ORPHAN NUCLEAR RECEPTOR HNF-4 BINDS TO THE HUMAN COAGULATION-FACTOR-VII PROMOTOR

The human coagulation protease factor VII plays a pivotal role in the initiation of the coagulation cascade by both the extrinsic and the intrinsic pathway, Although the gene, encoding factor MI, is expressed predominantly in the Liver, the mechanisms underlying this tissue-specific expression have not been elucidated, In this study, we have analyzed the contribution of 5 kilobases upstream of the ATG translational initiation codon upon hepatic factor MI gene transcription, Transient transfection assays of a set of nested deletions in both liver and non-liver cell lines, HepG2 and HeLa respectively, indicate that several regions are involved in liver-specific expression, A slight negative effect on factor VII promoter activity in HepG2 cells is mediated by sequences upstream of position -1212. DNase I protection experiments reveal six footprints, FPVII1 through FPVII6, within the proximal 714 base pairs but a minimal promoter of 165 base pairs containing only FPVII3-6 is sufficient to confer liver-specific expression in HepG2 cells, Interestingly, FPVII6, at position -14 to +10 on the sense strand, would indicate that an as yet unknown transcription factor covers the ATG translational initiation codon. Gel retardation experiments show that the liver-enriched transcription factor HNF-4 binds specifically to footprint FPVII4 at position -71 to -49, Furthermore, a T --> A transversion, that in the HNF-4 binding site of factor IX causes a severe bleeding disorder, was introduced into the HNF-4-binding site of factor VII and reduced promoter activity by 20-50%. Coordinate HNF-4-mediated regulation of several blood protease genes as well as genes involved in lipid metabolism might account for the positive correlation of these factors with increased risk of occlusive heart diseases.