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Saturday, February 27, 2010

AGBT 2010 - Bing Ren - UCSD

Epigenomic Landscapes of Pluripotent and Lineage-Committed Human Cells

Sequencing of the human genome has led to
* identification of disease causing genes
* Personalized medicine
* advanced sequencing technologies
* Foundation for understanding the construction of human beings

But DNA is only half the story
* variations in DNA alone not account for all variations in phenotypic traits
* organisms with identical DNA often exhibit distinct phenotypes (eg plants, insects, mammals)
* Epigenetic changes contribute to human diseases, phenotypes, etc

We know about the mechanisms
* DNA is wrapped around histone proteins which can be modified
* DNA is itself modified (methylation)

[paraphrased] DNA is hardware, epigenome is the software (Duke university quote... missed author's name)

* very complex
* varies among different cell types
* generally reprogrammed during the life cycle of tan organism
* Epigenome is also affected by environmental clues

How do we ecipher the "epigentic code"?
* sytematic approach
* large scale profindg of chromatin modification
* finding common modifications
* validation

* ChIP-Seq based. (started with Tiling arrays)
* use antibodies that recognize chromatin modification.

[beautiful pictures]
* Chromatin signature for the promoter and gene body
* H3K4me3 marks active promoters
* H3K36me3 marks gene body of active genes
* Signature has led to identification of thousands of long non-coding RNA genes.

Chromatin signatures of enhancers
* Can use information about modifications to model patterns
* predict enhancers in the human genome.
* 36,589 enhancer predictions were made
* 56% found in intergenic regions
* test a few with reporter assays - show that 80% of predicted enhancers do drive reporter genes. (Far fewer of the control sequences do - missed number)

Finding chromatin modification patterns in the genome de novo
(Hon et al, PLoS Comp Bio 2009)
* 16 different patterns of chromosome modification
* some are enhancers,
* others have no associations
* one has pattern highly enriched for exons.. regulates alt splicing.

* chromatin modification patterns could be used to annotate ...
* Epigenome Roadmap project (Generate reference epigenome maps for a large number of primary human cells and tissues)

Datasets are available at GEO. (NCBI)

Mapping of DNA methyltion and 53 histone modifications in human cells
* Human embryonic stem cells (H1)
* Fetal fibroblast cell line

Method for mapping DNA methylation
* Ryan Lister and Joe Ecker (Salk)
* sodium bisulfite (C to U), if not methylated
* Must do deep sequencing. If using HiSeq - could do it in 10 days. Used to take 20 runs
* Methylation status for more than 94% of cytosines determined.
* 75.5% in H1, 99.98% in Fibroblast
* DNA methylation is depletee from functional sequences
* no-CpG methlyation is enriched in gene body of transcribed genes suggesting link to the transcription process

11 chromatin modification marks
* comparing cells: different results
* K9me3 and K27me3 become dramatically extended (7% in ES to more than 30% in fibroblast.)
* genes with above marks are highly enriched in developmental genes.

Reduction of repressive chromatins in induced pluripotent cells

Repressive chromatin domains occupy small fraction of genome which is maintained as open structure in stem cells

Repressive chromatin domains occupy large fraction of genome, keeping genes involved in development silenced in differentiated cells.

* widespread difference in epigenomes of ES and fibroblasts
* stem cells are characterized by abundant non-CpG methylation
* Expansion of repressive domains may be a key characteristic of cellular differentiation
* [Missed 2]



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