Dual functions of Tet1 in transcriptional regulation in mouse embryonic stem cells. Nature

Department of Molecular & Medical Pharmacology, UCLA David Geffen School of Medicine, Los Angeles, California 90095, USA.
Nature (Impact Factor: 41.46). 03/2011; 473(7347):389-93. DOI: 10.1038/nature09934
Source: PubMed


Epigenetic modification of the mammalian genome by DNA methylation (5-methylcytosine) has a profound impact on chromatin structure, gene expression and maintenance of cellular identity. The recent demonstration that members of the Ten-eleven translocation (Tet) family of proteins can convert 5-methylcytosine to 5-hydroxymethylcytosine raised the possibility that Tet proteins are capable of establishing a distinct epigenetic state. We have recently demonstrated that Tet1 is specifically expressed in murine embryonic stem (ES) cells and is required for ES cell maintenance. Using chromatin immunoprecipitation coupled with high-throughput DNA sequencing, here we show in mouse ES cells that Tet1 is preferentially bound to CpG-rich sequences at promoters of both transcriptionally active and Polycomb-repressed genes. Despite an increase in levels of DNA methylation at many Tet1-binding sites, Tet1 depletion does not lead to downregulation of all the Tet1 targets. Interestingly, although Tet1-mediated promoter hypomethylation is required for maintaining the expression of a group of transcriptionally active genes, it is also involved in repression of Polycomb-targeted developmental regulators. Tet1 contributes to silencing of this group of genes by facilitating recruitment of PRC2 to CpG-rich gene promoters. Thus, our study not only establishes a role for Tet1 in modulating DNA methylation levels at CpG-rich promoters, but also reveals a dual function of Tet1 in promoting transcription of pluripotency factors as well as participating in the repression of Polycomb-targeted developmental regulators.

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Available from: Kairong Cui, Jun 02, 2015
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    • "5mC is necessary for the correct embryonic development (Bestor, 2000; Fouse et al., 2008; Li, 2002; Neri et al., 2013b; Reik and Walter, 2001), and its deregulation has been linked to cancer development and progression (Costello et al., 2000; Easwaran et al., 2012; Jones, 2002; Neri et al., 2014; Teng et al., 2011). On the other hand, active DNA demethylation is mediated by ten eleven translocation (Tet) proteins, which progressively oxidize 5mC to 5-hydroxy- methylcytosine (5hmC), 5-formylcytosine (5fC), and 5-carbox- ylcytosine (5caC), and by the thymine-DNA glycosidase (Tdg) protein that binds and excises 5fC and 5caC residues to allow the restoration of unmodified C by the base excision and repair machinery (Cortá zar et al., 2011; Cortellino et al., 2011; He et al., 2011; Ito et al., 2010, 2011; Maiti and Drohat, 2011; Neri et al., 2013a; Williams et al., 2011; Wu et al., 2011). Several methods, such as bisulfite sequencing (bisulfite-seq), Tet-assisted bisulfite sequencing (TAB-seq), and oxidative bisulfite sequencing, have been established to map 5mC and 5hmC residues at single-base resolution on a genome-wide scale (Booth et al., 2012; Meissner et al., 2008; Yu et al., 2012). "
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    ABSTRACT: Ten eleven translocation (Tet) proteins oxidize 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC), and 5-carboxylcytosine (5caC). 5fC and 5caC can be further excised by thymine-DNA glycosylase (Tdg). Here, we present a genome-wide approach, named methylation-assisted bisulfite sequencing (MAB-seq), that enables single-base resolution mapping of 5fC and 5caC and measures their abundance. Application of this method to mouse embryonic stem cells (ESCs) shows the occurrence of 5fC and 5caC residues on the hypomethylated promoters of highly expressed genes, which is increased upon Tdg silencing, revealing active DNA demethylation on these promoters. Genome-wide mapping of Tdg reveals extensive colocalization with Tet1 on active promoters. These regions were found to be methylated by Dnmt1 and Dnmt3a and demethylated by a Tet-dependent mechanism. Our work demonstrates the DNA methylation dynamics that occurs on the promoters of the expressed genes and provides a genomic reference map of 5fC and 5caC in ESCs. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.
    Cell Reports 02/2015; 10(5). DOI:10.1016/j.celrep.2015.01.008 · 8.36 Impact Factor
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    • "5hmC was particularly enriched at the start sites of " bivalent genes " , which bear dual histone 3 lysine 27 trimethylation (H3K27me3) and histone 3 lysine 4 trimethylation (H3K4me3) marks and are usually found at developmentallyregulated genes in ES cells [58]. Indeed TET1 has been shown to bind to these bivalent domains (repressed genes) as well as H3K4me3-only promoters (actively-transcribed genes), mediating both expression of pluripotency-associated genes and repression of Polycomb-targeted developmental regulators [82]. Although Tet1-null ES cells show reduced levels of 5hmC and subtle changes in gene expression, they are pluripotent and support development of live-born mice in tetraploid complementation assay but display skewed differentiation toward trophectoderm in vitro. "
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    ABSTRACT: Epigenetic modifications of the genome play important roles in controlling gene transcription thus regulating several molecular and cellular processes. A novel epigenetic modification -5-hydroxymethylcytosine (5hmC) – has been recently described and attracted a lot of attention due to its possible involvement in the active DNA demethylation mechanism. TET enzymes are dioxygenases capable of oxidizing the methyl group of 5-methylcytosines (5mC) and thus converting 5mC into 5hmC. Although most of the work on TET enzymes and 5hmC has been carried out in embryonic stem (ES) cells, the highest levels of 5hmC occur in the brain and in neurons, pointing to a role for this epigenetic modification in the control of neuronal differentiation, neural plasticity and brain functions. Here we review the most recent advances on the role of TET enzymes and DNA hydroxymethylation in neuronal differentiation and function.
    Genomics 11/2014; 104(5). DOI:10.1016/j.ygeno.2014.08.018 · 2.28 Impact Factor
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    • "Mapping 5fC/5caC accumulation in TDG-deficient cells at single-base resolution may contribute to a better understanding of Tet and TDGdependent active DNA demethylation. Previous studies have shown that Tet proteins are required for the maintenance of low methylation status at a subset of promoters in ESCs (Wu et al. 2011; Xu et al. 2011; Dawlaty et al. 2014). In this study, we not only provided a complete list of promoters undergoing active DNA demethylation but also found that enhancer regions are the most profound Tet-dependent active demethylation loci. "
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    ABSTRACT: DNA methylation at the C-5 position of cytosine (5mC) is one of the best-studied epigenetic modifications and plays important roles in diverse biological processes. Iterative oxidation of 5mC by the ten-eleven translocation (Tet) family of proteins generates 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC), and 5-carboxylcytosine (5caC). 5fC and 5caC are selectively recognized and excised by thymine DNA glycosylase (TDG), leading to DNA demethylation. Functional characterization of Tet proteins has been complicated by the redundancy between the three family members. Using CRISPR/Cas9 technology, we generated mouse embryonic stem cells (ESCs) deficient for all three Tet proteins (Tet triple knockout [TKO]). Whole-genome bisulfite sequencing (WGBS) analysis revealed that Tet-mediated DNA demethylation mainly occurs at distally located enhancers and fine-tunes the transcription of genes associated with these regions. Functional characterization of Tet TKO ESCs revealed a role for Tet proteins in regulating the two-cell embryo (2C)-like state under ESC culture conditions. In addition, Tet TKO ESCs exhibited increased telomere-sister chromatid exchange and elongated telomeres. Collectively, our study reveals a role for Tet proteins in not only DNA demethylation at enhancers but also regulating the 2C-like state and telomere homeostasis.
    Genes & Development 09/2014; 28(19). DOI:10.1101/gad.248005.114 · 10.80 Impact Factor
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