Structure of the 5 '-flanking region of the rat prostaglandin F-2 alpha receptor gene and its transcriptional control functions in hepatocytes

Prostaglandin F-2 alpha (PGF(2 alpha)), modulates hepatocyte functions via a heptahelical G(q)-coupled PGF(2 alpha)-receptor (FP-R) which in liver is expressed exclusively in hepatocytes. The aim of the present study was to i solate the 5'-flanking region of the rat FP-R gene and to elucidate its bas al and IL-6-modulated transcription control function in rat hepatocytes. Th e 5'-nontranslated region of the rat hepatocyte FP-R mRNA differed from the corresponding region in rat fetal astrocyte or corpus luteum. It was encod ed by exons la and 2 which were separated by a 1.4 kb intron containing the exons Ib and Ic coding for the 5'-untranslated region of rat fetal astrocy te and corpus luteum FP-R mRNA, respectively. The transcription initiation site in hepatocytes was localized 263 bp upstream of the start ATG by 5'-RA CE. A DNA-fragment covering the 5'-flanking region of the rFP-R gene from - 1 of the transcription initiation site to -2590 bp was cloned and sequenced . Its 3'-two thirds had a 65% sequence identity to the mouse FP-R promoter however no homology to the bovine FP-R promoter. In the overlapping sequenc e most of the putative transcription factor binding sites were conserved be tween mouse and rat. The rat promoter contained no classical TATA- or CAAT- boxes but putative binding sites for the transcription factors C/EBP, GATA- 1, HNF-1, HNF-3 beta, SP-1, and USF. Luciferase reporter gene constructs co ntaining portions of the 5'-flanking region were transfected into rat hepat ocytes. Luciferase expression ranked -181 greater than or equal to -608 < - 1418 > -1821 greater than or equal to -2590. The strongest transcriptional activity was conferred by the region between -608 and -1418 containing a cl uster of potential HNF-1 and HNF-3 beta binding sites that might allow the exclusive expression of FP-R mRNA in hepatocytes. The amount of FP-R mRNA a nd the luciferase expression under control of the -2590 promoter fragment w ere reduced by IL-6 in hepatocytes. (C) 2000 Academic Press.