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High-resolution enzymatic mapping of genomic 5-hydroxymethylcytosine in mouse embryonic stem cells

ABSTRACT: We describe the use of a novel DNA modification-dependent restriction endonuclease AbaSI coupled with sequencing (Aba-seq) to map high-resolution hydroxymethylome of mouse E14 embryonic stem cells. The specificity of AbaSI enables sensitive detection of 5hmC at low occupancy regions. Bioinformatic analysis suggests 5hmCs in genic regions closely follows the 5mC distribution. 5hmC is generally depleted in CpG islands and only enriched in a small set of repetitive elements. A regularly spaced and oscillating 5hmC pattern was observed at the binding sites of CTCF. 5hmC is enriched at the poised enhancers with the mono-methylated histone H3 lysine 4 (H3K4me1) marks, but not at the active enhancers with the acetylated histone H3 lysine 27 (H3K27Ac) marks. Non-CG hydroxymethylation appears to be prevalent in the mitochondrial genome. We propose that some amounts of transiently stable 5hmCs may indicate a poised epigenetic state or de-methylation intermediate, while others may suggest a locally accessible chromosomal environment to the TET enzymatic apparatus.

ORGANISM(S): Mus musculus

PROVIDER: GSE42898 | GEO | 2013/01/01

SECONDARY ACCESSION(S): PRJNA183732

REPOSITORIES: GEO

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Project description:Cell fate transitions involve integration of genomic information encoded by regulatory elements, such as enhancers, with the cellular environment. However, identification of the genomic sequences that control the earliest steps of human embryonic development represents a formidable challenge. Here we show that in human embryonic stem cells (hESCs) unique chromatin signatures identify two distinct classes of genomic elements, both of which are marked by the presence of chromatin regulators p300 and BRG1, and monomethylation of histone H3 at lysine 4 (H3K4me1). In addition, the elements of the first class are distinguished by the acetylation of histone H3 at lysine 27 (H3K27ac), overlap with previously characterized enhancers active in hESCs, and are generally located proximally to genes expressed in hESCs and in the epiblast. In contrast, the elements within the second class, which we termed M-bM-^@M-^\poised enhancersM-bM-^@M-^], are distinguished by the absence of H3K27ac, enrichment of histone H3 lysine 27 trimethylation (H3K27me3) and are linked to genes inactive in hESCs and involved in orchestrating early steps in mammalian development, such as gastrulation, mesoderm formation and neurulation. Consistent with the poised identity, during differentiation of hESCs to neuroepithelium, a neuroectoderm-specific subset of these elements acquires a chromatin signature associated with active enhancers. Remarkably, when assayed in zebrafish embryos, human poised enhancer elements are able to direct cell type and stage specific expression patterns characteristic of their proximal developmental gene, even in the absence of sequence conservation in the fish genome. Our data demonstrate that enhancers are epigenetically pre-marked and suggest a heretofore unappreciated role of H3K27me3 at distal regulatory elements. Moreover, the unique chromatin signature associated with poised enhancers allowed us to uncover over 2,000 putative developmental regulatory sequences, thereby creating an invaluable resource for future studies and isolation of transient, rare cell populations representing early steps of human development. For GSM602289-90: RNA-seq experiments in human ESC and neuroectodermal (NEC) speheres For GSM602291-303: Genome-wide analysis of p300, H3K4me3, H3K4me1, H3K27me3 and H3K27ac in human embryonic stem cells (ESC) and neuroectoderm cells (NEC). Additionally, in H9 ESC ChIP-seq were alsoe obtained for BRG1 and FAIRE-seq was also performed.

Project description:Cell fate transitions involve integration of genomic information encoded by regulatory elements, such as enhancers, with the cellular environment. However, identification of the genomic sequences that control the earliest steps of human embryonic development represents a formidable challenge. Here we show that in human embryonic stem cells (hESCs) unique chromatin signatures identify two distinct classes of genomic elements, both of which are marked by the presence of chromatin regulators p300 and BRG1, and monomethylation of histone H3 at lysine 4 (H3K4me1). In addition, the elements of the first class are distinguished by the acetylation of histone H3 at lysine 27 (H3K27ac), overlap with previously characterized enhancers active in hESCs, and are generally located proximally to genes expressed in hESCs and in the epiblast. In contrast, the elements within the second class, which we termed “poised enhancers”, are distinguished by the absence of H3K27ac, enrichment of histone H3 lysine 27 trimethylation (H3K27me3) and are linked to genes inactive in hESCs and involved in orchestrating early steps in mammalian development, such as gastrulation, mesoderm formation and neurulation. Consistent with the poised identity, during differentiation of hESCs to neuroepithelium, a neuroectoderm-specific subset of these elements acquires a chromatin signature associated with active enhancers. Remarkably, when assayed in zebrafish embryos, human poised enhancer elements are able to direct cell type and stage specific expression patterns characteristic of their proximal developmental gene, even in the absence of sequence conservation in the fish genome. Our data demonstrate that enhancers are epigenetically pre-marked and suggest a heretofore unappreciated role of H3K27me3 at distal regulatory elements. Moreover, the unique chromatin signature associated with poised enhancers allowed us to uncover over 2,000 putative developmental regulatory sequences, thereby creating an invaluable resource for future studies and isolation of transient, rare cell populations representing early steps of human development.

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High-resolution enzymatic mapping of genomic 5-hydroxymethylcytosine in mouse embryonic stem cells

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