Cycloheximide induces nitric oxide synthase mRNA in vascular smooth muscle cells by prolonging mRNA lifetime.
Bacterial lipopolysaccharide (LPS) and other immunostimulants induce an isoform of NO synthase (iNOS) in vascular smooth muscle (VSM) which produces large quantities of NO and profound vasodilation; this process has been implicated as the cause of gram-negative septic shock. Although regulation of iNOS has been considered to occur at the level of transcription, it is unclear whether post-transcriptional events also contribute to changes in iNOS mRNA expression. We show that cycloheximide (CH), an inhibitor of protein synthesis, induces iNOS mRNA in VSM and potentiates the induction of iNOS mRNA caused by LPS. For many early-response genes, protein-synthesis inhibitors enhance mRNA levels by increasing mRNA stability. Since iNOS mRNA contains multiple copies of a consensus sequence (AUUUA) in common with these eary-response genes and responsible for mRNA destabilization, we tested whether CH induces iNOS mRNA by prolonging mRNA lifetime. In the absence of CH, iNOS mRNA decayed with biphagic kinetics and a half-life of 2 h. In the presence of CH, half-life was prolonged to approximately 7 h. Our observations indicate that in VSM the stability of iNOS mRNA may be under the control of a labile protein factor which awaits identification and characterization. Regulation of mRNA lifetime represents a novel site for control of iNOS gene expression.
Hattori
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Department of Endcrinology, Dokkyo University School of Medicine, Tochigi, Japan.
Gross
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SS
D000818:Animals; D001011:Aorta / Q000187:drug effects; D001483:Base Sequence; D002478:Cells, Cultured; D003513:Cycloheximide / Q000494:pharmacology*; D003609:Dactinomycin / Q000494:pharmacology; D004357:Drug Synergism; D006207:Half-Life; D008070:Lipopolysaccharides / Q000494:pharmacology*; D008297:Male; D008969:Molecular Sequence Data; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000235:genetics*; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology*; D012333:RNA, Messenger / Q000096:biosynthesis / Q000187:drug effects*; D051381:Rats; D017208:Rats, Wistar; D013997:Time Factors
D008070:Lipopolysaccharides; D011500:Protein Synthesis Inhibitors; D012333:RNA, Messenger; D003609:Dactinomycin; D003513:Cycloheximide; D019001:Nitric Oxide Synthase