Epigenetics Slides by GC Yuan 3/28/12

One genome, Many cell-types The genome is constant across all cell types. Tissue specific gene expression cannot be explained by the DNA sequence alone. Epigenetics provides an addition layer of regulatory control. errraticwisdom.com

Epigenetics Epigenetics refers to changes in phenotype (appearance) or gene expression caused by mechanisms OTHER THAN changes in the underlying DNA sequence. In the most stringent sense, epigenetics only refers to heritable changes --- those can be passed on to daughter cells (somatic) or next generation (germline). In common usage of the term, epigenetics does not have to be heritable.

Epigenetic mechanisms Nucleosome positions Histone modification DNA methylation

Chromatin DNA is packaged into chromatin. Nucleosome is the fundamental unit of chromatin. It wraps 146 bp DNA. Higher-order organization is still poorly understood. Felsenfeld and Groudine 2003

1bp (0.3nm) 10,000 nm 30nm 11 nm DNA in the nucleus is complexed with histones to form nucleosomes

Richard Wheeler Complete nucleosome with DNA Individual histone proteins

Histone modification The amino acids on the N-terminus of histones can be covalently modified at multiple locations in multiple ways. Felsenfeld and Groudine 2003

Histonations??? Acetylation Methylation (me1, me2, me3) Phosphorylation Ubiquitination Sumoylation Citrulination ADP-ribosylation

Histone variants The core histone proteins have a number of variants. H2A.Z (early embryonic development?) H2A.X (maybe DSB repair???) H3.3 (???) CENP-A (centromeres) …

Mapping histones by ChIP-seq Park 2009, Nature Rev Gen Extract protein- bound DNA by chromatin immunoprecipitation Detect the nucleotide sequence of the ChIP-DNA by next generation sequencing technology.

Euchromatin and heterochromatin Euchromatin Heterochromatin Heterochromatin is enriched near centromeres and telomeres.

The action’s in the middle – sequence signatures are used to regulate nucleosome placement Segal 2006

The action’s in the middle – sequence signatures are used to regulate nucleosome placement Segal 2006

DNA methylation Alberts et al. Molecular Biology of the Cell DNA methylation normally occurs only at CpG dinucleotides and can be inherited during cell-division.

More “track” data, typically % methylation Weber 2005 Lister/Ecker 2011 Rauch 2007

Genome-wide DNA methylation detection Bisulfite sequencing –Convert unmethylated C to U (RNA version of T). –Sequence the original and converted sequences and look for differences. Methylated DNA immunoprecipitation (MeDIP) –Similar to ChIP, but use antibodies against methyl-C instead, followed by microarray or sequencing. Endonuclease digestion –Digest by methylation-sensitive, sequence-specific enzymes followed by sequencing.

Epigenetic Regulation of Transcription

Promoter DNA methylation is negatively correlated with gene expression Lister et al. 2009, Nature Abs. mC % Rel. mC % CpG CpNG CpN

DNA methylation pattern in genomic regions (of ES cells) is unique Lister et al. 2009

Genome-wide nucleosome positions Yeast Human Genome-wide nucleosome positions can be identified by MNase based tiling array or DNA sequencing methods Most promoters contain nucleosome depleted regions before TSS. Lee et al. 2007Schones et al. 2008

Transcription factor binding sites (TFBSs) are likely to be nucleosome-depleted TFBSs tend to be nucleosome-depleted. Motif sites that are unbound in our condition but bound in other conditions also tend to be nucleosome depleted. Motif sites that are always unbound do not have nucleosome- depletion property. Yuan et al. 2005

Transcriptional regulation by nucleosome and histone modification Nucleosome positioning is mainly repressive Histone modification can be either active or repressive TF TF target site TF Ace TF H3K27me3 H3K9ac TF

Correlation between gene expression and different histone modification patterns Barski et al. 2007

Histone code hypothesis “… multiple histone modifications, acting in a combinatorial or sequential fashion on one or multiple histone tails, specify unique downstream functions …” ― Strahl and Allis, Nature, (2000)

Role of Epigenetics in Development

Sparmann and Lohuizen Nature Reviews Cancer 6, 846–856 (November 2006) | doi: /nrc1991 Polycomb family proteins silence developmental genes (DNA methyltransferases)

Polycomb binding represses gene expression me Ac Ezh2 Suz12 Eed Jmj PRC2 PRC1 Bmi1 HPCRing1 HPH Polycomb complexes repress gene expression through H3K27me3.

Polycomb targets developmental genes in ES Boyer et al. 2006, Cell Polycomb Oct4 Nanog Sox2 expressed repressed Kim et al. 2008, Cell Enrichment in Polycomb ChIP-bound gene functions

Mikkelsen et al Bivalent domains in ES cells ES cells contain many bivalent domains: H3K4me3 (active mark) and H3K27me3 (repressive mark) These genes are “poised for activation.” High/intermediate/low CpG content

Lineage-specific epigenetic patterns Epigenetic patterns change during cell differentiation. activation repression Housekeeping Stem Neural progenitor Fibroblasts Neural TF Neurogenesis Adipogenesis Neural progenitor Brain/lung TF, multiple TSSs Mikkelsen et al. 2007

Epigenetics and Diseases Cancer

General DNA methylation signature of cancer (Many) CpG islands are (usually) unmethylated in normal cells. In cancer, some of the CpG islands are hypermethylated, repressing some tumor repressor genes. But the overall level of DNA methylation is typically decreased.

The CpG Island Methylator Phenotype (CIMP) Toyota M et al. PNAS 1999;96: Jean-Pierre Issa. Nature Reviews Cancer 4,

Methylator subtype of brain cancer Laird 2010

Methylator subtype is associated with survival Laird 2010

Over-expression of Ezh2 in advanced prostate cancer Varambelly et al Polycomb-group gene silencer by H3K27m3 Oncogenes/ metastasis promoters

Why does this matter?

Epigenetic reprogramming can modify tumorigenesis Hochedlinger et al. Genes & Dev, (2004) In SCID mice Partially tumor-derived Control Genetics are tough to modify; epigenetics aren’t (as much). 1.Gives us a way to “cure cancer” and 2.Links cancer to developmental biology!

An ESC signature of high-grade breast cell cancers Ben-Porath et al “Worse” tumors → More “stemmy” More epigenetic suppression

ES-signature is negatively associated with survival Ben-Porath et al Stemness is good when you’re an embryo, bad when you’re a tumor

Maybe

New debate of ESC signature of cancer “ESC” signature associated with cancer can be explained by Myc activity ESC signature contains three modules: core, Polycomb, and Myc. The ESC signature associated with cancer can be explained by Myc signature. Kim et al. 2010

But in all fairness, epigenetics matter for more than just cancer… Lung Cardiovascular Diabetes Ageing Post Traumatic Disease Social Behavior …

Epigenetics and Environment

Epigenetic changes are reversible Epigenetic changes can be induced by environmental changes. Unlike genetic mutations, epigenetic changes are reversible. Ideal for therapeutic targets, IF we know how epigenetic patterns are regulated. Jirtle and Skinner 2007

Baccarelli 2010 Diet Environment Immune function Environment

Baccarelli 2010

Nutrition affects skin color Genetically identical Skin color is controlled by DNA methylation Mother mouse was fed with methyl-enriched diet during pregnancy Jirtle and Skinner 2007

NIH Epigenome Roadmap Five-year initiative starting from 2008 It will generate reference epigenome for hundreds of cell-types in human The data will be made publicly available immediately. New technologies will be developed to detect novel epigenomic marks Population-level studies will be carried out to detect epigenetic changes associated with various diseases. This is an exciting time to do research in epigenomics!

To learn more... Bernstein et al. “The mammalian epigenome” Cell Feb 23;128(4): Jones and Baylin, “The epigenomics of cancer”, Cell Feb 23;128(4): Jirtle and Skinner, “Environmental epigenomics and disease susceptibility.”, Nat Rev Genet Apr;8(4): NIH Epigenome Roadmap – NIH ENCODE Project –