Distinct and overlapping control of 5-methylcytosine and 5-hydroxymethylcytosine by the TET proteins in human cancer cells

摘要

Background: The TET family of dioxygenases catalyze conversion of 5-methylcytosine (5m C) to 5-hydroxymethylcytosine (5hm C), but their involvement in establishing normal 5m C patterns during mammalian development and their contributions to aberrant control of 5m C during cellular transformation remain largely unknown. We depleted TET1, TET2, and TET3 in a pluripotent embryonic carcinoma cell model and examined the impact on genome-wide 5m C, 5hm C, and transcriptional patterns. Results: TET1 depletion yields widespread reduction of 5hm C, while depletion of TET2 and TET3 reduces 5hm C at a subset of TET1 targets suggesting functional co-dependence. TET2 or TET3 depletion also causes increased 5hm C, suggesting these proteins play a major role in 5hm C removal. All TETs prevent hypermethylation throughout the genome, a finding dramatically illustrated in Cp G island shores, where TET depletion results in prolific hypermethylation. Surprisingly, TETs also promote methylation, as hypomethylation was associated with 5hm C reduction. TET function is highly specific to chromatin environment: 5hm C maintenance by all TETs occurs at polycomb-marked chromatin and genes expressed at moderate levels; 5hm C removal by TET2 is associated with highly transcribed genes enriched for H3K4me3 and H3K36me3. Importantly, genes prone to hypermethylation in cancer become depleted of 5hm C with TET deficiency, suggesting that TETs normally promote 5hm C at these loci. Finally, all three TETs, but especially TET2, are required for 5hm C enrichment at enhancers, a condition necessary for expression of adjacent genes. Conclusions: These results provide novel insight into the division of labor among TET proteins and reveal important connections between TET activity, the chromatin landscape, and gene expression.