CONSTITUTIVE EXPRESSION OF MURINE COMPLEMENT FACTOR-B GENE IS REGULATED BY THE INTERACTION OF ITS UPSTREAM PROMOTER WITH HEPATOCYTE NUCLEARFACTOR-4

Factor B (Bf) is a constituent of the alternative pathway of complemen t activation encoded within the major histocompatibility complex, Tran scription of the murine gene from two initiation sites generates two B f mRNA species differing in size and tissue distribution, Striking gen etic, tissue-specific differences in Bf mRNA levels at extrahepatic si tes (kidney and intestine) among mouse strains correlate with a DNA se quence polymorphism in the 5'-flanking region of the gene and differen tial nuclear protein binding at the Bf upstream transcriptional initia tion site (UIS), To ascertain the functional consequences of this poly morphism in the Bf promoter, we analyzed the effects of strain-specifi c sequences in the Bf 5' region on the expression of a chloramphenicol acetyltransferase (CAT) reporter gene transfected in human and mouse hepatoma cells, The CAT activity and mRNA level produced when transcri ption was driven by the sequence of strains with high extrahepatic exp ression were reduced to background levels when the sequence specific t o the low expressor strains was used, Eighty percent of this differenc e was accounted for by a point substitution that affects DNA-protein i nteraction at the UIS, the sequence of higher affinity conferring high er expression, Hepatocyte nuclear factor 4 (HNF-4), derived from HepG2 , mouse liver and kidney or cell-free translation of HNF-4 RNA, is the nuclear protein that preferentially binds to the high expressor UIS, Bf-CAT is not expressed in cells that lack HNF-4 (CV-1), However, co-t ransfection of HNF-4 into CV-I cells drives Bf-CAT expression and repr oduces the differences derived from the substitution that affect HNF-4 binding in vitro. These data show that interaction of HNF-4 with poly morphic variants of the upstream Bf promoter is the major determinant of strain-specific extrahepatic factor B expression.