PRC2 Promotes Canalisation During Endodermal Differentiation [ChIP-Seq]

In this study we employed a mouse embryonic stem cell (ESCs) differentiation system to convert pluripotent ESCs into Anterior Definitive Endoderm (ADE) to model primitive streak formation and early gastrulation. To monitor the progression of the differentiation we utilised a dual reporter mESC line (B6) bearing fluorescent reporters knocked in to the Gsc (marker of the PS/mesendoderm tagged with GFP) and Hhex (marker of definitive endoderm lineages tagged with Redstar) loci. To monitor changes in the chromatin landscape during this differentiation we performed ChIP-seq for H3K27me3 and H3K4me3 (modifications associated with gene repression and activation respectively) in each of nine FACs sorted ADE populations. Using this approach, in combination with genome-wide gene expression analysis, we; i). Found that changes in gene expression/transcription were both predictive of and predicted by dynamic changes in the levels of these histone modifications at gene promoters; ii). Found that changes in H3K27me3 levels at TSS are more dynamic than the levels of H3K4me3; iii). Determined that transcriptional upregulation of genes did not require the prior loss of H3K27me3 at gene TSSs. ChIP-seq was performed on nine ADE populations isolated by FACS sorting based on GSC-GFP and HHEX-RedStar expression levels. Two or three independent experiments were pooled and sequenced for each ChIP and input sample.