Differentiation-dependent changes in the epigenomic landscape of human myeloid leukemia cells implicate signaling pathways and highlight chromosomal amplifications
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE66765 | GEO | 2017/03/16
SECONDARY ACCESSION(S): PRJNA277878
REPOSITORIES: GEO
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Project description:Cytosine DNA bases can be methylated by DNA methyltransferases and subsequently oxidized by TET proteins. The resulting 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC), and 5-carboxylcytosine (5caC) are considered demethylation intermediates as well as stable epigenetic marks. To dissect the contribution of these cytosine modifying enzymes, we generated combinations of Tet knockout (KO) embryonic stem cells (ESCs) and systematically measured protein and DNA modification levels at the transition from naive to primed pluripotency. Whereas the increase of genomic 5-methylcytosine (5mC) levels during exit from pluripotency correlated with an upregulation of the de novo DNA methyltransferases DNMT3A and DNMT3B, the subsequent oxidation steps turned out to be far more complex. The strong increase of oxidized cytosine bases (5hmC, 5fC, and 5caC) was accompanied by a drop in TET2 levels, yet the analysis of KO cells suggested that TET2 is responsible for most 5fC formation. The comparison of modified cytosine and enzyme levels in Tet KO cells revealed distinct and differentiation-dependent contributions of TET1 and TET2 to 5hmC and 5fC formation arguing against a processive mechanism of 5mC oxidation. The apparent independent steps of 5hmC and 5fC formation suggest yet to be identified mechanisms regulating TET activity and may constitute another layer of epigenetic regulation.
2020-07-24 | PXD019514 | Pride
Project description:DNA methylation (5mC) plays important roles in epigenetic regulation of genome function, and recently the TET1-3 hydroxylases have been found to oxidize 5mC to hydroxymethylcytosine (5hmC), formylcytosine (5fC), and carboxylcytosine (5caC) in DNA. These derivatives have a role in demethylation of DNA but in addition may have epigenetic signaling functions in their own right. A recent study identified proteins with preferential binding to 5-methylcytosine (5mC) and its oxidized forms where readers for 5mC and 5hmC (5-hydroxymethylcytosine) showed little overlap while further oxidation forms enriched for repair proteins and transcription regulators. We extend this study by using promoter sequences as baits and compare protein binding patterns to unmodified or modified cytosine containing DNA using mouse embryonic stem cell (mESCs) extracts. The dataset contains 3 biological replicates each of mouse ES cell nuclear proteins binding to Pax6 and FGF15 promoter sequences containing different modified forms of cytosine. Data analysis: Mass spectrometric data were processed using Proteome Discoverer v1.3 and searched against a mammalian entries in Uniprot 2011.09 using Mascot v2.3 with the following parameters: Enzyme - trypsin; max 1 missed cleavage; Precursor Mass Tolerance - 10 ppm; Fragment Mass Tolerance - 0.6 Da; Dynamic Modification - Oxidation (M); Static Modification - Carbamidomethyl at C.
2013-10-21 | PXD000524 | Pride
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
Project description:Oxidative modification of 5-methylcytosine (5mC) by TET DNA dioxygenases generates 5-hydroxymethylcytosine (5hmC), the most abundant form of oxidized 5mC. Existing single-cell bisulfite sequencing methods cannot resolve 5mC and 5hmC, leaving the cell-type-specific regulatory mechanisms of TET and 5hmC largely unknown. Here we present Joint single-nucleus (hydroxy)methylcytosine sequencing (Joint-snhmC-seq), a scalable and quantitative approach that simultaneously profiles 5hmC and true 5mC in single cells by harnessing differential deaminase activity of APOBEC3A towards 5mC and chemically protected 5hmC. Joint-snhmC-seq profiling of single nuclei from the mouse brains reveals an unprecedented level of epigenetic heterogeneity of both 5hmC and true 5mC at single-cell resolution. We show that cell-type-specific profiles of 5hmC or true 5mC improve multi-modal single-cell data integration, enable accurate identification of neuronal subtypes, and uncover context-specific regulatory effects of cell-type-specific genes by TET enzymes.
2023-07-14 | GSE236776 | GEO
Project description:Oxidative modification of 5-methylcytosine (5mC) by TET DNA dioxygenases generates 5-hydroxymethylcytosine (5hmC), the most abundant form of oxidized 5mC. Existing single-cell bisulfite sequencing methods cannot resolve 5mC and 5hmC, leaving the cell-type-specific regulatory mechanisms of TET and 5hmC largely unknown. Here we present Joint single-nucleus (hydroxy)methylcytosine sequencing (Joint-snhmC-seq), a scalable and quantitative approach that simultaneously profiles 5hmC and true 5mC in single cells by harnessing differential deaminase activity of APOBEC3A towards 5mC and chemically protected 5hmC. Joint-snhmC-seq profiling of single nuclei from the mouse brains reveals an unprecedented level of epigenetic heterogeneity of both 5hmC and true 5mC at single-cell resolution. We show that cell-type-specific profiles of 5hmC or true 5mC improve multi-modal single-cell data integration, enable accurate identification of neuronal subtypes, and uncover context-specific regulatory effects of cell-type-specific genes by TET enzymes.
2023-07-14 | GSE236789 | GEO
Project description:Oxidative modification of 5-methylcytosine (5mC) by TET DNA dioxygenases generates 5-hydroxymethylcytosine (5hmC), the most abundant form of oxidized 5mC. Existing single-cell bisulfite sequencing methods cannot resolve 5mC and 5hmC, leaving the cell-type-specific regulatory mechanisms of TET and 5hmC largely unknown. Here we present Joint single-nucleus (hydroxy)methylcytosine sequencing (Joint-snhmC-seq), a scalable and quantitative approach that simultaneously profiles 5hmC and true 5mC in single cells by harnessing differential deaminase activity of APOBEC3A towards 5mC and chemically protected 5hmC. Joint-snhmC-seq profiling of single nuclei from the mouse brains reveals an unprecedented level of epigenetic heterogeneity of both 5hmC and true 5mC at single-cell resolution. We show that cell-type-specific profiles of 5hmC or true 5mC improve multi-modal single-cell data integration, enable accurate identification of neuronal subtypes, and uncover context-specific regulatory effects of cell-type-specific genes by TET enzymes.
2023-07-14 | GSE236784 | GEO
Project description:Oxidative modification of 5-methylcytosine (5mC) by TET DNA dioxygenases generates 5-hydroxymethylcytosine (5hmC), the most abundant form of oxidized 5mC. Existing single-cell bisulfite sequencing methods cannot resolve 5mC and 5hmC, leaving the cell-type-specific regulatory mechanisms of TET and 5hmC largely unknown. Here we present Joint single-nucleus (hydroxy)methylcytosine sequencing (Joint-snhmC-seq), a scalable and quantitative approach that simultaneously profiles 5hmC and true 5mC in single cells by harnessing differential deaminase activity of APOBEC3A towards 5mC and chemically protected 5hmC. Joint-snhmC-seq profiling of single nuclei from the mouse brains reveals an unprecedented level of epigenetic heterogeneity of both 5hmC and true 5mC at single-cell resolution. We show that cell-type-specific profiles of 5hmC or true 5mC improve multi-modal single-cell data integration, enable accurate identification of neuronal subtypes, and uncover context-specific regulatory effects of cell-type-specific genes by TET enzymes.
2023-07-14 | GSE236769 | GEO
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. The mechanisms controlling Tet activity and 5hmC levels are poorly understood. In particular, it is not known how the neuronal Tet3 isoform lacking a DNA binding domain is targeted to the DNA. To identify factors binding to Tet3 we screened for proteins that co-precipitate with Tet3 from mouse retina and identified the transcriptional repressor Rest as a highly enriched Tet3-specific interactor. Rest was able to enhance Tet3 hydroxylase activity after co-expression and overexpression of Tet3 activated transcription of Rest-target genes. Moreover, we found that Tet3 also interacts with Nsd3 and two other H3K36 methyltransferases and is able to induce H3K36 trimethylation. We propose a mechanism for transcriptional activation in neurons that involves Rest-guided targeting of Tet3 to the DNA for directed 5hmC-generation and Nsd3-mediated H3K36 trimethylation.
2015-04-02 | PXD001857 | Pride
Project description:In human and mouse stem cells and brain, 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) can occur outside of CG dinucleotides. Using protein binding microarrays (PBMs) containing 60-mer DNA probes, we evaluated the effect of 5mC and 5hmC on one DNA strand on the double-stranded DNA binding of the mouse B-ZIP transcription factors (TFs) CREB1, ATF1, and JUND. 5mC inhibited CREB1 binding to the canonical CRE half-site |GTCA, but increased binding to the C/EBP half-site |GCAA. 5hmC inhibited CREB1 binding to all 8-mers except TGAT|GCAA, where binding is enhanced. We observed similar DNA binding patterns with the closely related TF: ATF1. In contrast, both 5mC and 5hmC inhibited binding of JUND. These results identify new DNA sequences that are well-bound by CREB1 and ATF1 only when they contain 5mC or 5hmC. Analysis of two x-ray structures examines the consequences of 5mC and 5hmC on DNA binding by CREB and FOS|JUN.
2016-10-19 | GSE88897 | GEO
Project description:Prenatal exposure to neurotoxicants such as lead (Pb) may cause stable changes in the DNA methylation (5mC) profile of the fetal genome. However few studies have examined its effect on the DNA de-methylation pathway, specifically the dynamic changes of the 5-hydroxymethylcytosine (5hmC) profile. Therefore, in this study, we investigate the relationship between Pb exposure and 5mC and 5hmC modifications during early development. To study the changes in the 5hmC profile, we use a novel modification of the Infinium™ Human methylation 450K assay (Illumina, Inc.), which we named HMeDIP-450K assay, in an in vitro human embryonic stem cell model of Pb-exposure. We model Pb-exposure associated 5hmC changes as clusters of correlated, adjacent CpG sites, which are co-responding to Pb. We further extend our study to look at Pb-dependent changes in high density 5hmC regions in umbilical cord blood DNA from 48 mother-infant pairs from the Early Life Exposure in Mexico to Environmental Toxicants (ELEMENT) cohort. For our study, we randomly selected UCB from 24 male and 24 female children from the 1st and 4th quartiles of Pb levels. Our data show that Pb-associated changes in the 5hmC and 5mC profiles can be divided into sex-dependent and sex-independent categories. Interestingly, differential 5mC sites are better markers of Pb-associated sex-dependent changes compared to differential 5hmC sites. In this study we identified several 5hmC and 5mC genomic loci, which we believe might have some potential as early biomarkers of prenatal Pb-exposure. Human Methylation 450K array coupled with 5-hydroxymethylcytosine (5hmC) IP or HMeDIP-450K array, was used to determine the high density 5hmC profile of 46 Umbilical cord blood samples from the ELLEMENT cohort. HM450K array, without IP was used to determine the 5-methylcytosine(5mC) plus 5hmC profile. The 5hmC region detected from the HMeDIP-450K array was subtracted from the HM450K(without IP) to get putative high density 5mC regions.
2015-06-08 | E-GEOD-69633 | biostudies-arrayexpress