THE 3'-UNTRANSLATED REGION OF THE MOUSE CHOLESTEROL 7-ALPHA-HYDROXYLASE MESSENGER-RNA CONTAINS ELEMENTS RESPONSIVE TO POSTTRANSCRIPTIONAL REGULATION BY BILE-ACIDS

To investigate the importance of the 3'-untranslated region (UTR) of t he mouse cholesterol 7 alpha-hydroxylase (cyp7) mRNA in post-transcrip tional regulation of expression of the cyp7 gene, chimaeric genes enco ding mRNA containing the structural sequence of chloramphenicol acetyl transferase (CAT) linked to either the 3'-UTR of the mouse cyp7 mRNA o r the SV40 early gene mRNA were constructed. The human cytomegalovirus (CMV) promoter was used to drive the expression of all the chimaeric genes. Thus the transgenes had identical sequences in the promoter, th e regions encoding the 5'-UTR and translated sequence but differed in the region encoding the 3'-UTR of their respective mRNA species. The t ransgene containing the entire cyp7 3'-UTR (designated CMV.CAT.CYP7) g ave rise to CAT activity in transfected hepatoma cells that was one-qu arter of that obtained in cells transfected with the transgene contain ing the SV40 3'-UTR (designated CMV.CAT.SV40). The 3'-UTR of the cyp7 mRNA contains sequences resembling AU-rich elements (AREs). Deleting e ight of nine putative AREs from the CYP7 3'-UTR sequence increased the CAT activity to a level greater than that observed for CMV.CAT.SV40, whereas deletion of the intron region had no effect. These results sho w that the AREs of the 3'-UTR of the cyp7 mRNA decrease transgene expr ession. Bile acids are known to repress the expression of the cyp7 gen e. To test whether the 3'-UTR of the cyp7 mRNA has a role in this proc ess, the expression of the chimaeric genes was evaluated in hepatoma c ells competent for bile acid uptake. Conjugated bile acids, but not un conjugated bile acids, further decreased the expression of the CMV.CAT .CYP7 transgene. The same bile acids had no effect on the expression o f the CMV.CAT.SV40 transgene. Deletion of the intron from the cyp7 seq uence did not alter the CAT activity compared with the parental plasmi d, and also did not alter the sensitivity of the transgene to the conj ugated bile acids. Deletion of the AREs from the cyp7 3'-UTR, which in creased the expression of the transgene, did not abolish the sensitivi ty of the transgene to repression by conjugated bile acids. Thus the 3 '-UTR of the mouse cyp7 mRNA also contains elements that facilitate th e further repression of transgene expression in the presence of conjug ated bile acids. The results indicate that the 3'-UTR of the mouse cyp 7 mRNA contains information specifying regulation at the post-transcri ptional level.