Acute Depletion Redefines the Division of Labor among DNA Methyltransferases in Methylating the Human Genome

• Epigenome profiling after combination depletion identifies DNMT interactions
• DNMT3B regulates initiation of differentiation pathways
• DNMT1 and DNMT3B inversely coregulate 5mC and 5hmC at conserved loci
• DNMT3B promotes non-CpG methylation; DNMT3L regulates CpG versus non-CpG choice

SummaryGlobal patterns of DNA methylation, mediated by the DNA methyltransferases (DNMTs), are disrupted in all cancers by mechanisms that remain largely unknown, hampering their development as therapeutic targets. Combinatorial acute depletion of all DNMTs in a pluripotent human tumor cell line, followed by epigenome and transcriptome analysis, revealed DNMT functions in fine detail. DNMT3B occupancy regulates methylation during differentiation, whereas an unexpected interplay was discovered in which DNMT1 and DNMT3B antithetically regulate methylation and hydroxymethylation in gene bodies, a finding confirmed in other cell types. DNMT3B mediated non-CpG methylation, whereas DNMT3L influenced the activity of DNMT3B toward non-CpG versus CpG site methylation. Altogether, these data reveal functional targets of each DNMT, suggesting that isoform selective inhibition would be therapeutically advantageous.

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