Statins modulate transcriptional activity of heme-oxygenase-1 promoter in NIH 3T3 Cells.
Statins, inhibitors of HMG CoA reductase, have pleiotropic effects independent of their capacity to lower cholesterol. Heme-oxygenase-1(HO-1) plays an important role as an anti-oxidant and anti-inflammatory enzyme. In the present study, we used NIH 3T3 cells which express HO-1 to investigate the molecular mechanisms of HO-1 induction by statins. Simvastatin or fluvastatin induced a significant increase in HO-1 protein expression and mRNA levels. Both statins stimulated activity of a mouse HO-1 promoter (-1,287 to +73 bp)/luciferase reporter gene, 3.25 ± 0.23 (Mean ± S.E.M., n = 15, P < 0.001, t-test) and 3.13 ± 0.33 (Mean ± S.E.M., n = 6, P < 0.001, t-test), respectively. This effect was more pronounced in the short proximal promoter than the full promoter of HO-1. Gel retardation experiments for C/EBP and upstream stimulatory factor (USF) DNA-binding activities using simvastatin- or fluvastatin-treated cells showed significant nuclear protein-DNA complexes which were supershifted with antibodies specific for C/EBP β and δ or USF-1 and USF-2. Point mutations of the proximal HO-1 promoter (-149 to +73 bp) for the myc/max which binds USF or the C/EBP binding sequences showed a reduction in statin-induced reporter activity whereas no role of the distal C/EBP binding elements located at -4 kb was observed. Moreover, overexpression of mutated C/EBP β and USF factor or the siRNA for both factors supported a role of these transcription factors in statin-dependent induction of HO-1, with a clearer effect for C/EBP.
Department of Biochemistry and Molecular Genetics, American University of Beirut, Lebanon.
May F
MF
Mrad
Charbel A
CA
Mouawad
Moustafa
M
Al-Hariri
Assaad A
AA
Eid
Jawed
J
Alam
Aïda
A
Habib
D000818:Animals; D001665:Binding Sites; D022782:CCAAT-Enhancer-Binding Protein-beta / Q000235:genetics / Q000378:metabolism; D051743:CCAAT-Enhancer-Binding Protein-delta / Q000235:genetics / Q000378:metabolism; D005229:Fatty Acids, Monounsaturated / Q000494:pharmacology*; D000077340:Fluvastatin; D015870:Gene Expression / Q000187:drug effects; D017930:Genes, Reporter; D051547:Heme Oxygenase-1 / Q000235:genetics* / Q000378:metabolism; D007211:Indoles / Q000494:pharmacology*; D008156:Luciferases; D051379:Mice; D041681:NIH 3T3 Cells; D011401:Promoter Regions, Genetic; D011485:Protein Binding / Q000187:drug effects; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics*; D015398:Signal Transduction / Q000187:drug effects; D019821:Simvastatin / Q000494:pharmacology*; D015533:Transcriptional Activation / Q000187:drug effects; D014162:Transfection; D015854:Up-Regulation / Q000187:drug effects; D051720:Upstream Stimulatory Factors / Q000235:genetics / Q000378:metabolism
D022782:CCAAT-Enhancer-Binding Protein-beta; C545013:Cebpb protein, mouse; C497253:Cebpd protein, mouse; D005229:Fatty Acids, Monounsaturated; D007211:Indoles; D012333:RNA, Messenger; D051720:Upstream Stimulatory Factors; C497183:Usf1 protein, mouse; C497186:Usf2 protein, mouse; D051743:CCAAT-Enhancer-Binding Protein-delta; D000077340:Fluvastatin; D019821:Simvastatin; D008156:Luciferases; D051547:Heme Oxygenase-1