Differential RNA expression profiling in response to TET inhbition
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE137282 | GEO | 2019/12/31
REPOSITORIES: GEO
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Project description:p53 is one of the most frequently mutated genes in human cancers, which could regulate multiple target genes involved in a widespread of cellular processes. The regulation of DNA methylation dynamics is critical for maintaining genome stability and proper gene expression. Recently a family of Ten-Eleven-Translocation (TET) proteins has been shown to possess the enzymatic activity to oxidize 5mC to 5hmC, and contribute to DNA demethylation. Here we explore the relationship between p53 binding, 5hmC modification and gene activation using human p53-deficient cells (H1299). H1299 Cells with and without p53
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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.
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Project description:p53 is one of the most frequently mutated genes in human cancers, which could regulate multiple target genes involved in a widespread of cellular processes. The regulation of DNA methylation dynamics is critical for maintaining genome stability and proper gene expression. Recently a family of Ten-Eleven-Translocation (TET) proteins has been shown to possess the enzymatic activity to oxidize 5mC to 5hmC, and contribute to DNA demethylation. Here we explore the relationship between p53 binding, 5hmC modification and gene activation using human p53-deficient cells (H1299).
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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.
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Project description:Enzymatic oxidation of 5-methylcytosine (5mC) in DNA by the Tet (Ten-eleven translocation) family of dioxygenases reprograms genome function during pre-implantation development and in primordial germ cells. Tet-oxidized derivatives of 5mC such as 5-hydroxymethylcytosine (5hmC) act as transient intermediates in DNA demethylation or serve as stable epigenetic marks, yet how these alternate fates are specified at individual CpGs is not understood. Here, we report that the Sos-response associated peptidase (SRAP) domain-containing protein Srap1, the mammalian orthologue of an ancient protein superfamily operonically associated with the DNA damage response in bacteria and bacteriophage, binds directly and selectively to Tet-oxidized forms of 5mC in DNA and catalyses turnover of these bases to unmodified cytosine by an autopeptidase cleavage-coupled endonuclease. Biallelic inactivation of Srap1 in mouse embryonic stem (ES) cells results in the global accumulation of Tet-oxidized 5mC derivatives, with ectopic 5hmC in promoters of highly expressed genes, across gene bodies and within enhancer elements. Srap1 deficiency causes partially penetrant lethality in late blastocysts prior to uterine implantation, while surviving midgestation embryos exhibit aberrant patterns of DNA methylation associated with altered gene expression, including hypermethylation in regions closely linked to ectopic 5hmC detected earlier in development. Thus, highly specific removal of 5hmC and its oxidized derivatives by Srap1 sustains genome-wide DNA methylation homeostasis. These findings establish a function for a previously unknown structural class of DNA base modification-specific eraser enzymes that combine the catalytic properties of an autopeptidase with endonuclease activation, and position Srap1 as a key determinant of 5mC demethylation trajectories during mammalian embryonic development.
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Project description:DNA methylation at 5-position of cytosine (5mC) is one of the best studied epigenetic modifications that 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), which can be further processed by DNA repair proteins leading to DNA demethylation. Functional characterization of the Tet proteins has been complicated by the redundancy between the three Tet proteins. Using the CRISPR/Cas9 technology, we have generated mouse embryonic stem cells (ESCs) deficient for all three Tet proteins (TKO). Whole genome bisulphite sequencing (WGBS) analysis revealed that Tet-mediated DNA demethylation mainly occurs distal enhancers as well as promoters that significantly overlap with 5hmC, 5fC and 5caC. Characterization of the Tet TKO ESCs revealed a function for Tet proteins in cell fate restriction as Tet TKO ESCs tend to adopt both primed pluripotent stem cell-like state and 2-cell embryo-like state. In addition, Tet TKO ESCs exhibit elongated telomeres. Thus, our study reveals a role of Tet proteins not only in DNA demethylation, but also in cell fate restriction and telomere maintenance. 2 samples for WGBS and 2 samples for RNA-seq
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