DNA methylation controls unmethylated transcription start sites in the genome in trans

The inverse correlation between DNA methylation and steady state mRNA levels is imperfect; a large number of genes, which are unmethylated in the promoter region, are silenced. We show here that either pharmacological inhibition of DNA methyltransferase (DNMT1) with 5-aza-2'-deoxycytidine (5-azaCdR) or knockdown with shRNA dramatically expands the number of transcription initiation positions in the genome including many unmethylated promoters. Induction of unmethylated promoters in response to inhibition of DNMT1 is a result of activation by demethylation of methylated upstream regulators such as the transcription factor HNF4A. Our results imply that the landscape of genes that are regulated by DNA methylation is more wide-ranging than genes silenced by methylation of their own cis regulatory sequences. We propose a hierarchical model of transcriptional control by DNA methylation whereby regulation of unmethylated promoters is dependent on the methylation state of trans upstream regulators. RNAPII ser 5 ChIP DNA from primary liver culture and SKHep1 cancer adenocarcinoma cells, treated with scrambled control, 1uM of 5azaCdR and ShDNMT1