Genome-wide mapping of 5-hydroxymethylcytosine in embryonic stem cells

By William A. Pastor, U. J. Pape, Yun Huang, H. Henderson, Ryan Lister, M. Ko, E. Mcloughlin, Y. Brudno, Sahasransu Mahapatra, Philipp Kapranov and 7 more. Nature, 2011

796 citations recorded by the source corpus.

5-hydroxymethylcytosine (5hmC) is a modified base present at low levels in diverse cell types in mammals. 5hmC is generated by the TET family of Fe(II) and 2-oxoglutarate-dependent enzymes through oxidation of 5-methylcytosine (5mC). 5hmC and TET proteins have been implicated in stem cell biology and cancer, but information on the genome-wide distribution of 5hmC is limited. Here we describe two novel and specific approaches to profile the genomic localization of 5hmC. The first approach, termed GLIB (glucosylation, periodate oxidation, biotinylation) uses a combination of enzymatic and chemical steps to isolate DNA fragments containing as few as a single 5hmC. The second approach involves conversion of 5hmC to cytosine 5-methylenesulphonate (CMS) by treatment of genomic DNA with sodium bisulphite, followed by immunoprecipitation of CMS-containing DNA with a specific antiserum to CMS. High-throughput sequencing of 5hmC-containing DNA from mouse embryonic stem (ES) cells showed strong enrichment within exons and near transcriptional start sites. 5hmC was especially enriched at the start sites of genes whose promoters bear dual histone 3 lysine 27 trimethylation (H3K27me3) and histone 3 lysine 4 trimethylation (H3K4me3) marks. Our results indicate that 5hmC has a probable role in transcriptional regulation, and suggest a model in which 5hmC contributes to the ‘poised’ chromatin signature found at developmentally-regulated genes in ES cells.

Subfields: Genetics, Molecular Biology

View Paper: https://www.semanticscholar.org/paper/d27849e7cba3f214f1389825846cde7e7d0ba842

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In Genetics

1. Yi Zhang
2. Hao Wu
3. E. Nestler
4. Chuan He
5. K. Helin
6. Peng Jin
7. Qing Dai
8. Chun-Xiao Song

In Molecular Biology

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4. K. Helin
5. J. Wysocka
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Citations in the corpus

• DNA Demethylation Dynamics (2011, methodology): “Thus, contrary to expectations, the 5mC to 5hmC modification is clearly not the functional equivalent of 5mC to cytosine, which is associated with derepression of certain gene promoters (Pastor et al., 2011).”
• Development and maintenance of regulatory T cells. (2013, background): “Genomic DNA is mainly methylated by DNA methyltransferases (Dnmt family members), whereas it can be demethylated bymultiple steps, includingmethylcytosine hydroxylation mediated by TET family members (Bhutani et al., 2011; Pastor et al., 2011).”
• DNA Methylation and Its Basic Function (2013, background): “…DNA can be enriched using antibodies that bind specifically to 5hmC or by biotinylation of modified 5hmC and precipitated sequences can be identified using microarray chips or by DNA sequencing (Szwagierczak et al, 2010; Ficz et al, 2011; Jin et al, 2011; Pastor et al, 2011; Wu and Zhang, 2011).”
• Role of TET enzymes in DNA methylation, development, and cancer (2016, background): “There has been substantial interest inmapping cytosine oxidation products in the genome with the aim of understanding their role in gene regulation and DNA methylation dynamics (Pastor et al. 2011; Stroud et al. 2011; Szulwach et al. 2011; Williams et al. 2011; Wu et al. 2011a; Xu et al. 2011c; Séra”
• Modification of enhancer chromatin: what, how and why? (2013, background): “…proteins have been reported to interact with cell-type-specific and signaling-dependent TFs, including Pax proteins, nuclear receptors, and beta-catenin, suggesting a TF-mediated recruitment mechanism (Kawabe et al., 2012; Lee et al., 2006; Mo et al., 2006; Patel et al., 2007; Sierra et al., 2006).”