TETs compete with DNMT3 activity in pluripotent cells at thousands of methylated somatic enhancers

Mammalian cells stably maintain high levels of DNA methylation despite expressing both positive (DNMT3A/B) and negative (TET1–3) regulators. Here, we used wildtype and TET triple knockout human embryonic stem cells (ESCs), generated DNMT3-null as well as TET and DNMT3-null pentuple knockouts, and compared methylation patterns using whole genome bisulfite sequencing (WGBS). The greatest impact on global methylation levels was observed in DNMT3-deficient cells, including reproducible focal demethylation at thousands of normally methylated loci. This demethylation depends upon TET expression and only occurs when both DNMT3s are absent. Dynamic loci are enriched for hydroxymethylcytosine and overlap with subsets of putative somatic enhancers that are methylated in ESCs and can be activated upon differentiation. We observe similar dynamics in mouse ESCs that were less frequenct in epiblast stem cells (EpiSCs) and scarce in somatic tissues, suggesting a conserved pluripotency-linked mechanism. Taken together, our data reveal tightly regulated competition between DNMT3s and TETs at thousands of somatic regulatory sequences within pluripotent cells.