, des, sep=".")) top.Ts.up.df.msn5 <- data.frame(probeIDs=top.Ts.up.msn5.id, symbols, ORF, des, sym.orf.des);dim(top.Ts.up.df.msn5) Ts.up.msn5.2 <data.frame(exprs(pm.msn5.and.wt.exprs)[top.Ts.up.msn5.id, ]) output.Ts.up.msn5 <- merge(top.Ts.up.df.msn5 ,Ts.up.msn5.2, by.x="probeIDs", by.y=0, all=T);dim(output.Ts.up.msn5) output.Ts.up.msn5.2 <- cbind(output.Ts.up.msn5, top.Ts.up.msn5) ;dim(output.Ts.up.msn5.2) write.table(output.Ts.up.msn5.2, file="annot_limma_P0.05_Ts.up.msn5_vs_WT_usesmall dataset.txt", sep="\t",quote=FALSE) ### Generate msn5 annotation table with Total.starved down-regulated ###genes top.Ts.down.msn5.id <- as.vector(top.Ts.down.msn5$ID) ;length(top.Ts.down.msn5.id) symbols <- as.vector(unlist(mget(top.Ts.down.msn5.id, yeast2GENENAME))) ORF <- as.vector(unlist(mget(top.Ts.down.msn5.id, yeast2ORF))) des <- as.vector(unlist(mget(top.Ts.down.msn5.id, yeast2DESCRIPTION))) sym.orf.des <- as.vector(paste(symbols, ORF, des, sep=".")) top.Ts.down.df.msn5 <- data.frame(probeIDs=top.Ts.down.msn5.id, symbols, ORF, des, sym.orf.des);dim(top.Ts.down.df.msn5) Ts.down.msn5.2 <data.frame(exprs(pm.msn5.and.wt.exprs)[top.Ts.down.msn5.id, ]) output.Ts.down.msn5 <- merge(top.Ts.down.df.msn5 ,Ts.down.msn5.2, by.x="probeIDs", by.y=0, all=T);dim(output.Ts.down.msn5) output.Ts.down.msn5.2 <- cbind(output.Ts.down.msn5, top.Ts.down.msn5) ;dim(output.Ts.down.msn5.2) write.table(output.Ts.down.msn5.2, file="annot_limma_P0.05_Ts.down.msn5_vs_WT_usesmall dataset.txt", sep="\t",quote=FALSE) ##### Step 3 ##### Generate graphs of DEGs ##### ### ### Volcano plot ### ## Step 3.1 ## average PM values wt.pm.f <- exprs(esetsub[, 1:2])- exprs(esetsub[, 3:4]) ; dim(wt.pm.f) wt.pm.f.avg <- rowMeans(wt.pm.f) wt.pm.s <- exprs(esetsub[, 5:6])- exprs(esetsub[, 7:8]) ; dim(wt.pm.s) wt.pm.s.avg <- rowMeans(wt.pm.s) wt.pm.sf <- wt.pm.s - wt.pm.f ; dim(wt.pm.sf) 322 wt.pm.sf.avg <- rowMeans(wt.pm.sf) msn5.pm.f <- exprs(esetsub[, 9:10]) - exprs(esetsub[, 11:12]) msn5.pm.f.avg <- rowMeans(msn5.pm.f) msn5.pm.s <- exprs(esetsub[, 13:14])- exprs(esetsub[, 15:16]) msn5.pm.s.avg <- rowMeans(msn5.pm.s) msn5.pm.sf <- msn5.pm.s - msn5.pm.f msn5.pm.sf.avg <- rowMeans(msn5.pm.sf) mtr10.pm.f <- exprs(esetsub[, 17:18]) - exprs(esetsub[, 19:20]) mtr10.pm.f.avg <- rowMeans(mtr10.pm.f) mtr10.pm.s <- exprs(esetsub[, 21:22])- exprs(esetsub[, 23:24]) mtr10.pm.s.avg <- rowMeans(mtr10.pm.s) mtr10.pm.sf <- mtr10.pm.s - mtr10.pm.f mtr10.pm.sf.avg <- rowMeans(mtr10.pm.sf) ## mtr10 mtr10.pm.f.dif <- mtr10.pm.f.avg-wt.pm.f.avg; length(mtr10.pm.f.dif) mtr10.pm.s.dif <- mtr10.pm.s.avg-wt.pm.s.avg; length(mtr10.pm.s.dif) logp.mtr10.pm.f <- -log10(fit3.mtr10$p.value[,1]) adj.p.val logp.mtr10.pm.s <- -log10(fit3.mtr10$p.value[,2]) ## msn5 msn5.pm.f.dif msn5.pm.s.dif logp.msn5.pm.f logp.msn5.pm.s <<- <- msn5.pm.f.avg-wt.pm.f.avg <- msn5.pm.s.avg-wt.pm.s.avg -log10(fit3.msn5$p.value[,1]) -log10(fit3.msn5$p.value[,2]) ### NOT ### NOT adj.p.val ## PT values ## mtr10 mtr10.pt.f.dif <- mtr10.pt.f.avg-wt.pt.f.avg; length(mtr10.pt.f.dif) mtr10.pt.s.dif <- mtr10.pt.s.avg-wt.pt.s.avg; length(mtr10.pt.s.dif) logp.mtr10.pt.f <- -log10(pt.fit3$p.value[,1]) ### NOT adj.p.val logp.mtr10.pt.s <- -log10(pt.fit3$p.value[,2]) ## Txn values ## mtr10 mtr10.Tf.dif mtr10.Ts.dif logp.mtr10.Tf adj.p.val logp.mtr10.Ts ## msn5 msn5.Tf.dif msn5.Ts.dif <- mtr10.t.f.avg - wt.t.f.avg <- mtr10.t.s.avg - wt.t.s.avg <- -log10(fit3.txn.mtr10$p.value[,1]) <- -log10(fit3.txn.mtr10$p.value[,2]) <- msn5.t.f.avg - wt.t.f.avg <- msn5.t.s.avg - wt.t.s.avg 323 ### NOT logp.msn5.Tf adj.p.val logp.msn5.Ts <- -log10(fit3.txn.msn5$p.value[,1]) <- -log10(fit3.txn.msn5$p.value[,2]) ### NOT ## Step 3.2 ## Making plots pdf("Volcano plot of limma_PM values_mtr10 vs wt.pdf") plot(mtr10.pm.f.dif, -log10(fit3.mtr10$p.value[,1]), xlab="diff(logFC)",ylab="-log10 p value", cex=0.5, col="#A0A0A0", main = "Volcano plot of limma -- Pf/Mf_mtr10 vs wt") points(mtr10.pm.f.dif[top.pm.f.down.mtr10$ID], logp.mtr10.pm.f[top.pm.f.down.mtr10$ID], pch = 20, col= "blue3") points(mtr10.pm.f.dif[top.pm.f.up.mtr10$ID], logp.mtr10.pm.f[top.pm.f.up.mtr10$ID], pch = 20, col= "red3") abline(h =2.7, col="darkolivegreen4", lty=2) ## h value is smallest of -log10(p.value) abline(v = 0.6, col="darkolivegreen4", lty=2) abline(v = -0.6, col="darkolivegreen4", lty=2) plot(mtr10.pm.s.dif, -log10(fit3.mtr10$p.value[,2]), xlab="diff(logFC)",ylab="-log10 p value", cex=0.5, col="#A0A0A0", main = "Volcano plot of limma -- Ps/Ms_mtr10 vs wt") points(mtr10.pm.s.dif[top.pm.s.down.mtr10$ID], logp.mtr10.pm.s[top.pm.s.down.mtr10$ID], pch = 20, col= "blue3") points(mtr10.pm.s.dif[top.pm.s.up.mtr10$ID], logp.mtr10.pm.s[top.pm.s.up.mtr10$ID], pch = 20, col= "red3") abline(h =4.2, col="darkolivegreen4", lty=2) abline(v = 0.6, col="darkolivegreen4", lty=2) abline(v = -0.6, col="darkolivegreen4", lty=2);dev.off() pdf("Volcano plot of limma_PT values_mtr10 vs wt.pdf") plot(mtr10.pt.f.dif, -log10(pt.fit3$p.value[,1]), xlab="diff(logFC)",ylab="-log10 p value", cex=0.5, col="#A0A0A0", main = "Volcano plot of limma -- Pf/Tf_mtr10 vs wt") points(mtr10.pt.f.dif[top.pt.f.mtr10.down$ID], logp.pt.f[top.pt.f.mtr10.down$ID], pch = 20, col= "blue3") points(mtr10.pt.f.dif[top.pt.f.mtr10.up$ID], logp.pt.f[top.pt.f.mtr10.up$ID], pch = 20, col= "red3") abline(v = 0.6, col="darkolivegreen4", lty=2) abline(v = -0.6, col="darkolivegreen4", lty=2) plot(mtr10.pt.s.dif, -log10(pt.fit3$p.value[,2]), xlab="diff(logFC)",ylab="-log10 p value", cex=0.5, col="#A0A0A0", main = "Volcano plot of limma -- Pf/Tf_mtr10 vs wt") points(mtr10.pt.s.dif[top.pt.s.mtr10.down$ID], logp.pt.s[top.pt.s.mtr10.down$ID], pch = 20, col= "blue3") points(mtr10.pt.s.dif[top.pt.s.mtr10.up$ID], logp.pt.s[top.pt.s.mtr10.up$ID], pch = 20, col= "red3") abline(h = 3.1, col="darkolivegreen4", lty=2) abline(v = 0.6, col="darkolivegreen4", lty=2) abline(v = -0.6, col="darkolivegreen4", lty=2);dev.off() 324 ## plots of Txn values pdf("Volcano plot of limma_Txn values_mtr10 vs wt.pdf") plot(mtr10.Tf.dif, -log10(fit3.txn.mtr10$p.value[,1]), xlab="diff(logFC)",ylab="-log10 p value", cex=0.5, col="#A0A0A0", main = "Volcano plot of limma -- Tf_mtr10 vs wt") points(mtr10.Tf.dif[top.Tf.down.mtr10 $ID], logp.mtr10.Tf[top.Tf.down.mtr10 $ID], pch = 20, col= "blue3") points(mtr10.Tf.dif[top.Tf.up.mtr10 $ID], logp.mtr10.Tf[top.Tf.up.mtr10$ID], pch = 20, col= "red3") abline(h = 4, col="darkolivegreen4", lty=2) abline(v = 0.6, col="darkolivegreen4", lty=2) abline(v = -0.6, col="darkolivegreen4", lty=2) plot(mtr10.Ts.dif, -log10(fit3.txn.mtr10$p.value[,2]), xlab="diff(logFC)",ylab="-log10 p value", cex=0.5, col="#A0A0A0", main = "Volcano plot of limma -- Ts_mtr10 vs wt") points(mtr10.Ts.dif[top.Ts.down.mtr10 $ID], logp.mtr10.Ts[top.Ts.down.mtr10 $ID], pch = 20, col= "blue3") points(mtr10.Ts.dif[top.Ts.up.mtr10 $ID], logp.mtr10.Ts[top.Ts.up.mtr10$ID], pch = 20, col= "red3") abline(h = 3.63, col="darkolivegreen4", lty=2) abline(v = 0.6, col="darkolivegreen4", lty=2) abline(v = -0.6, col="darkolivegreen4", lty=2);dev.off() pdf("Volcano plot of limma_Txn values_msn5 vs wt.pdf") plot(msn5.Tf.dif, -log10(fit3.txn.msn5$p.value[,1]), xlab="diff(logFC)",ylab="-log10 p value", cex=0.5, col="#A0A0A0", main = "Volcano plot of limma -- Tf_msn5 vs wt") points(msn5.Tf.dif[top.Tf.down.msn5 $ID], logp.msn5.Tf[top.Tf.down.msn5 $ID], pch = 20, col= "blue3") points(msn5.Tf.dif[top.Tf.up.msn5 $ID], logp.msn5.Tf[top.Tf.up.msn5$ID], pch = 20, col= "red3") abline(h = 2.1, col="darkolivegreen4", lty=2) abline(v = 0.6, col="darkolivegreen4", lty=2) abline(v = -0.6, col="darkolivegreen4", lty=2) plot(msn5.Ts.dif, -log10(fit3.txn.msn5$p.value[,2]), xlab="diff(logFC)",ylab="-log10 p value", cex=0.5, col="#A0A0A0", main = "Volcano plot of limma -- Ts_msn5 vs wt") points(msn5.Ts.dif[top.Ts.down.msn5 $ID], logp.msn5.Ts[top.Ts.down.msn5 $ID], pch = 20, col= "blue3") points(msn5.Ts.dif[top.Ts.up.msn5 $ID], logp.msn5.Ts[top.Ts.up.msn5$ID], pch = 20, col= "red3") abline(h = 1.95, col="darkolivegreen4", lty=2) abline(v = 0.6, col="darkolivegreen4", lty=2) abline(v = -0.6, 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