Post-transcriptional regulation of endothelial nitric oxide synthase mRNA stability by Rho GTPase.
The mechanism by which 3-hydroxy-3-methylglutaryl (HMG)-CoA reductase inhibitors increase endothelial nitric oxide synthase (eNOS) expression is unknown. To determine whether changes in isoprenoid synthesis affects eNOS expression, human endothelial cells were treated with the HMG-CoA reductase inhibitor, mevastatin (1-10 microM), in the presence of L-mevalonate (200 microM), geranylgeranylpyrophosphate (GGPP, 1-10 microM), farnesylpyrophosphate (FPP, 5-10 microM), or low density lipoprotein (LDL, 1 mg/ml). Mevastatin increased eNOS mRNA and protein levels by 305 +/- 15% and 180 +/- 11%, respectively. Co-treatment with L-mevalonate or GGPP, but not FPP or LDL, reversed mevastatin's effects. Because Rho GTPases undergo geranylgeranyl modification, we investigated whether Rho regulates eNOS expression. Immunoblot analyses and [35S]GTPgammaS-binding assays revealed that mevastatin inhibited Rho membrane translocation and GTP binding activity by 60 +/- 5% and 78 +/- 6%, both of which were reversed by co-treatment with GGPP but not FPP. Furthermore, inhibition of Rho by Clostridium botulinum C3 transferase (50 microg/ml) or by overexpression of a dominant-negative N19RhoA mutant increased eNOS expression. In contrast, activation of Rho by Escherichia coli cytotoxic necrotizing factor-1 (200 ng/ml) decreased eNOS expression. These findings indicate that Rho negatively regulates eNOS expression and that HMG-CoA reductase inhibitors up-regulate eNOS expression by blocking Rho geranylgeranylation, which is necessary for its membrane-associated activity.
Laufs
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Cardiovascular Division, Brigham & Women's Hospital and Harvard Medical School, Boston, Massachusetts 02115, USA.
Liao
J K
JK
D036002:ADP Ribose Transferases / Q000378:metabolism / Q000494:pharmacology; D001427:Bacterial Toxins / Q000494:pharmacology; D001905:Botulinum Toxins; D002478:Cells, Cultured; D003600:Cytosol / Q000378:metabolism; D003603:Cytotoxins / Q000494:pharmacology; D004730:Endothelium, Vascular / Q000201:enzymology*; D029968:Escherichia coli Proteins; D020558:GTP Phosphohydrolases / Q000235:genetics / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism; D006801:Humans; D019161:Hydroxymethylglutaryl-CoA Reductase Inhibitors / Q000494:pharmacology; D007700:Kinetics; D008077:Lipoproteins, LDL / Q000494:pharmacology / Q000502:physiology; D008148:Lovastatin / Q000031:analogs & derivatives / Q000494:pharmacology; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D008798:Mevalonic Acid / Q000494:pharmacology; D019001:Nitric Oxide Synthase / Q000235:genetics*; D052250:Nitric Oxide Synthase Type III; D011106:Polyisoprenyl Phosphates / Q000494:pharmacology; D012323:RNA Processing, Post-Transcriptional / Q000187:drug effects; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*; D012717:Sesquiterpenes; D018631:ras Proteins / Q000378:metabolism; D020742:rhoA GTP-Binding Protein; D020743:rhoB GTP-Binding Protein
D001427:Bacterial Toxins; D003603:Cytotoxins; D029968:Escherichia coli Proteins; D019161:Hydroxymethylglutaryl-CoA Reductase Inhibitors; D008077:Lipoproteins, LDL; D008565:Membrane Proteins; D011106:Polyisoprenyl Phosphates; D012333:RNA, Messenger; D012717:Sesquiterpenes; C060296:cytotoxic necrotizing factor type 1; C012258:mevastatin; D016244:Guanosine 5'-O-(3-Thiotriphosphate); C004808:farnesyl pyrophosphate; D006160:Guanosine Triphosphate; D008148:Lovastatin; C496314:NOS3 protein, human; D019001:Nitric Oxide Synthase; D052250:Nitric Oxide Synthase Type III; D036002:ADP Ribose Transferases; C063007:exoenzyme C3, Clostridium botulinum; D001905:Botulinum Toxins; D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins; D018631:ras Proteins; D020742:rhoA GTP-Binding Protein; D020743:rhoB GTP-Binding Protein; C002963:geranylgeranyl pyrophosphate; D008798:Mevalonic Acid