3'-UNTRANSLATED SEQUENCES MEDIATE POSTTRANSCRIPTIONAL REGULATION OF 3-HYDROXY-3-METHYLGLUTARYL-COA REDUCTASE MESSENGER-RNA BY 25-HYDROXYCHOLESTEROL

In an earlier study [Choi, Lundquist and Peffley (1993) Biochem. J. 29 6, 859-866], we determined that 25-hydroxycholesterol regulates 3-hydr oxy-3-methylglutaryl-CoA (HMG-CoA) reductase mRNA through a post-trans criptional mechanism that requires protein synthesis. To investigate w hether 3'-untranslated sequences play a role in 25-hydroxycholesterol- mediated post transcriptional control, we ligated approx. 1400 bp of t he 3'-untranslated region of HMG-CoA reductase cDNA to the coding regi on of human beta-globin DNA. beta-Globin-3'-untranslated reductase fus ion constructs were then transiently expressed in Chinese hamster ovar y fibroblasts under conditions known to regulate reductase mRNA. There were no differences in beta-globin RNA levels in transfected cells in cubated with or without lovastatin, a competitive inhibitor of reducta se. However, in the presence of lovastatin and an oxysterol, 25-hydrox ycholesterol, beta-globin RNA levels were decreased approx. 2-fold. In hibition of protein synthesis with cycloheximide blocked the effects o f 25-hydroxycholesterol on beta-globin RNA. Moreover, replacing the 3' -untranslated sequences with 1367 bp of the simian virus 40 enhancer r egion eliminated the regulatory effect of 25-hydroxycholesterol. Becau se the fusion construct has no sterol regulatory elements necessary fo r transcription, our results indicate that the change in beta-globin R NA occurred at a post-transcriptional level. In addition, we have show n that the 3'-untranslated region of HMG-CoA reductase cDNA imparted o xysterol-mediated posttranscriptional regulation to beta-globin RNA, a n effect that required protein synthesis.