Nitric oxide-dependent suppression of thioredoxin-interacting protein expression enhances thioredoxin activity

Objective - Cellular redox balance is regulated by enzymatic and nonenzymatic systems and freely diffusible nitric oxide ( NO) promotes antioxidative mechanisms. We show the NO-dependent transcriptional regulation of the antioxidative thioredoxin system. Methods and Results - Incubation of rat pulmonary artery smooth muscle cells (RPaSMC) with the NO donor compound S-nitroso-glutathione (GSNO, 100 mu mol/L) suppressed thioredoxin-interacting protein (Txnip), an inhibitor of thioredoxin function, by 71 +/- 18% and enhanced thioredoxin reductase 2.7 +/- 0.2 fold (n = 6; both P < 0.001 versus control). GSNO increased thioredoxin activity (1.9 +/- 0.5-fold after 4 hours; P < 0.05 versus control). Promoter deletion analysis revealed that NO suppression of Txnip transcription is mediated by cis-regulatory elements between -1777 and -1127 bp upstream of the start codon. Hyperglycemia induced Txnip promoter activity (3.9 +/- 0.2-fold; P < 0.001) and abolished NO effects (-37.4 +/- 1.0% at 5.6 mmol/L glucose versus 12.4 +/- 2.1% at 22.4 mmol/L glucose; P < 0.05). Immunoprecipitation experiments demonstrated that GSNO stimulation and mutation of thioredoxin at Cys69, a site of nitrosylation, had no effect on the Txnip/thioredoxin interaction. Conclusions - NO can regulate cellular redox state by changing expression of Txnip and thioredoxin reductase. This represents a novel antioxidative mechanism of NO independent of posttranslational protein S-nitrosylation of thioredoxin.