Abstract
De novo establishment of DNA methylation is accomplished by DNMT3A and DNMT3B. Here, we analyze de novo DNA methylation in mouse embryonic fibroblasts (2i-MEFs) derived from DNA-hypomethylated 2i/L ES cells with genetic ablation of Dnmt3a or Dnmt3b. We identify 355 and 333 uniquely unmethylated genes in Dnmt3a and Dnmt3b knockout (KO) 2i-MEFs, respectively. We find that Dnmt3a is exclusively required for de novo methylation at both TSS regions and gene bodies of Polycomb group (PcG) target developmental genes, while Dnmt3b has a dominant role on the X chromosome. Consistent with this, tissue-specific DNA methylation at PcG target genes is substantially reduced in Dnmt3a KO embryos. Finally, we find that human patients with DNMT3 mutations exhibit reduced DNA methylation at regions that are hypomethylated in Dnmt3 KO 2i-MEFs. In conclusion, here we report a set of unique de novo DNA methylation target sites for both DNMT3 enzymes during mammalian development that overlap with hypomethylated sites in human patients.
Highlights
De novo establishment of DNA methylation is accomplished by DNMT3A and DNMT3B
That mouse embryonic stem (ES) cells cultured in serum and leukemia inhibitory factor (LIF) (S/L ES cells) exhibit global DNA hypermethylation relative to inner cell mass (ICM) cells, which are the in vivo counterpart of ES cells[17]
We found that DNMT3B has a higher activity of de novo methylation than DNMT3A, which is remarkably pronounced at CGIs on X chromosome
Summary
We analyze de novo DNA methylation in mouse embryonic fibroblasts (2i-MEFs) derived from DNA-hypomethylated 2i/L ES cells with genetic ablation of Dnmt3a or Dnmt3b. Three conserved DNA methyltransferases, DNMT3A, DNMT3B, and DNMT1, are responsible for de novo establishment and maintenance of DNA methylation patterns[4]. The genomic enrichment patterns of DNMT3A differ from those of DNMT3B in mouse embryonic stem (ES) cells, suggesting that each de novo methyltransferase has unique targets during development[14,15]. Individual genetic ablation of these methyltransferases results in lethal phenotypes at different developmental stages[5] It remains unclear whether the differential targeting of DNMT3 enzymes is responsible for locus-specific de novo DNA methylation during embryonic development. In combination with genetic ablation of Dnmt3a or Dnmt3b, here we report unique de novo DNA methylation target sites for both DNMT3 enzymes during embryonic development
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