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.