Molecular mechanisms of iNOS induction by IL-1 beta and IFN-gamma in rat aortic smooth muscle cells.
In rat aortic smooth muscle cells (RASMC), interferon (IFN)-gamma enhanced nitrite accumulation and type II nitric oxide synthase (iNOS) protein expression induced by interleukin (IL)-1 beta. IFN-gamma alone had no effect on nitrite accumulation or iNOS protein. IL-1 beta, but not IFN-gamma, induced nuclear factor (NF)-kappa B and CCAAT box/enhancer binding protein (C/EBP) nuclear binding. Conversely, IFN-gamma, but not IL-1 beta, induced signal transducer and activator of transcription (STAT) 1 and interferon regulatory factor (IRF)-1 binding. In a -1.4-kb rat iNOS promoter segment, deletion of an IFN-gamma-activated site (GAS) increased IL-1 beta-induced activity but inhibited IFN-gamma-enhanced activity, suggesting a two-way effect of the GAS site on iNOS induction: enhancing induction through STAT1 activation and inhibiting induction through a non-IFN-gamma-mediated mechanism. Deletion of both an IRF and a C/EBP site reduced the IL-1 beta-induced and the IFN-gamma-enhanced activities. However, IRF site mutations decreased the IFN-gamma-enhanced activity without affecting the IL-1 beta-induced activity. Insertion of two IRF sites increased the IFN-gamma-enhanced, but not the IL-1 beta-induced, activity. Mutations of a reverse NF-kappa B site did not significantly change IFN-gamma-enhanced activity. We conclude that in RASMC, NF-kappa B and C/EBP mediate the IL-1 beta-induced iNOS expression, whereas IRF-1 and STAT1 mediate the IFN-gamma-enhanced iNOS induction.
Teng
Xingwu
X
Vascular Biology Center and Department of Pharmacology and Toxicology, Medical College of Georgia, Augusta, Georgia 30912-2500, USA.
Zhang
Hanfang
H
Snead
Connie
C
Catravas
John D
JD
D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D001011:Aorta / Q000166:cytology; D001483:Base Sequence; D022762:CCAAT-Enhancer-Binding Proteins / Q000378:metabolism; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects / Q000502:physiology*; D050856:Interferon Regulatory Factor-1; D007371:Interferon-gamma / Q000494:pharmacology*; D007375:Interleukin-1 / Q000494:pharmacology*; D008969:Molecular Sequence Data; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000201:enzymology*; D016296:Mutagenesis / Q000502:physiology; D016328:NF-kappa B / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000235:genetics*; D052247:Nitric Oxide Synthase Type II; D010750:Phosphoproteins / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000502:physiology; D051381:Rats; D017208:Rats, Wistar; D050794:STAT1 Transcription Factor; D015534:Trans-Activators / Q000378:metabolism; D014162:Transfection
D000970:Antineoplastic Agents; D022762:CCAAT-Enhancer-Binding Proteins; D017931:DNA Primers; D004268:DNA-Binding Proteins; D050856:Interferon Regulatory Factor-1; D007375:Interleukin-1; C494259:Irf1 protein, rat; D016328:NF-kappa B; D010750:Phosphoproteins; D050794:STAT1 Transcription Factor; C494081:Stat1 protein, rat; D015534:Trans-Activators; D007371:Interferon-gamma; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496318:Nos2 protein, rat