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 m any hepatic cytochrome P450 genes (CYP). Its functional importance for rabbit CYP2C genes has led to the proposal that the HPF1 motif acts a s a common regulator for the liver-specific expression of CYP2 genes w ith hepatic nuclear factor (HNF)-4 being the corresponding trans-activ ator, The HPF1-like elements in the rat CYP2C genes 2C7, 2C11, 2C12, a nd 2C13 have been studied with regard to functional importance and bin ding of the orphan receptors HNF-4, apoAI reglatory protein-1 (ARP-1), and v-erbA-related receptors (EAR) 3 and 2. Binding activity in rat l iver nuclear extracts includes these orphan receptors as judged from e lectromobility supershift experiments and from results obtained with e xpressed receptors, although the element in (CYP2C11 did not bind HNF- 4, Mutations of the HPF1-like elements in the CYP2C7, CYP2C11, and CYP 2C12 promoters had marginal effects on the expression of luciferase re porter gene constructs transiently transfected into HepG2 cells, where as for CYP2C13 the activity was reduced to 60% of the wild type constr uct, Coexpression of HNF-4 in COS-7 cells had limited effect on the lu ciferase activity generated from the 2C promoters, maximally 3 fold. O ur data indicate that the HPF1 elements in the rat CYP2C genes have li mited functional importance and that HNF-4 is not a major trans-activa tor for any of these genes.