Genome-wide analysis of 5-hydroxymethylcytosine distribution reveals its dual function in transcriptional regulation in mouse embryonic stem cells
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE27613 | GEO | 2011/03/30
SECONDARY ACCESSION(S): PRJNA138263
REPOSITORIES: GEO
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Project description:Recent studies have demonstrated that the Ten-eleven translocation (Tet) family proteins can enzymatically convert 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC). While 5mC has been studied extensively, little is known about the distribution and function of 5hmC. Here we present a genome-wide profile of 5hmC in mouse embryonic stem (ES) cells. A combined analysis of global 5hmC distribution and gene expression profile in wild-type and Tet1-depleted ES cells revealed suggests that 5hmC is enriched at both gene bodies of actively transcribed genes and extended promoter regions of Polycomb-repressed developmental regulators. Thus, our study reveals the first genome-wide 5hmC distribution in pluripotent stem cells and supports its dual function in regulating gene expression. Genomic DNA extracted from control (Con) or Tet1 knockdown (KD) mouse ES cells was immunoprecipitated with 5-hydroxymethylcytosine (5hmC) antibodies and analyzed by NimbleGen 2.1M mouse whole genome tiling microarrays (a 4-array set covering the entired non-repetitive portion of mouse genome). Whole cell extract (WCE) was used as input controls in IP/input experiments.
2011-03-30 | E-GEOD-27613 | biostudies-arrayexpress
Project description:DNA methylation of C5-cytosine (5mC) in the mammalian genome is a key epigenetic event that is critical for various cellular processes. However, how the genome-wide 5mC pattern is dynamically regulated remains a fundamental question in epigenetic biology. The TET family of 5mC hydroxylases, which convert 5mC to 5-hydroxymethylcytosine (5hmC), have provided a new potential mechanism for the dynamic regulation of DNA methylation. The extent to which individual Tet family members contribute to the genome-wide 5mC and 5hmC patterns and associated gene network remains largely unknown. Here we report genome-wide mapping of Tet1 and 5hmC in mESCs and reveal a mechanism of action by which Tet1 controls 5hmC and 5mC levels in mESCs. In combination with microarray and mRNA-seq expression profiling, we identify a comprehensive yet intricate gene network influenced by Tet1. We propose a model whereby Tet1 controls DNA methylation both by binding to CpG-rich regions to prevent unwanted DNA methyltransferase activity, and by converting the existing 5mC to 5hmC through its enzymatic activity. This Tet1-mediated antagonism of CpG methylation imparts differential maintenance of DNA methylation status at Tet1 target loci, thereby providing a new regulatory mechanism for establishing the epigenetic landscape of mESCs, which ultimately contributes to mESC differentiation and the onset of embryonic development. To determine the genome-wide distribution of Tet1 and 5hmC in mouse ES cells, as well as identify the gene transcription changes after Tet1 depletion. GSM706669-GSM706671: We used GST pull-down followed by deep sequencing to map the DNA bound by the Tet1 CXXC domain in vitro. We made two mutants that have a single point mutation (Cys574 to Ala or Cys586 to Ala) in the core CXXC domain to ascertain the essential role of the CXXC domain in DNA binding by comparing the sequencing profile of DNA bound by wild type CXXC with the profiles of the CXXC mutants. GSM706672-GSM706673: Tet1 ChIP-seq was performed to identify the genome-wide distribution of Tet1 in mouse ES cells. GSM706674-GSM706679: We performed hydroxymethylated DNA immunoprecipitation (hMeDIP)-seq combined with a shRNA-mediated gene depletion strategy. To identify the loci specific 5hmC regulation by Tet1, we compared the 5hmC genome-wide distributions in control (Luc shRNA) and Tet1-depleted (Tet1 shRNA2863) mouse ES cells. GSM706680-GSM706682: To identify the gene regulation network by Tet1, we compared the gene expression profiles of control (scramble shRNA) and Tet1-depleted (Tet1 shRNA 2863 and Tet1 shRNA 3387) mouse ES cells determined by mRNA-seq.
2011-04-21 | E-GEOD-28532 | biostudies-arrayexpress
Project description:DNA methylation of C5-cytosine (5mC) in the mammalian genome is a key epigenetic event that is critical for various cellular processes. However, how the genome-wide 5mC pattern is dynamically regulated remains a fundamental question in epigenetic biology. The TET family of 5mC hydroxylases, which convert 5mC to 5-hydroxymethylcytosine (5hmC), have provided a new potential mechanism for the dynamic regulation of DNA methylation. The extent to which individual Tet family members contribute to the genome-wide 5mC and 5hmC patterns and associated gene network remains largely unknown. Here we report genome-wide mapping of Tet1 and 5hmC in mESCs and reveal a mechanism of action by which Tet1 controls 5hmC and 5mC levels in mESCs. In combination with microarray and mRNA-seq expression profiling, we identify a comprehensive yet intricate gene network influenced by Tet1. We propose a model whereby Tet1 controls DNA methylation both by binding to CpG-rich regions to prevent unwanted DNA methyltransferase activity, and by converting the existing 5mC to 5hmC through its enzymatic activity. This Tet1-mediated antagonism of CpG methylation imparts differential maintenance of DNA methylation status at Tet1 target loci, thereby providing a new regulatory mechanism for establishing the epigenetic landscape of mESCs, which ultimately contributes to mESC differentiation and the onset of embryonic development. To determine the genome-wide DNA methylation changes caused by Tet1 depletion in mouse ES cells. Tet1 protein was depleted by specific siRNA treatment. The DNA methylation levels in control and Tet1 siRNA-transfected ES cells were determined by targeted bisulfite sequencing.
2011-04-21 | E-GEOD-28533 | biostudies-arrayexpress
Project description:We performed a meta analysis of publicly available TET1, 5mC, 5hmC and genome wide bisulfite profiling data mostly from mouse embryonic stem cells (ESC). Genome wide chromatin immunoprecipitation combined with deep sequencing (ChIP-seq) has revealed binding of the TET1 protein at CpG-island (CGI) promoters and at bivalent promoters. We show that TET1 also coincides with DNAseI hypersensitive sites (HS). Presence of TET1 at these THREE locations suggests that it may play a dual role: an active role at CpG-islands and DNAseI hypersensitive sites and a repressive role at bivalent loci. In line with the presence of TET1, significant enrichment of 5hmC but not 5mC is detected at bivalent promoters and DNaseI HS. Surprisingly, 5hmC is not detected or present at very low levels at CGI promoters notwithstanding the presence of TET1 at these loci. Our meta analysis suggest that asymmetric methylation is present at CA- and CT-repeats in the genome of some human ESC. Examination of the distribution of 5-methylcytosine and 5-hydroxymethylcytosine in the genome of mouse embryonic stem cells.
2011-11-24 | E-GEOD-31343 | biostudies-arrayexpress
Project description:DNA methylation of C5-cytosine (5mC) in the mammalian genome is a key epigenetic event that is critical for various cellular processes. However, how the genome-wide 5mC pattern is dynamically regulated remains a fundamental question in epigenetic biology. The TET family of 5mC hydroxylases, which convert 5mC to 5-hydroxymethylcytosine (5hmC), have provided a new potential mechanism for the dynamic regulation of DNA methylation. The extent to which individual Tet family members contribute to the genome-wide 5mC and 5hmC patterns and associated gene network remains largely unknown. Here we report genome-wide mapping of Tet1 and 5hmC in mESCs and reveal a mechanism of action by which Tet1 controls 5hmC and 5mC levels in mESCs. In combination with microarray and mRNA-seq expression profiling, we identify a comprehensive yet intricate gene network influenced by Tet1. We propose a model whereby Tet1 controls DNA methylation both by binding to CpG-rich regions to prevent unwanted DNA methyltransferase activity, and by converting the existing 5mC to 5hmC through its enzymatic activity. This Tet1-mediated antagonism of CpG methylation imparts differential maintenance of DNA methylation status at Tet1 target loci, thereby providing a new regulatory mechanism for establishing the epigenetic landscape of mESCs, which ultimately contributes to mESC differentiation and the onset of embryonic development. Tet1 protein was depleted in J1 or E14 mouse ES cells by siRNA or shRNA treatment. Total RNA was purified and used to determine the global gene transcription profiles by microarray assays. The Tet1-regulated genes were identified by comparing the gene expression profiles of control and Tet1-depleted ES cells.
2011-04-21 | E-GEOD-28530 | biostudies-arrayexpress
Project description:Ten-eleven translocation (Tet) hydroxylases (Tet1-3) oxidize 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC). In neurons increased 5hmC levels within gene bodies correlate positively with gene expression. Here, we studied 5hmC profiles (hMeDIP) during retinal maturation between postnatal week 2 and postnatal week 3 using HiSeq 2000 instrument. Study of retinal 5hmC profile dynamics in 2-week old and 3-week old wild type (WT) mouse.
2015-04-02 | E-GEOD-65860 | biostudies-arrayexpress
Project description:Ten-eleven translocation (TET) family enzymes can convert 5-methylcytosine (5mC) into 5-hydroxymethylcytosine (5hmC) in DNA and have been proposed as potential DNA demethylase candidates1. Evidences from recent studies indicated that Tet1 is predominantly expressed in ES cells and plays dual functions in promoting transcription of pluripotency genes and as well as participating in the repression of developmental genes by facilitating recruitment of PRC21-5. These studies further raised the possibility that Tet1 might play a role in somatic cell reprogramming. Here, we provide evidence showing that Tet1 can substitute for pluripotent transcription factors in reprogramming differentiated somatic cells to pluripotent stem cells. Tet1 can replace any one of the four traditional transcription factors including Oct4, Sox2, Klf4 and c-Myc during somatic cell reprogramming. Subsequently, the chimeric mice with germline transmission capacity could be efficiently produced from all induced pluripotent stem (iPS) cell lines reprogrammed by OT (Oct4, Tet1), TSKM (Tet1, Sox2, Klf4, c-Myc), OTK, OTKM and OSTM combinations. Furthermore, the TSKM-reprogrammed iPS cells without using Oct4 could produce viable full-term iPS mice with normal fertility through tetraploid complementation and secondary iPS cells could be induced subsequently from the somatic cells retrieved from the iPS mice. Moreover, we demonstrated that conversion of 5mC into 5hmC in Nanog promoter occurred during reprogramming, which might account in part for the mechanism of Tet1 mediated reprogramming. To our knowledge, our study provides the first evidence demonstrating that DNA modifying enzyme Tet1 can replace the pluripotent transcription factors to reprogram differentiated somatic cells to iPS cells. Gene expression profile of iPS cells and ES cells were generated by Affymetrix Mouse Gene 1.0 ST Array. The Gene expression profile of ES cell R1 was used as control. Three biological repeats were included for each line.
2011-05-07 | E-GEOD-29102 | biostudies-arrayexpress
Project description:Ten eleven translocation enzymes (TET1-3) convert 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC), inducing DNA demethylation and gene regulation. They are often downregulated in cancer, yet the upstream cause of this event remains largely unknown. Here, we show increased TET1 expression in basal-like breast cancer (BLBC), specifically in tumors with low expression of immune markers and low infiltration by immune cells. Furthermore, we identify an anti-correlation between TET1 expression and NF-κB, a major immune regulatory family, in BLBC tissues. In vitro and in vivo, TET1 is downregulated in breast cells upon activation of NF-κB, through binding of p65 to its consensus sequence in the TET1 promoter. Moreover, we uncover genome-wide 5hmC changes related to TET1 regulation in BLBC. Finally, these findings are extended to other cancer types, including melanoma, lung and thyroid cancers. Collectively, our data suggest a novel mode of regulation for TET1, linking TETs and immunity, two major features of cancer.
2018-07-05 | GSE101444 | GEO
Project description:We performed a meta analysis of publicly available TET1, 5mC, 5hmC and genome wide bisulfite profiling data mostly from mouse embryonic stem cells (ESC). Genome wide chromatin immunoprecipitation combined with deep sequencing (ChIP-seq) has revealed binding of the TET1 protein at CpG-island (CGI) promoters and at bivalent promoters. We show that TET1 also coincides with DNAseI hypersensitive sites (HS). Presence of TET1 at these THREE locations suggests that it may play a dual role: an active role at CpG-islands and DNAseI hypersensitive sites and a repressive role at bivalent loci. In line with the presence of TET1, significant enrichment of 5hmC but not 5mC is detected at bivalent promoters and DNaseI HS. Surprisingly, 5hmC is not detected or present at very low levels at CGI promoters notwithstanding the presence of TET1 at these loci. Our meta analysis suggest that asymmetric methylation is present at CA- and CT-repeats in the genome of some human ESC.
2011-11-24 | GSE31343 | GEO
Project description:Methylation at the 5-position of cytosine is a well-studied epigenetic pathway. In addition to 5-methylcytosine (5mC), substantial amounts of 5-hydroxymethylcytosine (5hmC) also referred to as the 6th DNA base, have been detected in certain tissues, most notably the brain. However, the genomic distribution of this cytosine modification is unknown. Here, we have developed an immunoprecipitation technique (5hmC-IP) to examine the occurrence of 5hmC in human DNA from brain frontal lobe tissue. The distribution of 5hmC was compared to that of 5mC. We show that 5hmC is more selectively targeted to genes than is 5mC. 5hmC is found at promoters and is particularly enriched in intragenic regions (gene bodies) but is largely absent from non-gene regions. 5hmC peaks at transcription start sites did not correlate with gene expression levels or H3K4me3 peaks. However, presence of 5hmC in gene bodies was more positively correlated with gene expression levels than was presence of 5mC. Promoters of testis-specific genes showed strong 5mC peaks in brain DNA but were almost completely devoid of 5hmC. Our data provide a first overview of the genomic distribution of 5hmC in human brain and will set the stage for further functional characterization of this novel DNA modification. Comparison between 5hmC and 5mC profiles
2011-03-01 | E-GEOD-27051 | biostudies-arrayexpress