Characterization of orphan nuclear receptor binding elements in sex-differentiated members of the CYP2C gene family expressed in rat liver.

The HepG2-specific P450 2C factor motif (HPF1-motif) is conserved in many hepatic cytochrome P450 genes (CYP). Its functional importance for rabbit CYP2C genes has led to the proposal that the HPF1 motif acts as a common regulator for the liver-specific expression of CYP2 genes with hepatic nuclear factor (HNF)-4 being the corresponding trans-activator. The HPF1-like elements in the rat CYP2C genes 2C7, 2C11, 2C12, and 2C13 have been studied with regard to functional importance and binding of the orphan receptors HNF-4, apoAI regulatory protein-1 (ARP-1), and v-erbA-related receptors (EAR) 3 and 2. Binding activity in rat liver nuclear extracts includes these orphan receptors as judged from electromobility supershift experiments and from results obtained with expressed receptors, although the element in CYP2C11 did not bind HNF-4. Mutations of the HPF1-like elements in the CYP2C7, CYP2C11, and CYP2C12 promoters had marginal effects on the expression of luciferase reporter gene constructs transiently transfected into HepG2 cells, whereas for CYP2C13 the activity was reduced to 60% of the wild type construct. Coexpression of HNF-4 in COS-7 cells had limited effect on the luciferase activity generated from the 2C promoters, maximally 3-fold. Our data indicate that the HPF1 elements in the rat CYP2C genes have limited functional importance and that HNF-4 is not a major trans-activator for any of these genes.