Mouse ChIP-seq, MeDIP- and hMeDIP-seq

Cell fate determination is influenced by interactions between master transcription factors (TFs) and cis-regulatory elements. Hepatocyte nuclear factor 4 alpha (HNF4A), a liver-enriched TF, acts as a master controller in the specification of hepatic progenitor cells by regulating a network of TFs that controls the onset of hepatocyte cell fate. Using analysis of genome-wide histone modifications, DNA methylation and hydroxymethylation in mouse hepatocytes, we show that HNF4A occupies active enhancers in hepatocytes and essential for active histone (H3K27ac) and DNA signatures, especially 5-hydroxymethylcytosine (5hmC). In mice lacking HNF4A protein in hepatocytes, we observed a decrease in both H3K27ac and hydroxymethylation at regions bound by HNF4A. Mechanistically, HNF4A-associated hydroxymethylation (5hmC) requires its interaction with TET3, a protein responsible for the oxidation from 5mC to 5hmC. Furthermore, HNF4A regulates TET3 expression in liver by directly binding to an enhancer region. In conclusion, we identified that HNF4A is required for the active epigenetic state at enhancers that facilitates transcription from genes in hepatocytes.