An octamer motif is required for activation of the inducible nitric oxide synthase promoter in pancreatic beta-cells.
Nitric oxide, generated by the inducible form of nitric oxide synthase (iNOS), is a potential mediator of cytokine-induced beta-cell dysfunction in type 1 diabetes mellitus. We have previously shown that cytokine-induced iNOS expression is cycloheximide (CHX) sensitive and requires nuclear factor-kappa B (NF-kappa B) activation. In the present study, we show that an octamer motif located 20 bp downstream of the proximal NF-kappa B binding site in the rat iNOS promoter is critical for IL-1 beta and interferon-gamma induction of promoter activity in rat primary beta-cells and insulin-producing RINm5F cells. In gel shift assays, the octamer motif bound constitutively the transcription factor Oct1. Neither Oct1 nor NF-kappa B binding activities were blocked by CHX, suggesting that other factor(s) synthesized in response to IL-1 beta contribute to iNOS promoter induction. The high mobility group (HMG)-I(Y) protein also bound the proximal iNOS promoter region. HMG-I(Y) binding was decreased in cells treated with CHX and HMG-I(Y) silencing by RNA interference reduced IL-1 beta-induced iNOS promoter activity. These results suggest that Oct1, NF-kappa B, and HMG-I(Y) cooperate for transactivation of the iNOS promoter in pancreatic beta-cells.
Darville
Martine I
MI
Laboratory of Experimental Medicine, Université libre de Bruxelles, B-1070 Brussels, Belgium. mdarvill@ulb.ac.be
Terryn
Sara
S
Eizirik
Décio L
DL
D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology; D045744:Cell Line, Tumor; D004268:DNA-Binding Proteins / Q000378:metabolism; D005434:Flow Cytometry; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects / Q000502:physiology; D020868:Gene Silencing; D024243:HMGB1 Protein / Q000235:genetics / Q000378:metabolism; D051863:Host Cell Factor C1; D007340:Insulinoma; D007371:Interferon-gamma / Q000494:pharmacology; D007375:Interleukin-1 / Q000494:pharmacology; D007515:Islets of Langerhans / Q000502:physiology*; D016296:Mutagenesis; D016328:NF-kappa B / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000235:genetics*; D052247:Nitric Oxide Synthase Type II; D050811:Octamer Transcription Factor-1; D011401:Promoter Regions, Genetic / Q000502:physiology*; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D014157:Transcription Factors / Q000378:metabolism
D000970:Antineoplastic Agents; D004268:DNA-Binding Proteins; D024243:HMGB1 Protein; D051863:Host Cell Factor C1; D007375:Interleukin-1; D016328:NF-kappa B; D050811:Octamer Transcription Factor-1; C494168:Pou2f1 protein, rat; D012333:RNA, Messenger; D014157:Transcription Factors; D007371:Interferon-gamma; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496318:Nos2 protein, rat