Download introns within ribosomal protein genes regulate the production and

INTRONS WITHIN RIBOSOMAL PROTEIN GENES REGULATE THE PRODUCTION AND FUNCTION OF YEAST RIBOSOMES •  Dplicaed ribosomal protein genes are assymetrically expressed and regulated. •  Introns within ribosomal protein genes have an influence on gene expression. •  Introns influence cell survival under stress. •  Introns of ribosomal protein genes modulate the processing of pre-­‐mRNA. •  How introns regulate the funcOon of yeast ribosomes INTRODUCTION Introns are the non coding sequences of a gene that are removed from pre-­‐mRNA during alternaOve splicing during transcripOon.In buddying yeast the pre-­‐mRNA coding for the ribosomal proteins are the most spliced.The impact of these introns on growth,cell fitness,stress,pre-­‐rRNA processing and gene expression has been studied bz eliminaOng(deleOon)of the introns from the ribosomal proteins and determining their effect. NB:most of the ribosomal protein gene are duplicated bakers yeast organism.proposed that the duplicaOon permits adjusOng the dose of Rp to match that or rRna synthesis thereby ensuring opOmal ribosome assembly. IDENTIFICATION AND DELETION OF INTRONS ASSOCIATED WITH Rp genes •  Done in saccromyces cerevasie genome database ,the posiOons of introns within the Rp genes was idenOfied using the ares lab yeast intron database and MIPs yeast genome database. •  Intron distribuOon of Ribosomal protein genes is not subunit specific 70%(LSU) and 78%(SSU) Rp genes. •  Introns were found in 80% of the duplicated Rp genes but only 37% of the unique gene. •  To evaluate the fucOon of the introns deleOons were performed single,double deleOon and from both copies. •  None of the single or double deleOons had any effect on growth under standard labalatory condiOons. INTRONS FINE TUNE THE EXPRESSION OF RP GENES •  Impact of intron deleOon on the expression of the host gene was monitored using qPRC and was calculated relaOve to the expression of housekeeping gene SPT15. •  84% of all intron deleted strains changed the expression of the host genes. •  Gene expression was considered changed in an intron dependent intargenic regulatory manner.Intron dependent negaOve reglaOon was more prevelent in LSU proteins where as posiOve regulaOon was prevelant in SSU protein. Duplicated Rp Genes are asymetrically expressed and regulated •  Hypothesis:a change in the expression of any one single copy of the duplicated Rp gene pair is expected to be compensated by a change in the other copy. •  63% of all intron deleted strains in duplicated Rp genes that showed intron-­‐dependent intragenic regulaOon also affected the expression of the other copy in same or opposite way.however in 80% of these cases intragenic regulaOon was not compensetory in nature. Introns of Rp genes modulate the processing of pre-­‐rRNA •  Processing intermediated were detected using qPCR. •  About 88% of all intron deleOons increased or decreasd at least one pre-­‐rRna processing intermediate. •  MaturaOon of the primary precursor 35s that carry all rRna transcirpts and ends at the terminaOon site T2 was increased by 15%. •  The majority of the intron deleOons delaying the processing of 35s was foud in genes associated with the LSU where as those decreasing the precursor was found in SSU-­‐associated genes. Introns regulate cell fitness and drug response •  To determine the impact of introns on condiOonal growth a pilot study was done on strains carrying intron deleOons on essenOal RP genes . •  9 intron deleted strains were subjected to comprehensive funcOonal assays that include growth on 8 different carbon sources,16 different drugs and 3 different temperatures. •  The strongest effect was observed in the presence of the known apoptosis-­‐inducing drug straurosporine that inhibited 21% of all strains with bias in LSU Rp gene. •  Introns negaOvely and posiOvely regulate growth under stress and that the deleOons in essenOal genes had a stronger effect on growth in presence of drug compared to non essenOal genes. Introns were grown in compeOOon with wild Type cells to monitor cell fifness.17% of all Intron deleted strains decreased cell fifness And 25% of the deleOons enhanced cell fitness In rich media. Rp introns play an important role in ModulaOng the compeOOve advantage of Yeast cells. TAKE HOME MESSEGE •  The presence of introns does not necessarily inhibit gene expression as one might expect but can sOmulate gene expression. •  Introns are required for fine-­‐tuning the expression of duplicated Rp genes thereby offering means to exploit their funcOonal difference and to alter ribosome homeostasis. •  Introns are not essenOal for cell growth under labalatory condiOons however they influence cell survival under stress and compeOoOon for limited resources.