Oestradiol decreases rat apolipoprotein AI transcription via promoter siteB

Oestrogens protect against ischaemic heart disease in the post-menopausal f emale by increasing serum concentrations of apolipoprotein (apo) AI and the abundance of high-density lipoprotein particles. In men and experimental m ale animals, the administration of oestrogen has variable effects on apo AI expression. As the major mode of oestrogen action on target genes involves regulating promoter activity and hence transcription, oestrogen is expecte d to alter transcription of the apo AI gene. To test this hypothesis, the e ffect of 17 beta -oestradiol (E-2), on rat apo AI promoter activity in male hepatoma HuH-7 cells, was tested by cotransfecting a reporter template, pA I.474.CAT containing - 474 to - 7 of the rat apo AI promoter and an oestrog en receptor (ER) expression vector, pCMV-ER. Transfected cells exposed to E 2 showed a dose-dependent decrease in chloramphenicol acetyltransferase (CA T)-activity, with a maximum 91 +/- 1.5% reduction at 1 muM E-2. Deletional analysis of the promoter localized the inhibitory effect of ER and E-2 to s ite B (-170 to - 144) with an adjacent 5' contiguous motif, site S (-186 to - 171) acting as an amplifier. HuH-7 cell nuclear extracts showed binding activities with both sites S and B, but recombinant human ER did not. Furth ermore, nuclear extracts from E-2-treated HuH-7 cells showed weaker binding activity to site B, but not to site S. In summary, the inhibitory effect o f ER and E-2 on rat apo AI gene activity is mediated by a promoter element, site B. This inhibitory effect arises from a mechanism that does not invol ve direct ER binding to the B-element. The conclusion that E-2 inhibits apo AI transcription was confirmed in vivo. Treatment of male adult Sprague-Da wley rats with up to 200 mug E-2 for 7 days decreased apo AI protein and he patic mRNA by 72 +/- 21% and 68 +/- 1.4% respectively. Results of 'run-on' transcription of the apo AI gene in isolated hepatic nuclei showed a 55% de crease in hormone-treated male rats. These findings suggest that E-2, exert s primarily an inhibitory effect within male hepatic nuclei.