reChIP-seq reveals widespread bivalency of H3K4me3 and H3K27me3 in CD4+ memory T cells

Abstract

The combinatorial action of co-localizing chromatin modifications and regulators determines chromatin structure and function. However, identifying co-localizing chromatin features in a high-throughput manner remains a technical challenge. Here we describe a novel reChIP-seq approach and tailored bioinformatic analysis tool, normR that allows for the sequential enrichment and detection of co-localizing DNA-associated proteins in an unbiased and genome-wide manner. We illustrate the utility of the reChIP-seq method and normR by identifying H3K4me3 or H3K27me3 bivalently modified nucleosomes in primary human CD4+ memory T cells. We unravel widespread bivalency at hypomethylated CpG-islands coinciding with inactive promoters of developmental regulators. reChIP-seq additionally uncovered heterogeneous bivalency in the population, which was undetectable by intersecting H3K4me3 and H3K27me3 ChIP-seq tracks. Finally, we provide evidence that bivalency is established and stabilized by an interplay between the genome and epigenome. Our reChIP-seq approach augments conventional ChIP-seq and is broadly applicable to unravel combinatorial modes of action.

The International Human Epigenome Consortium: A Blueprint for Scientific Collaboration and Discovery

“…To address this challenge we have developed a series of molecular and computational approaches to deconvolute epigenomic signatures from heterogeneous populations. Three independent strategies are presented to explore the heterogeneity at bivalent domains, a "poised state" marking important developmental genes characterized by an active (histone H3 lysine 4 trimethylation, H3K4me3) and a repressive (H3K27me3) mark on the same histone, and reveal that this combinatory epigenetic signature is both lost and gained at key regulatory genes during development (Hirst 2016, Kinkley, Helmuth et al 2016, Weiner, Lara-Astiaso et al 2016. Further these methods define previously undescribed cooccurrence patterns of histone modifications on single nucleosomes and in relationship with enzyme accessibility of chromatin.…”

Dynamics of RNA Polymerase II Pausing and Bivalent Histone H3 Methylation during Neuronal Differentiation in Brain Development

“…Interestingly, genes containing overlapping H3K4me3 and H3K27me3 marks only express at a low level, which leads to the thought that such bivalent genes may be poised for activation or repression (or maintained in the bivalent silent state) during the progression of embryogenesis. In support of this idea, the bivalent H3K4me3 and H3K27me3 chromatin domains are found to be preferentially enriched at developmental regulatory genes in the mouse ESCs (Bernstein et al, 2006), germline cells (Sachs et al, 2013), pre-implantation embryos (Liu et al, 2016b) and several adult tissues (Cui et al, 2009; Cui et al, 2012; Hammoud et al, 2009; Kinkley et al, 2016; Mikkelsen et al, 2007). Therefore, bivalent histone H3 modification also presents a suitable regulatory mechanism for modulating gene expression levels during development.…”

Epigenomic Profiling of Human CD4+ T Cells Supports a Linear Differentiation Model and Highlights Molecular Regulators of Memory Development

“…For NOMe-seq, nuclei of fixed cells were extracted and DNA-meth on GpC motifs in accessible chromatin regions was introduced using the M. CviPI methyltransferase, followed by WGBS analysis. ChIP-seq for histone modifications was carried out as previously described (Arrigoni et al, 2016;Kinkley et al, 2016). RNA was extracted using the miRNeasy Micro Kit (QIAGEN) and three Illumina sequencing libraries were prepared (small RNA sequencing library, one stranded total RNA, and one stranded mRNA library).…”