TGF-β-Dependent Active Demethylation and Expression of the p15ink4b Tumor Suppressor Are Impaired by the ZNF217/CoREST Complex

SummaryIn this study we examine the mechanisms of dynamic DNA methylation of the p15ink4b tumor suppressor gene. Using conventional ChIP and ChiPseq, we identify the p15ink4b promoter as a target for the ZNF217 oncogene, the CoREST complex, and DNMT3A. Treatment of cells with TGF-β triggers active demethylation involving loss of ZNF217/CoREST/DNMT3A and the corecruitment of SMAD2/3, CBP, and the DNA glycosylase TDG. Knockdown of TDG, or its functional homolog MBD4, prevents TGF-β-dependent demethylation of p15ink4b. DNA immunoprecipitation of 5mC and 5hmC indicates that 5mC undergoes conversion to 5hmC prior to activation of p15ink4b. Remarkably, overexpression of ZNF217 inhibits active demethylation and expression of the p15ink4b gene by preventing recruitment of SMAD2/3 and TDG. These findings suggest that active demethylation is essential for regulating a subset of TGF-β-dependent genes. Importantly, disruption of active demethylation by the ZNF217 oncogene may be a paradigm for other oncogenic signals on DNA methylation dynamics.Highlights•p15ink4b gene is a target for ZNF217/CoREST and DNMT3A•TGF-β triggers active DNA demethylation of p15ink4b•TGF-β-dependent active demethylation requires TDG or MBD4•Overexpressing ZNF217 inhibits active DNA demethylation and expression of p15ink4b