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BNFO 615 Data Analysis in Bioinformatics Instructor Zhi Wei Outline Cell Genome Gene mRNA Proteins Systems biology Outline Cell Genome Gene mRNA Proteins Systems biology Cells Fundamental working units of every living system. Every organism is composed of one of two radically different types of cells: prokaryotic cells or eukaryotic cells. Prokaryotes and Eukaryotes are descended from the same primitive cell. All extant prokaryotic and eukaryotic cells are the result of a total of 3.5 billion years of evolution. Prokaryotes v.s. Eukaryotes Different Structures Different Components Different biological processes Prokaryotes vs Eukaryotes Prokaryotes Eukaryotes Single cell Single or multi cell No nucleus Nucleus No organelles Organelles One piece of circular DNA Chromosomes No mRNA post transcriptional modification Exons/Introns splicing Prokaryotes v.s. Eukaryotes Prokaryotes Eukaryotes bacteria, archaea Ecoli cell plants, animals, protista, and fungi Yeast cell 5X106 base pairs > 90% of DNA encode protein 5400 genes Lacks a membrane-bound nucleus. Circular DNA Histones are unknown 12.4x106 base pairs A small fraction of the total DNA encodes protein. Many repeats of non-coding sequences 5800 genes All chromosomes are contained in a membrane bound nucleus DNA is divided between 16 chromosomes A set of five histones: DNA packaging and gene expression regulation Cells chemical composition Chemical composition-by weight 70% water 7% small molecules salts Lipids amino acids nucleotides 23% macromolecules Proteins Polysaccharides lipids We have different cells Cells differ in size, shape and weights Q: what is the biggest cell in the human body? Cell Cycle Born, eat, replicate, and die Lodish et al. Molecular Biology of the Cell (5th ed.). W.H. Freeman & Co., 2003. Sexual Reproduction v.s. Cell Division Cell Division: Cells reproduce by duplicating their contents and dividing in two. Sexual Reproduction Formation of new individual by a combination of two haploid sex cells (gametes). Gametes for fertilization usually come from separate parents Both gametes are haploid, with a single set of chromosomes. The new individual is called a zygote, with two sets of chromosomes (diploid). Meiosis is a process to convert a diploid cell to a haploid gamete, and cause a change in the genetic information to increase diversity in the offspring. Meiosis v.s. Mitotic cell division Outline Cell Genome Gene mRNA Proteins Systems biology Genome A genome is an organism’s complete set of DNA (including its genes). However, in humans less than 3% of the genome actually encodes for genes. A part of the rest of the genome serves as a control regions (though that’s also a small part) The function of the rest of the genome is unknown (junk DNA? An open question). Comparison of Different Organisms Genome size (bp) Num. of genes E. Coli .05*108 5,400 Yeast .12*108 5,800 Worm .15*108 18,400 Fly 1.8*108 13,600 Human 30*108 25,000 Plant 1.3*108 25,000 Outline Cell Genome Gene mRNA Proteins Systems biology What is a gene? Promoter Protein coding sequence Terminator Genomic DNA DNA: Deoxyribo Nucleic Acid Example of a Gene: Gal4 DNA ATGAAGCTACTGTCTTCTATCGAACAAGCATGCGATATTTGCCGACTTAAAAAGCTCAAG TGCTCCAAAGAAAAACCGAAGTGCGCCAAGTGTCTGAAGAACAACTGGGAGTGTCGCTAC TCTCCCAAAACCAAAAGGTCTCCGCTGACTAGGGCACATCTGACAGAAGTGGAATCAAGG CTAGAAAGACTGGAACAGCTATTTCTACTGATTTTTCCTCGAGAAGACCTTGACATGATT TTGAAAATGGATTCTTTACAGGATATAAAAGCATTGTTAACAGGATTATTTGTACAAGAT AATGTGAATAAAGATGCCGTCACAGATAGATTGGCTTCAGTGGAGACTGATATGCCTCTA ACATTGAGACAGCATAGAATAAGTGCGACATCATCATCGGAAGAGAGTAGTAACAAAGGT CAAAGACAGTTGACTGTATCGATTGACTCGGCAGCTCATCATGATAACTCCACAATTCCG TTGGATTTTATGCCCAGGGATGCTCTTCATGGATTTGATTGGTCTGAAGAGGATGACATG TCGGATGGCTTGCCCTTCCTGAAAACGGACCCCAACAATAATGGGTTCTTTGGCGACGGT TCTCTCTTATGTATTCTTCGATCTATTGGCTTTAAACCGGAAAATTACACGAACTCTAAC GTTAACAGGCTCCCGACCATGATTACGGATAGATACACGTTGGCTTCTAGATCCACAACA TCCCGTTTACTTCAAAGTTATCTCAATAATTTTCACCCCTACTGCCCTATCGTGCACTCA CCGACGCTAATGATGTTGTATAATAACCAGATTGAAATCGCGTCGAAGGATCAATGGCAA ATCCTTTTTAACTGCATATTAGCCATTGGAGCCTGGTGTATAGAGGGGGAATCTACTGAT ATAGATGTTTTTTACTATCAAAATGCTAAATCTCATTTGACGAGCAAGGTCTTCGAGTCA A sequence of A,C,G,T Example of a Gene: Gal4 AA MKLLSSIEQACDICRLKKLKCSKEKPKCAKCLKNNWECRYSPKTKRSPLTRAHLTEVESR LERLEQLFLLIFPREDLDMILKMDSLQDIKALLTGLFVQDNVNKDAVTDRLASVETDMPL TLRQHRISATSSSEESSNKGQRQLTVSIDSAAHHDNSTIPLDFMPRDALHGFDWSEEDDM SDGLPFLKTDPNNNGFFGDGSLLCILRSIGFKPENYTNSNVNRLPTMITDRYTLASRSTT SRLLQSYLNNFHPYCPIVHSPTLMMLYNNQIEIASKDQWQILFNCILAIGAWCIEGESTD IDVFYYQNAKSHLTSKVFESGSIILVTALHLLSRYTQWRQKTNTSYNFHSFSIRMAISLG LNRDLPSSFSDSSILEQRRRIWWSVYSWEIQLSLLYGRSIQLSQNTISFPSSVDDVQRTT TGPTIYHGIIETARLLQVFTKIYELDKTVTAEKSPICAKKCLMICNEIEEVSRQAPKFLQ MDISTTALTNLLKEHPWLSFTRFELKWKQLSLIIYVLRDFFTNFTQKKSQLEQDQNDHQS YEVKRCSIMLSDAAQRTVMSVSSYMDNHNVTPYFAWNCSYYLFNAVLVPIKTLLSNSKSN AENNETAQLLQQINTVLMLLKKLATFKIQTCEKYIQVLEEVCAPFLLSQCAIPLPHISYN NSNGSAIKNIVGSATIAQYPTLPEENVNNISVKYVSPGSVGPSPVPLKSGASFSDLVKLL SNRPPSRNSPVTIPRSTPSHRSVTPFLGQQQQLQSLVPLTPSALFGGANFNQSGNIADSS A sequence of 20 amino acids {A,R,N,D,C,E,Q,G,H,I,L,K,M,F,P,S,T,W,Y,V} The Central Dogma DNA RNA: Gene Transcription promoter 5’ 3’ G A T T A C A . . . C T A A T G T . . . 3’ 5’ Gene Transcription transcription factor, binding site, RNA polymerase 5’ 3’ G A T T A C A . . . C T A A T G T . . . 3’ 5’ Transcription factors recognize transcription factor binding sites and bind to them, forming a complex. RNA polymerase binds the complex. Gene Transcription 5’ 3’ 3’ 5’ The two strands are separated Gene Transcription 5’ 3’ 3’ 5’ An RNA copy of the 5’→3’ sequence is created from the 3’→5’ template Gene Transcription G A T T A C A . . . 5’ 3’ 3’ 5’ C T A A T G T . . . pre-mRNA 5’ G A U U A C A . . . 3’ RNA Processing (Eukaryotes) 5’ cap, polyadenylation, exon, intron, splicing, UTR 5’ cap poly(A) tail exon intron mRNA 5’ UTR 3’ UTR Mammalian Gene Structure introns 5’ 3’ coding promoter 5’ UTR exons 3’ UTR non-coding Regulatory regions: up to 50 kb upstream of +1 site Exons: protein coding and untranslated regions (UTR) 1 to 178 exons per gene (mean 8.8) 8 bp to 17 kb per exon (mean 145 bp) Introns: splice acceptor (GU) and donor (AG) sites, junk DNA average 1 kb – 50 kb per intron Gene size: Largest – 2.4 Mb (Dystrophin). Mean – 27 kb. Only 1.5% DNA for coding in Human! Identifying Genes in Sequence Data Predicting the start and end of genes as well as the introns and exons in each gene is one of the basic problems in computational biology. Gene prediction methods look for ORFs (Open Reading Frame). These are (relatively long) DNA segments that start with the start codon, end with one of the end codons, and do not contain any other end codon in between. Splice site prediction has received a lot of attention in the literature. Comparative genomics Outline Cell Genome Gene mRNA Proteins Systems biology RNA RNA is similar to DNA chemically. It is usually only a single strand. T(hyamine) is replaced by U(racil) Some forms of RNA can form secondary structures by “pairing up” with itself. This can have change its properties dramatically. DNA and RNA can pair with each other. tRNA linear and 3D view: http://www.cgl.ucsf.edu/home/glasfeld/tutorial/trna/trna.gif RNA, continued Several types exist, classified by function mRNA – this is what is usually being referred to when a Bioinformatician says “RNA”. This is used to carry a gene’s message out of the nucleus. tRNA – transfers genetic information from mRNA to an amino acid sequence rRNA – ribosomal RNA. Part of the ribosome which is involved in translation. Messenger RNA Basically, an intermediate product Transcribed from the genome and translated into protein Number of copies correlates well with number of proteins for the gene. Unlike DNA, the amount of messenger RNA (as well as the number of proteins) differs between different cell types and under different conditions. Complementary base-pairing mRNA is transcribed from the DNA mRNA (like DNA, but unlike proteins) binds to its complement Quantify mRNA levels Outline Cell Genome Gene mRNA Proteins Systems biology Proteins: Workhorses of the Cell Proteins are polypeptide chains of amino acids. 20 different amino acids different chemical properties cause the protein chains to fold up into specific three-dimensional structures that define their particular functions in the cell. Proteins do all essential work for the cell build cellular structures digest nutrients execute metabolic functions Mediate information flow within a cell and among cellular communities. Genes Make Proteins genome-> genes ->protein(forms cellular structural & life functional)->pathways & physiology Genes Encode for Proteins Second letter U C A G UUU Phenylalanine (Phe) UCU Serine (Ser) UAU Tyrosine (Tyr) UGU Cysteine (Cys) U UUC Phe UCC Ser UAC Tyr UGC Cys C UUA Leucine (Leu) UCA Ser UAA STOP UGA STOP A UUG Leu UCG Ser UAG STOP UGG Tryptophan (Trp) G CUU Leucine (Leu) CCU Proline (Pro) CAU Histidine (His) CGU Arginine (Arg) U CUC Leu CCC Pro CAC His CGC Arg C CUA Leu CCA Pro CAA Glutamine (Gln) CGA Arg A CUG Leu CCG Pro CAG Gln CGG Arg G AUU Isoleucine (Ile) ACU Threonine (Thr) AAU Asparagine (Asn) AGU Serine (Ser) U AUC Ile ACC Thr AAC Asn AGC Ser C AUA Ile ACA Thr AAA Lysine (Lys) AGA Arginine (Arg) A AUG Methionine (Met) or START ACG Thr AAG Lys AGG Arg G GUU Valine (Val) GCU Alanine (Ala) GAU Aspartic acid (Asp) GGU Glycine (Gly) U GUC Val GCC Ala GAC Asp GGC Gly C GUA Val GCA Ala GAA Glutamic acid (Glu) GGA Gly A GUG Val GCG Ala GAG Glu GGG Gly G C A G Triplet one Amino Acid 4^3 combinations mapped to 20 Amino Acids Third letter First letter U Open Reading Frames G C U U G U U U A C G A A U U A G G C U U G U U U A C G A A U U A G G C U U G U U U A C G A A U U A G G C U U G U U U A C G A A U U A G Synonymous Mutation G G C U U G U U U A C G A A U U A G G C U U G U U U A C G A A U U A G Ala Cys Leu Arg Ile G C U U G U U U G C G A A U U A G Ala Cys Leu Arg Ile Missense Mutation G G C U U G U U U A C G A A U U A G G C U U G U U U A C G A A U U A G Ala Cys Leu Arg Ile G C U U G G U U A C G A A U U A G Ala Trp Leu Arg Ile Nonsense Mutation A G C U U G U U U A C G A A U U A G G C U U G U U U A C G A A U U A G Ala Cys Leu Arg Ile G C U U G A U U A C G A A U U A G Ala STOP Frameshift G C U U G U U U A C G A A U U A G G C U U G U U U A C G A A U U A G Ala Cys Leu Arg Ile G C U U G U U A C G A A U U A G Ala Tyr Cys Glu Leu Protein Structure Proteins work together with other proteins or nucleic acids as "molecular machines" structures fit together and function in highly specific, lock-and-key ways. Four levels of structure: Primary Structure: The sequence of the protein Secondary structure: Local structure in regions of the chain. (alpha helix, beta sheet) Tertiary Structure: Three dimensional structure Quaternary Structure: multiple subunits Assigning Function to Proteins While 25000 genes have been identified in the human genome, relatively few have known functional annotation. Determining the function of the protein can be done in several ways. Sequence similarity to other (known) proteins Using domain information Using three dimensional structure Based on high throughput experiments (when does it functions and who it interacts with) Summary: DNA(Gene) RNA Protein Replication Transcription Translation Outline Cell Genome Gene mRNA Proteins Systems biology Biological pathway/gene networks Instead of having brains, cells make decision through complex networks of interactions, called pathways Synthesize new materials Break other materials down for spare parts Signal to eat or die In order to fulfill their function, proteins interact with other proteins in a number of ways including: Regulation Signaling Pathways, for example A -> B -> C Post translational modifications Forming protein complexes An Example Systems Biology We now have many sources of data, each providing a different view on the activity in the cell Sequence (genes) DNA motifs Gene expression Protein interactions Protein-DNA interaction Etc. Putting it all together: Systems Biology Next week Introduction to R programming You need to do in-class exercises Acknowledgments Ziv Bar-Joseph: for some of the slides adapted or modified from his lecture slides at Carnegie Mellon University Neil Jones: for some of the slides adapted or modified from his slides for the book An Introduction to Bioinformatics Algorithms