Repression of cholesterol 7 alpha-hydroxylase transcription by bile acids is mediated through protein kinase C in primary cultures of rat hepatocytes.

ABSTRACT

Inhibitors of protein kinases were screened for the ability to prevent the repression of cholesterol 7 alpha-hydroxylase mRNA by taurocholate in primary cultures of adult rat hepatocytes. The addition of taurocholate (25 microM) for 6 h decreased cholesterol 7 alpha-hydroxylase mRNA by 64 +/- 3%. However, after a preincubation with the protein kinase C inhibitors calphostin C or chelerythrine, taurocholate had no significant effect on cholesterol 7 alpha-hydroxylase mRNA, or decreased levels by only 23 +/- 8%, respectively. Protein kinase C activation with phorbol 12-myristate, 13-acetate (100 nM) decreased cholesterol 7 alpha-hydroxylase mRNA and transcriptional activity by 71 +/- 5% and 60 +/- 16%, respectively, within 3 h. mRNA levels recovered to control levels by 18-24 h, however, consistent with downregulation of protein kinase C. Furthermore, after depletion of protein kinase C with a 24-h preincubation with phorbol diesters, taurocholate (25 microM) repressed cholesterol 7 alpha-hydroxylase mRNA by only 14 +/- 17%. The addition of taurocholate (50 microM) to the culture medium transiently increased membrane-associated protein kinase C activity by approximately twofold, while causing a concomitant decrease in cytosolic activity. Other bile acids increased membrane-associated protein kinase C activity in approximate proportion to their relative hydrophobicity. Finally, immunoblotting experiments revealed translocation of the alpha isoform of protein kinase C to hepatocyte membranes in response to taurocholate. These data suggest that hydrophobic bile acids repress cholesterol 7 alpha-hydroxylase transcription through a protein kinase C-dependent mechanism.