Differential regulation of constitutive androstane receptor expression by hepatocyte nuclear factor4α lsoforms

Constitutive androstane receptor (CAR; NR1I3) controls the metabolism and elimination of endogenous and exogenous toxic compounds by up-regulating a battery of genes. In this work, we analyzed the expression of human CAR (hCAR) in normal liver during development and in hepatocellular carcinoma (HCC) and investigated the effect of hepatocyte nuclear factor 4 alpha isoforms (HNF4 alpha 1 and HNF4 alpha 7) on the hCAR gene promoter. By performing functional analysis of hCAR 5'-deletions including mutants, chromatin immunoprecipitation in human hepatocytes, electromobility shift and cotransfection assays, we identified a functional and species-conserved HNF4 alpha response element (DR1: ccAGGCCTtTGCCCTga) at nucleotide - 144. Both HNF4a isoforms bind to this element with similar affinity. However, HNF4 alpha 1 strongly enhanced hCAR promoter activity whereas HNF4a7 was a poor activator and acted as a repressor of HNF4 alpha l-mediated transactivation of the hCAR promoter. PGC1 alpha stimulated both HNF4al-mediated and HN174 alpha 7-mediated hCAR transactivation to the same extent, whereas SRC1 exhibited a marked specificity for HNF4 alpha l. Transduction of human hepatocytes by HNF4a7-expressing lentivitus confirmed this finding. In addition, we observed a positive correlation between CAR and HNF4a I mRNA levels in human liver samples during development, and an inverse correlation between CAR and HNF4a7 mRNA levels in HCC. These observations suggest that HNF4al positively regulates hCAR expression in normal developing and adult livers, whereas HNF4a7 represses hCAR gene expression in HCC.