An Octamer Motif Is Required for Activation of the Inducible Nitric Oxide Synthase Promoter in Pancreatic β-Cells

Abstract

Nitric oxide, generated by the inducible form of nitric oxide synthase (iNOS), is a potential mediator of cytokine-induced β-cell dysfunction in type 1 diabetes mellitus. We have previously shown that cytokine-induced iNOS expression is cycloheximide (CHX) sensitive and requires nuclear factor-κB (NF-κB) activation. In the present study, we show that an octamer motif located 20 bp downstream of the proximal NF-κB binding site in the rat iNOS promoter is critical for IL-1β and interferon-γ induction of promoter activity in rat primary β-cells and insulin-producing RINm5F cells. In gel shift assays, the octamer motif bound constitutively the transcription factor Oct1. Neither Oct1 nor NF-κB binding activities were blocked by CHX, suggesting that other factor(s) synthesized in response to IL-1β 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-1β-induced iNOS promoter activity. These results suggest that Oct1, NF-κB, and HMG-I(Y) cooperate for transactivation of the iNOS promoter in pancreatic β-cells.