Transcriptional regulation of the human iNOS gene in vascular-smooth-muscle cells and macrophages: evidence for tissue specificity.
We have cloned the 5' upstream -1034 to +88 fragment of the human inducible nitric oxide synthase (hiNOS) gene and demonstrate its competence to promote luciferase gene transcription in vascular-smooth-muscle (VSM) cells and macrophages. Sequential 5' end-deletions localized positive regulatory elements of hiNOS transcription in VSM A7r5 cells downstream of nucleotide -205 and demonstrated the functional importance of the resident NF-kappaB site (nucleotides -115 to -106). The hiNOS promoter/enhancer was induced strongly by LPS and IFN-gamma, and modestly by IL-1beta in RAW 264.7 cells, but not in VSM cells. Truncation of the NF-kappaB site markedly diminished, but did not eliminate, LPS-inducibility. Sodium salicylate and ibuprofen down-regulated the basal transcriptional activity of the hiNOS promoter/enhancer in VSM but not in RAW 264.7 cells. These results indicate that the transcriptional regulation of the hiNOS gene features considerable complexity and tissue specificity.
Kolyada
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AY
Department of Medicine, Tufts University School of Medicine, Boston, Massachusetts 02111, USA.
Savikovsky
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N
Madias
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NE
D000818:Animals; D001011:Aorta; D001483:Base Sequence; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D017931:DNA Primers; D004742:Enhancer Elements, Genetic; D015870:Gene Expression / Q000187:drug effects; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D007052:Ibuprofen / Q000494:pharmacology; D007527:Isoenzymes / Q000096:biosynthesis; D008070:Lipopolysaccharides / Q000494:pharmacology; D008156:Luciferases / Q000096:biosynthesis; D008264:Macrophages / Q000201:enzymology*; D051379:Mice; D008969:Molecular Sequence Data; D009131:Muscle, Smooth, Vascular / Q000201:enzymology*; D019001:Nitric Oxide Synthase / Q000096:biosynthesis*; D009928:Organ Specificity; D011401:Promoter Regions, Genetic; D051381:Rats; D017207:Rats, Sprague-Dawley; D011994:Recombinant Proteins / Q000096:biosynthesis; D015183:Restriction Mapping; D012501:Saphenous Vein; D012980:Sodium Salicylate / Q000494:pharmacology; D014158:Transcription, Genetic; D014162:Transfection
D017931:DNA Primers; D007527:Isoenzymes; D008070:Lipopolysaccharides; D011994:Recombinant Proteins; D008156:Luciferases; D019001:Nitric Oxide Synthase; D012980:Sodium Salicylate; D007052:Ibuprofen