Maintenance of DNA methylation in embryonic stem cells depends on the histone H3K9 methyltransferases
Ontology highlight
ABSTRACT:
ORGANISM(S): Mus musculus
SUBMITTER: Ulrich Wagner
PROVIDER: E-GEOD-47894 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
Publications
Regulation of DNA methylation turnover at LTR retrotransposons and imprinted loci by the histone methyltransferase Setdb1.
Leung Danny D Du Tingting T Wagner Ulrich U Xie Wei W Lee Ah Young AY Goyal Preeti P Li Yujing Y Szulwach Keith E KE Jin Peng P Lorincz Matthew C MC Ren Bing B
Proceedings of the National Academy of Sciences of the United States of America 20140422 18
During mammalian development, DNA methylation patterns need to be reset in primordial germ cells (PGCs) and preimplantation embryos. However, many LTR retrotransposons and imprinted genes are impervious to such global epigenetic reprogramming via hitherto undefined mechanisms. Here, we report that a subset of such genomic regions are resistant to widespread erasure of DNA methylation in mouse embryonic stem cells (mESCs) lacking the de novo DNA methyltransferases (Dnmts) Dnmt3a and Dnmt3b. Intri ...[more]
Similar Datasets
Project description:During mammalian development DNA methylation patterns need to be reset in primordial germ cells (PGC) and preimplantation embryos. However, many retro-transposons and imprinted genes are resistant to such global epigenetic reprogramming via hitherto undefined mechanisms. Here, we report that some of these sequences are immune to widespread erasure of DNA methylation in the mouse embryonic stem cells (mESCs) lacking de novo DNA methyltransferases. Persistence of DNA methylation at these loci in mESCs depends on the histone H3K9 methyltransferase Setdb1, as deletion of Setdb1 results in reduction of H3K9me3 and DNA methylation levels concomitant with an increase in 5-hydroxymethylation (5hmC). In addition, depletion of H3K9 methyltransferase G9a leads to genome-wide reduction of DNA methylation but to a lesser extent at the above sequences. Taken together, these data reveal that Setdb1 ensures the fidelity of DNA methylation at specific loci in mESCs, which may reflect mechanisms functioning in vivo during key developmental stages. Examination of genome-wide DNA methylation status in 3 cell types.
2014-06-03 | E-GEOD-47890 | biostudies-arrayexpress
Project description:During mammalian development DNA methylation patterns need to be reset in primordial germ cells (PGC) and preimplantation embryos. However, many retro-transposons and imprinted genes are resistant to such global epigenetic reprogramming via hitherto undefined mechanisms. Here, we report that some of these sequences are immune to widespread erasure of DNA methylation in the mouse embryonic stem cells (mESCs) lacking de novo DNA methyltransferases. Persistence of DNA methylation at these loci in mESCs depends on the histone H3K9 methyltransferase Setdb1, as deletion of Setdb1 results in reduction of H3K9me3 and DNA methylation levels concomitant with an increase in 5-hydroxymethylation (5hmC). In addition, depletion of H3K9 methyltransferase G9a leads to genome-wide reduction of DNA methylation but to a lesser extent at the above sequences. Taken together, these data reveal that Setdb1 ensures the fidelity of DNA methylation at specific loci in mESCs, which may reflect mechanisms functioning in vivo during key developmental stages. Examination of genome-wide DNA hydroxy-methylation status in 3 cell types.
2014-06-03 | E-GEOD-47892 | biostudies-arrayexpress
Project description:During mammalian development DNA methylation patterns need to be reset in primordial germ cells (PGC) and preimplantation embryos. However, many retro-transposons and imprinted genes are resistant to such global epigenetic reprogramming via hitherto undefined mechanisms. Here, we report that some of these sequences are immune to widespread erasure of DNA methylation in the mouse embryonic stem cells (mESCs) lacking de novo DNA methyltransferases. Persistence of DNA methylation at these loci in mESCs depends on the histone H3K9 methyltransferase Setdb1, as deletion of Setdb1 results in reduction of H3K9me3 and DNA methylation levels concomitant with an increase in 5-hydroxymethylation (5hmC). In addition, depletion of H3K9 methyltransferase G9a leads to genome-wide reduction of DNA methylation but to a lesser extent at the above sequences. Taken together, these data reveal that Setdb1 ensures the fidelity of DNA methylation at specific loci in mESCs, which may reflect mechanisms functioning in vivo during key developmental stages. Examination of H3K9me3 histone modifications in 2 cell types.
2014-06-03 | E-GEOD-47887 | biostudies-arrayexpress
Project description:During mammalian development DNA methylation patterns need to be reset in primordial germ cells (PGC) and preimplantation embryos. However, many retro-transposons and imprinted genes are resistant to such global epigenetic reprogramming via hitherto undefined mechanisms. Here, we report that some of these sequences are immune to widespread erasure of DNA methylation in the mouse embryonic stem cells (mESCs) lacking de novo DNA methyltransferases. Persistence of DNA methylation at these loci in mESCs depends on the histone H3K9 methyltransferase Setdb1, as deletion of Setdb1 results in reduction of H3K9me3 and DNA methylation levels concomitant with an increase in 5-hydroxymethylation (5hmC). In addition, depletion of H3K9 methyltransferase G9a leads to genome-wide reduction of DNA methylation but to a lesser extent at the above sequences. Taken together, these data reveal that Setdb1 ensures the fidelity of DNA methylation at specific loci in mESCs, which may reflect mechanisms functioning in vivo during key developmental stages. Examination of genome-wide DNA methylation status in 2 cell types.
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Project description:During mammalian development DNA methylation patterns need to be reset in primordial germ cells (PGC) and preimplantation embryos. However, many retro-transposons and imprinted genes are resistant to such global epigenetic reprogramming via hitherto undefined mechanisms. Here, we report that some of these sequences are immune to widespread erasure of DNA methylation in the mouse embryonic stem cells (mESCs) lacking de novo DNA methyltransferases. Persistence of DNA methylation at these loci in mESCs depends on the histone H3K9 methyltransferase Setdb1, as deletion of Setdb1 results in reduction of H3K9me3 and DNA methylation levels concomitant with an increase in 5-hydroxymethylation (5hmC). In addition, depletion of H3K9 methyltransferase G9a leads to genome-wide reduction of DNA methylation but to a lesser extent at the above sequences. Taken together, these data reveal that Setdb1 ensures the fidelity of DNA methylation at specific loci in mESCs, which may reflect mechanisms functioning in vivo during key developmental stages. Examination of H3K9me3 histone modifications in 2 cell types.
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Project description:Genome modification in mouse embryonic stem cells (mESCs) has provided the versatile platform to analyzing gene functions in mammalian cells. Bloom helicase-deficient mESCs showed elevated rate of loss of heterozygosity (LOH) through mitotic recombination. By utilizing this characteristic of Blm deficiency, we established an efficient method to introduce homozygous mutations into mESCs, which can be used for phenotype-driven recessive screen. It is generally thought that mitotic recombination is initiated by double-strand break. DNA repair machinery, however, possesses mechanisms that suppress deleterious crossovers between homologous chromosomes. Bloom helicase plays a central role to suppress such crossover; hence its deficiency causes elevated frequency of mitotic recombination. To directly answer if DSB can initiate mitotic recombination, we established a reporter cell line, in which the recognition site of I-SceI endonuclease was introduced in the Aprt gene in Blm-deficient mESCs. We used two types of I-SceI expression vectors; one expresses a mammalian codon-optimized I-SceI (ISceIo) and the other expresses I-SceIo fused with N-terminal 110-amino acids of Geminin (Gemi). Geminin-fused I-SceI can be stabilized only in lateS/G2 phase. After I-SceI transfection and subsequent selection, we isolated Aprt-deficient mutants. Half of the mutants showed LOH in the whole telomeric region from I-SceI site, but the other half showed local LOH near the I-SceI site. Further analyses suggested that large deletion was introduced and caused LOH, which was not seen in spontaneous LOH. We designed CGH probes in 300-bp resolution in 8Mb telomeric region of chr 8 (The Aprt gene locates 7Mb from the telomere) to map the deleted regions in these mutants.
2012-11-30 | E-MTAB-1116 | biostudies-arrayexpress
Project description:During mammalian development DNA methylation patterns need to be reset in primordial germ cells (PGC) and preimplantation embryos. However, many retro-transposons and imprinted genes are resistant to such global epigenetic reprogramming via hitherto undefined mechanisms. Here, we report that some of these sequences are immune to widespread erasure of DNA methylation in the mouse embryonic stem cells (mESCs) lacking de novo DNA methyltransferases. Persistence of DNA methylation at these loci in mESCs depends on the histone H3K9 methyltransferase Setdb1, as deletion of Setdb1 results in reduction of H3K9me3 and DNA methylation levels concomitant with an increase in 5-hydroxymethylation (5hmC). In addition, depletion of H3K9 methyltransferase G9a leads to genome-wide reduction of DNA methylation but to a lesser extent at the above sequences. Taken together, these data reveal that Setdb1 ensures the fidelity of DNA methylation at specific loci in mESCs, which may reflect mechanisms functioning in vivo during key developmental stages. Examination of gene expression in 2 cell types by RNA-seq.
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2015-06-15 | E-GEOD-66022 | biostudies-arrayexpress
Project description:Argonaute (Ago) proteins mediate post-transcriptional gene repression by binding guide microRNAs (miRNAs) to regulate targeted RNAs. To confidently assess Agobound small RNAs, we adapted a mouse embryonic stem cell system to express a single inducible epitope-tagged Ago protein. Here, we report the small RNA profile of Agodeficient cells and determine Ago-dependent stability is a common feature of mammalian miRNAs. Considering both in vivo Ago-dependence for stability and Ago2 binding as defined by immunopurification, we have identified a novel class of non-canonical miRNAs derived from protein-coding gene promoters, which we name transcriptional start site miRNAs (TSS-miRNAs). A subset of promoter-proximal RNA polymerase II complexes produce hairpin RNAs that are processed in a DGCR8/Drosha-independent, but Dicer-dependent manner. TSS-miRNA activity is detectable endogenously, upon transfection of a mimic or by mRNA overexpression. Finally, we present evidence of differential expression and conservation in humans, suggesting important roles in gene regulation. Examination of Ago immunoprecipitations and mESC without Ago proteins
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Project description:Ten-eleven translocation (Tet) family of DNA dioxygenases converts 5-methylcytosine (5mC) into 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC), and 5- carboxylcytosine (5caC) through iterative oxidation reactions. While 5mC and 5hmC are relatively abundant, 5fC and 5caC are at very low levels in the mammalian genome. Thymine DNA glycosylase (TDG) and base excision repair (BER) pathways can actively remove 5fC/5caC to regenerate unmethylated cytosine, but it is unclear to what extent and at which part of the genome such active demethylation processes take place. Here, we have performed high-throughput sequencing analysis of 5mC/5hmC/5fC/5caC- enriched DNA using modification-specific antibodies and generated genome-wide distribution maps of these cytosine modifications in wild-type and Tdg-deficient mouse embryonic stem cells (ESCs). We observe that the steady state 5fC and 5caC are preferentially detected at repetitive sequences in wild-type mouse ESCs. Depletion of TDG causes marked accumulation of 5fC and 5caC at a large number of distal gene regulatory elements and transcriptionally repressed/poised gene promoters, suggesting that Tet/TDG-dependent dynamic cycling of 5mC oxidation states may be involved in regulating the function of these regions. Thus, comprehensive mapping of 5mC oxidation and BER pathway activity in the mammalian genome provides a promising approach for better understanding of biological roles of DNA methylation and demethylation dynamics in development and diseases. In this dataset, we include the DIP-seq data of 5mC, 5hmC, 5fC and 5caC in both control and Tdg knockdown mouse embryonic stem cells.
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