Transcriptional inhibition of the inducible nitric oxide synthase gene by competitive binding of NF-kappa B/Rel proteins.
The activity of the inducible nitric oxide synthase enzyme (iNOS) is tightly controlled, partly at the transcriptional level. We find NF-kappa B/Rel activation (p50-p50 and p50-p65) in RAW 264.7 macrophages after lipopolysaccharide treatment and binding to both NF-kappa B sites in the mouse iNOS promoter. To delineate the importance of NF-kappa B/Rel in iNOS gene transcription, we used an unusually direct approach to try to improve on the antioxidant-treatment or reporter techniques, namely the depletion of NF-kappa B/Rel activity through the use of a phosphorothioate-modified oligonucleotide containing three copies of the NF-kappa B consensus sequence. The reduction in NF-kappa B/Rel activity (particularly that binding to the downstream of the two sites) was associated with a 50% reduction in NO output and a reduction in the quantity of the iNOS protein expressed. These results point to the probability that physiologically relevant NF-kappa B/Rel activators or repressors other than lipopolysaccharide might crucially affect the macrophage NO response.
Goldring
C E
CE
Laboratoire de l'Ecole Pratique des Hautes Etudes, EPHE, Faculté de Médecine, Dijon, France.
Narayanan
R
R
Lagadec
P
P
Jeannin
J F
JF
D000594:Amino Acid Oxidoreductases / Q000235:genetics* / Q000378:metabolism; D000818:Animals; D001483:Base Sequence; D001667:Binding, Competitive; D015971:Gene Expression Regulation, Enzymologic; D051379:Mice; D008969:Molecular Sequence Data; D016328:NF-kappa B / Q000378:metabolism*; D052002:NF-kappa B p50 Subunit; D019001:Nitric Oxide Synthase; D009838:Oligodeoxyribonucleotides; D051996:Transcription Factor RelA; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic
D016328:NF-kappa B; D052002:NF-kappa B p50 Subunit; D009838:Oligodeoxyribonucleotides; D051996:Transcription Factor RelA; D014157:Transcription Factors; D019001:Nitric Oxide Synthase; D000594:Amino Acid Oxidoreductases