Integrative Genomics

Double-helix DNA strands are separated in the gene coding region Which enzyme detects the beginning of a gene ? RNA Polymerase (multi-subunit enzyme that synthesize RNA) binds to promoter RNA polymerase I – 28S, 5.8S and 18S rRNA genes RNA polymerase II – coding genes, snRNA RNA polymerase III – tRNA, 5S rRNA, snoRNA Other enzymes General (Basal) Transcription Factor (GTF) TFIIA, TFIIB, TFIID TFIID – recognize promoter sequence Transcription initiation

Transcription initiation in eukaryotes Initial contact is made by general transcription factor (GTF) TFIID, which consists of TATA-binding protein (TBP) and at least 12 TBP-associated factors (TAF)

Epigenomics Stephen Baylin & Peter Jones, A decade of exploring the cancer epigenome, 2011 Active gene promoters (CpG rich and lacking DNA meth) have nucleosome-depleted regions (NDRs) just upstream of TSS Nucleosomes flanking NDRs are marked by H3K4me3 and have extensive K ac and histone variant H2A.Z (destabilize nucleosomes to promote transcription) Gene body shows enrichment of specific covalent marks, including H3K36me3 (facilitating transcriptional elongation) Active gene has H3K4me1 and H3K27ac DNA meth silences in promoters that lack H2A.Z with repressive H3K9me2 or H3K9me3 Insulators such as CCCTC-binding factor (CTCF) organize DNA into repressive loops or active euchromatin

Integrative Genomics RD Hawkins, et al., “Next-generation genomics: an integrative approach,” Nature, 2010 Genomic Data Sets Available Sequence variation data from individual genomes Transcriptome Epigenomic data – methylation in MethDB Interactome – RNA-protein, protein-protein

Integrative Genomics Can address questions related to fundamental mechanisms of genome function and disease How might particular risk-associated SNPs (Single Nucleotide Polymorphism) affect cellular function, leading to a disease ? What functional sequences exist in human genome ? How are key development pathways regulated by epigenetic mechanisms ? Astrazeneca: sequence-2-million-genomes

Disease of premature aging Average life span – 12 yrs Mostly die of hardening of heart chamber Caused by a single neucleotide mutation SNP (Single Nucleotide Polymorphism)

Integrative Genomics Can answer What are the functional variants of gene-distal loci identified by associate studies ? Where are the regulatory elements ? To what extent does the activity of regulatory elements contribute to disease or to individual gene expression variation ? Became an essential part of experimental design

Result: Annotating functional features of genome Regulation elements are not fully understood Enhancers, insulators From characteristics of known regulatory elements, identify novel elements Chromatin signature of enhancers are used to find new enhancers

Result: Inferring function of genetic variants SNPs in non-coding regions are still poorly defined SNVs (single-nucleotide variants) in transcription factor binding sites or chromatin-marked regulatory elements may be used to determine regulatory SNPs SNP at Pol II bonding regions cause variability of gene expression levels

Result: Understanding Gene Regulation Correlating epigenomic info and transcriptome info: show combined action of insulators, enhancers, etc. Correlating histone modification data and transcriptome info: determine roles of histone modifications

Combinatorial Complexity Oliver Rando, Combinatorial complexity in chromatin structure and function, 2012 H4K5ac – 2-fold increase in transcription H4K8ac – 2-fold increase Does H4K5acK8ac produce 4-fold increase ?

Combinatorial Complexity Multivalent binding Arabidopsis CMT3 binds to H3K9me3K27me3, but not to either Chd1 binds to H3R2me2K4me3 than to H3K4me3 Binding of HP1 to H3K9me3 is inhibited by phosphorylation of adjacent H3S10

Combinatorial Complexity Genome-wide mapping studies cannot account for combinatorial complexity Studies of the effect of comb. Histone modifications to gene expressions find Different comb. of histone mutants exhibit relatively little phenotype complexity H3: Loss of H3K9 ac is little different from loss of H3K18 ac H4: 16 possible comb. Mutations among 4 K’s K5, K8, K12 have almost the same effect Variations of 41 chromatin marks have >56% captured using 3 PCA components