Transcriptional derepression of the murine Cyp1a-1 gene by mevinolin.
In mouse hepatoma Hepa-1c1c7 cultures, polycyclic aromatic compounds such as benzol[a]pyrene and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD; dioxin) activate the Cyp1a-1 (cytochrome P(1)450) and Nmo-1[NAD(P)H:menadione-oxidoreductase] genes, two members of the aromatic hydrocarbon (Ah)-responsive gene battery. Mevinolin is known to inhibit 3-hydroxy-3-methylglutaryl CoA (HMG-CoA) reductase (EC 1.1.1.34), the rate-limiting step in cholesterol biosynthesis. We show here that in the absence of TCDD, mevinolin markedly increases Cyp1a-1 transcription, CYP1A1 mRNA and protein levels and enzyme activity, and NMO1 mRNA concentrations. Addition of mevalonate, the product of HMG-CoA reductase activity, fails to reverse the effects of mevinolin. In fact, when used at high concentrations, mevalonate activates Cyp1a-1 transcription. Mevinolin-induced Cyp1a-1 gene activation: (1) occurs independently of the lipid content of the growth medium, (2) is not suppressed by adding 25-hydroxycholesterol, which blocks MHG-CoA reductase activity, and (3) requires a functional Ah receptor and unimpaired nuclear translocation of the receptor. It is possible that an unknown metabolite (or metabolites) of mevinolin activates Cyp1a-1 expression and that high concentrations of mevalonate act via the same mechanism. Using chimaeric plasmids that contain different lengths of Cyp1a-1 5' flanking regions fused to the bacterial neomycin (neo) gene, we find that the mevinolin effect on Cyp1a-1 induction requires the 5' flanking sequences between -1647 and -824, which are also needed for TCDD induction. Mevinolin, however, is not a ligand for the Ah receptor. Gel mobility shift assays revealed that Cyp1a-1 activation caused by mevinolin does not involve the ligand-dependent formation of a functional Ah receptor-dependent DNA-binding complex, but instead appears to be correlated with release of a putative repressor from its cognate DNA site. Our results suggest that the basel level of Cyp1a-1 transcription is maintained by an unknown negative regulatory factor. We propose that Cyp1a-1 transcriptional activation can result not only from induction by polycyclic aromatic compounds but also from derepression by mevinolin, independent of HMG-CoA reductase inhibition.
Puga
A
A
Laboratory of Developmental Pharmacology, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland 20892.
Raychaudhuri
B
B
Nebert
D W
DW
D000818:Animals; D002460:Cell Line; D002467:Cell Nucleus / Q000502:physiology; D003513:Cycloheximide / Q000494:pharmacology; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D015342:DNA Probes; D004789:Enzyme Activation; D004794:Enzyme Repression; D019161:Hydroxymethylglutaryl-CoA Reductase Inhibitors; D007700:Kinetics; D008114:Liver Neoplasms, Experimental; D008148:Lovastatin / Q000494:pharmacology*; D008798:Mevalonic Acid / Q000494:pharmacology; D051379:Mice; D010957:Plasmids; D000072317:Polychlorinated Dibenzodioxins / Q000494:pharmacology; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D018336:Receptors, Aryl Hydrocarbon; D011955:Receptors, Drug / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects*; D014162:Transfection
D015342:DNA Probes; D019161:Hydroxymethylglutaryl-CoA Reductase Inhibitors; D000072317:Polychlorinated Dibenzodioxins; D012333:RNA, Messenger; D018336:Receptors, Aryl Hydrocarbon; D011955:Receptors, Drug; D003577:Cytochrome P-450 Enzyme System; D003513:Cycloheximide; D008148:Lovastatin; D008798:Mevalonic Acid