Integrating 5-Hydroxymethylcytosine into the Epigenomic Landscape of Human Embryonic Stem Cells

Abstract

Covalent modification of DNA distinguishes cellular identities and is crucial for regulating the pluripotency and differentiation of embryonic stem (ES) cells. The recent demonstration that 5-methylcytosine (5-mC) may be further modified to 5-hydroxymethylcytosine (5-hmC) in ES cells has revealed a novel regulatory paradigm to modulate the epigenetic landscape of pluripotency. To understand the role of 5-hmC in the epigenomic landscape of pluripotent cells, here we profile the genome-wide 5-hmC distribution and correlate it with the genomic profiles of 11 diverse histone modifications and six transcription factors in human ES cells. By integrating genomic 5-hmC signals with maps of histone enrichment, we link particular pluripotency-associated chromatin contexts with 5-hmC. Intriguingly, through additional correlations with defined chromatin signatures at promoter and enhancer subtypes, we show distinct enrichment of 5-hmC at enhancers marked with H3K4me1 and H3K27ac. These results suggest potential role(s) for 5-hmC in the regulation of specific promoters and enhancers. In addition, our results provide a detailed epigenomic map of 5-hmC from which to pursue future functional studies on the diverse regulatory roles associated with 5-hmC.

Segmenting the human genome based on states of neutral genetic divergence

“…Whereas telomere-specific aberrant repair might elevate substitutions directly (29), the mutagenicity of recombination has been debated (2, 31). GC richnessmeasured at 45% vs. 39.8% genome median-agrees well with abundance of genes and functional elements in the hot state, as do abundance of open chromatin (32), enrichment for 5-hydroxy methylcytosines (33), and early replication timing (34) (Fig. 2B and Table 2; see also below).…”

Real-time dynamics of methyl-CpG-binding domain protein 3 and its role in DNA demethylation by fluorescence correlation spectroscopy

“…Immunostaining of 5hmC was done per the methods in the literature with suitable modification. 78 Generally, cells after different treatments were fixed with 4% paraformaldehyde for 15 min at 4 °C and washed 3 times with 1× PBS buffer (Gibco, 10010). Cell membrane was permeabilized by 0.4% Triton X-100 (Sigma, T8787) for 30 min and washed with PBS for 15 min.…”

Dynamics of 5-hydroxymethylcytosine during mouse spermatogenesis

“…Indeed, two recent studies showed that two methyl CpG binding proteins, MeCP2 and Mbd3, preferentially bind to 5hmC and may have important roles in transcription regulation 20,21 . Previous studies indicate that 5hmC is enriched in the exons of highly expressed genes in brain tissues, and in cis-regulatory elements such as active enhancers, insulatorbinding sites in ESCs [22][23][24][25][26][27][28] . Enrichment of 5hmC is also found in the promoter region of polycomb-repressed 'poised' genes in ESCs [23][24][25]27 , which disappeared in the adult brain 29 .…”

“…Previous studies indicate that 5hmC is enriched in the exons of highly expressed genes in brain tissues, and in cis-regulatory elements such as active enhancers, insulatorbinding sites in ESCs [22][23][24][25][26][27][28] . Enrichment of 5hmC is also found in the promoter region of polycomb-repressed 'poised' genes in ESCs [23][24][25]27 , which disappeared in the adult brain 29 . By comparing the genomic distribution of 5hmC in mouse hippocampus and cerebellum at three different ages, we have shown that it exhibits dynamic changes during brain development and aging 29 .…”