Histone deacetylase inhibitors reverse CpG methylation by regulating DNMT1 through ERK signaling

Methylation of CpG repeats in the upstream/promoter regions of genes is an established mechanism of gene silencing in many cell types. DNA methylation results in the recruitment of histone deacetylases (HDACs) to promoter regions, thereby repressing expression of genes. General inhibitors of class I and II HDACs (HDACi), such as sodium butyrate and suberoylanilide hydroxamic acid, suppress the growth of prostate cancer cells in vitro and in vivo. In this study, we investigated the mechanism of re-expression of silenced cell cycle inhibitors and retinoic acid receptor B2 (RARB2). HDACi inhibited cell cycle progression, and reversed promoter methylation and silencing of three tumor suppressor genes: RARB2 and the cell cycle regulating cyclin-dependent kinase inhibitors p16 and p21. HDACi repressed MAP kinase I (ERK) activation and down-regulated DNA (cytosine-5-)-methyltransferase 1 (DNMT1) levels. Direct inhibition of ERK activity similarly decreased DNMT1 protein levels and reversed the basal hypermethylation of the promoters and silencing of the RARB2, p21 and p16 tumor suppressor genes. Suppression of DNMT1 level by siRNA also reversed methylation of these tumor suppressor genes with similar kinetics. Collectively, these data demonstrate that HDACi, by inhibiting ERK activity, regulate DNMT1 and ultimately DNA methylation. These results demonstrate that HDACs regulate gene methylation, in addition to the established and reciprocal ability of CpG methylation to recruit HDACs to repress transcription.