Philippe CollasUniversity of Oslo

Institute of Basic Medical SciencesStem Cell Epigenetics Laboratory

Norwegian Center for Stem Cell Research

An epigenetic approach to understanding(and predicting?) environmental effects on

gene expression

www.collaslab.com

Untangled hair(DNA strands)

Head of Leda, ~yr 1500

Braids(nucleosomal arrays)

Coiled braids(folded chromatin)

Had Leonardo da Vinci figured it out?

High-order folding of the genome

DNA compaction in a eukaryotic cell

A high-order folding of the genome

DNA compaction in a eukaryotic cell

Epigenetic marks lie on top of the DNA code

DNA methylation

Histone modifications

Genome conformationcapture–seq (HiC)

3-D genome conformation also influences gene expression

Sequencing Chromosome interaction maps

Genome conformationcapture–seq (HiC)

3-D genome conformation also influences gene expression

SequencingIntegration of multiple information layers

3-D genome conformation also influencesgene expression – a dynamic system

Stem /progenitor cell

Epigenetic changes determine which genes are active or inactive during development

Identical genes Variable gene expression

patterns

Different sets of genes are active in different cell types

The environment can affect future generations through the epigenome

Mother:1st generation

Fetus:2nd generation

Germ cells:3rd generation

Direct exposure: up to 3 generations

Mother:1st generation

Fetus:2nd generation

Germ cells:3rd generation

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Transmission of epigenetic marks through the germline

The environment can affect future generations through the epigenome

4th generation?

Patterning developmental gene expression by epigenetic marks

Leif LindemanIngrid Andersen

• 3 h of development until zygotic genome activation (ZGA)• Essentially no transcription during this period• Great opportunity to test the hypothesis of an epigeneticpre-patterning of developmental gene expression

How does the zebrafish embryo programs itsdevelopmental gene expression pattern?

ZGA

Capturing epigenetic modifications during earlyzebrafish development

xPre-MBT2.5 hpf

MBT3.5 hpf

Post-MBT5.3 hpf

Lindeman et al., 2010. PLoS ONELindeman et al., 2010. Meth Mol BiolLindeman, Andersen et al., 2011. Dev CellAndersen et al., 2012. Genome Biol.

Chromatinimmunoprecipitation:

Immuno-affinity enrichment ofchromatin domains bearing a

specific modification

Marking of the genome by histone modificationsbefore, during and after onset of ZGA

RNA-seq(Aanes et al., 2011. Genome Res.)

Non-expressed genes

Maternal-zygotic genes Zygotic genes

Capturing epigenetic modifications during earlyzebrafish development

xPre-MBT2.5 hpf

MBT3.5 hpf

Post-MBT5.3 hpf

Lindeman, Andersen et al., 2011. Dev Cell

Homeostasis

Transcription regulationDevelopment

Development Transcription regulationSignaling(Metabolism)

Chromatin assembly

Gene ontologyPre‐ZGA

TSS

H3K4me3

H3K9me3

H3K27me3

SignalingSensory perception

The zebrafish genome is marked by modified histonesbefore onset of zygotic genome activation

Pre‐ZGA zebrafish embryos

Density profiles

H3K4me3H3K9me3

H3K27me3

TSS

Epigenetic fate map of genes marked by H3K4me3 pre-ZGA

Epigenetic fate map of genes marked by H3K4me3 pre-ZGA:Acquisition of bi- and trivalency

Pre‐MBT

H3K27me3H3K9me3H3K4me3

MBT

Post‐MBT

Pre‐MBT

Pre‐MBT H3K4me3‐marked genes are not expressed

H3K4me3-marked genes before ZGA are inactive... suggesting a predictive role of H3K4me3 marking

Enhanced propensity for expression Enhanced expression level post-MBT

H3K4me3 marking before ZGA correlates with transcriptionalactivation after ZGA onset

... suggesting an instructive role of epigenetic marking on (later) gene activation

Homeostasis

Transcription regulationDevelopment

Development Transcription regulationSignaling(Metabolism)

Chromatin assembly

Gene ontologyPre‐ZGA

TSS

H3K4me3

H3K9me3

H3K27me3

SignalingSensory perception

The zebrafish genome is marked by modified histonesbefore onset of zygotic genome activation

Pre‐ZGA zebrafish embryos

Human ES cells

Mikkelsen et al., 2007

Mouse embryos:H3K4/H3K27me3 bivalency

Dahl et al., 2010

Gene ontologyPre‐ZGA

Development Transcription regulationSignaling(Metabolism)

Common epigenetic pattern of ’transcriptional poising’ in zebrafish, mouse and man

Pre‐ZGA zebrafish embryos

Epigenetic patterning of zebrafish developmental gene expression before ZGA onset

Pre‐MBT MBT Post‐MBT

H3K4me3

H3K27me3H3K9me3

H3K36me3RNAPII

TSS

Pre‐patterning Priming Activation

Hypomethylation

Our data currently support two models:

• transgenerational inheritance of chromatin marks?

• post-fertilization removal & reconstitution of epigenetic marks based on DNA sequence rather than through a copy of histone modifications – inherent instructive property of DNA?

Both models are compatible with a de novo epigenetic marking of genes post-fertilization

?Where do chromatin modifications come from?

Predictive modeling of gene expression from chromatin states

Goal: to correlate epigenetic profiles at specific developmental stages to patterns of gene expression across stages

• Characterize epigenetic modifications throughout the genome• Find interrelations among the marks by assigning ‘chromatin states’• Do this for each developmental stage• Correlate developmental expression patterns with epigenetic patterns across the

states• Find epigenetic pattern changes which predict expression fate across

developmental stages• Find epigenetic patterns that are predictive of biological process (Gene Ontology)

and anatomically-specific (Zfin) expression

Atlas of zebrafish developmental epigenetics with prediction value

Apply to other species or fields

Developing innovative tools to look at things in another way

Andrew Reiner

Implications for CERAD

What you’re going to do to an adult organism (bearing gametes) or an embryo is going to affect, through changes in the epigenome, gene expression

patterns, phenotype, behavior, health, etc.

These epigenetic changes may have a predictive value on gene expression later in life

Need: a robust assay to survey epigenetic changes in early (fish) embryos

Fish’n ChIPs protocol – Dr. Leif Lindeman