Hepatocyte nuclear factor 4α (HNF4α) in coordination with retinoic acid receptors (RARs) increases all‐trans‐retinoic acid (RA)‐dependent CYP26A1 gene expression in hepatocytes

Abstract

CYP26A1, a cytochrome P450, catalyzes exclusively the oxidative inactivation of all‐trans‐RA, the major hormonal metabolite of vitamin A. RA is a pan‐agonist of RARs, promoting RAR binding to DNA response elements of numerous genes. CYP26A1 is very highly induced by RA in the liver, generally to a greater extent than in other tissues, suggesting that a liver‐specific factor may be required for its physiological transcriptional response. In this study, we cotransfected HNF4α, with or without RARs, and demonstrated RA‐dependent activation of a human CYP26A1 promoter‐luciferase construct in HepG2 cells. Analysis of a 2.5‐kbp putative CYP26A1 promoter sequence identified 5 putative HNF4α DNA response elements: H1 located in a proximal region overlapping with an RAR element‐1 (RARE1); H2 and H3 in the distal region, close to RARE2 and RARE3; and H4 and H5 in intermediary regions. Chromatin immunoprecipitation analysis showed significantly increased binding of HNF4α and RARs in the proximal and distal CYP26A1 promoter regions in RA‐treated cells, as compared to the control. Mutational analysis of the individual HNF4α DNA‐response elements identified H1 as the major site for HNF4α binding because H1 mutation inhibited the promoter activity by ~90%, followed by H2 and H3 mutation each with ~50% inhibition. Therefore, our results indicate that HNF4α, as a liver‐enriched nuclear transcription factor, may coordinate with RARs to strongly induce CYP26A1 gene by RA, which may explain the high level of response to RA observed in the liver. (NIH CA‐90214)