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Series GSE46402 Query DataSets for GSE46402
Status Public on Jun 30, 2013
Title Vitamin C induces Tet-dependent DNA demethylation in ES cells to promote a blastocyst-like methylome [MeDIP-Seq]
Organism Mus musculus
Experiment type Methylation profiling by high throughput sequencing
Summary DNA methylation is a heritable epigenetic modification involved in gene silencing, imprinting, and the suppression of retrotransposons. Global DNA demethylation occurs in the early embryo and the germline and may be mediated by Tet (ten-eleven-translocation) enzymes, which convert 5-methylcytosine (mC) to 5-hydroxymethylcytosine (hmC). Tet enzymes have been extensively studied in mouse embryonic stem (ES) cells, which are generally cultured in the absence of Vitamin C, a potential co-factor for Fe(II) 2-oxoglutarate dioxygenase enzymes like Tets. Here we report that addition of Vitamin C to ES cells promotes Tet activity leading to a rapid and global increase in hmC. This is followed by DNA demethylation of numerous gene promoters and up-regulation of demethylated germline genes. Tet1 binding is enriched near the transcription start site (TSS) of genes affected by Vitamin C treatment. Importantly, Vitamin C, but not other antioxidants, enhances the activity of recombinant human Tet1 in a biochemical assay and the Vitamin C-induced changes in hmC and mC are entirely suppressed in Tet1/2 double knockout (Tet DKO) ES cells. Vitamin C has the strongest effects on regions that gain methylation in cultured ES cells compared to blastocysts and in vivo are methylated only after implantation. In contrast, imprinted regions and intracisternal A-particle (IAP) elements, which are resistant to demethylation in the early embryo, are resistant to Vitamin C-induced DNA demethylation. Collectively, this study establishes that Vitamin C is a direct regulator of Tet activity and DNA methylation fidelity in ES cells.
Overall design Oct4-GiP mouse embryonic stem (ES) cells were cultured in the presence or absence of Vitamin C (L-ascorbic acid 2-phosphate, 100 μg/ml) for 12 or 72 hours. Cells were maintained in N2B27 medium supplemented with LIF (1000 U/ml), MEK inhibitor PD0325901 (1 μM), and GSK3β inhibitor CHIR99021 (3 μM). Genomic DNA was used for DNA immunoprecipitation with antibodies against 5-hydroxymethylcytosine (hmC) or 5-methylcytosine (mC). Immunoprecipitated DNA was adaptor-ligated for paired-end sequencing on an Illumina HiSeq and sequence reads were aligned to the mm9 mouse reference genome for analysis.
Contributor(s) Blaschke K, Ebata KT, Karimi MM, Hirst M, Lorincz MC, Ramalho-Santos M
Citation(s) 23812591
Submission date Apr 26, 2013
Last update date May 15, 2019
Contact name Martin Hirst
Phone 604-8226673
Organization name BCCA Genome Sciences Centre
Department Epigenomics
Street address 675 West 10th Avenue
City Vancouver
State/province BC
ZIP/Postal code V5Z 1L3
Country Canada
Platforms (1)
GPL13112 Illumina HiSeq 2000 (Mus musculus)
Samples (8)
GSM1129767 hmeDIP-Seq untreated 12hr
GSM1129768 hmeDIP-Seq untreated 72hr
GSM1129769 hmeDIP-Seq vitamin C treated 12hr
This SubSeries is part of SuperSeries:
GSE46403 Vitamin C induces Tet-dependent DNA demethylation in ES cells to promote a blastocyst-like methylome
BioProject PRJNA200282
SRA SRP021527

Download family Format
SOFT formatted family file(s) SOFTHelp
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Series Matrix File(s) TXTHelp

Supplementary file Size Download File type/resource
GSE46402_RAW.tar 642.9 Mb (http)(custom) TAR (of WIG)
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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