Upregulation of immunoproteasomes by nitric oxide: Potential antioxidative mechanism in endothelial cells

Nitric oxide (^•NO) was shown to stimulate the proteasomal function and the ubiquitin-proteasome pathway and to ameliorate endothelial apoptotic signaling induced by oxidants. Understanding the regulatory mechanisms by which ^•NO stimulates proteasomes and affords cytoprotection in endothelial cells has therapeutic implications, as many vascular diseases are characterized by a deficiency in ^•NO. Here we report that ^•NO/cGMP/cAMP-induced immunoproteasome subunit expression is responsible for the increased proteasomal activities. Cells pretreated with protein kinase G and protein kinase A inhibitors markedly attenuated ^•NO-dependent proteasome activation. Results show that the ^•NO/cGMP/cAMP signaling mechanism enhanced the phosphorylation of the transcription factor cAMP-response element-binding protein, elevated the cAMP-response element-promoter activity and induced the expression of immunoproteasomal subunits (LMP2 and LMP7). ^•NO-dependent proteasomal activity was abrogated in cells transfected with antisense LMP2 and LMP7 oligonucleotides. Lower levels of LMP2 and LMP7 were detected in aorta of iNOS^-/- mice compared to wild-type controls, suggesting that endogenous production of ^•NO is important in the basal regulation of immunoproteasome. The ^•NO/cGMP/cAMP signaling pathway mitigates transferrin-iron-mediated oxidative stress and apoptosis through induction of immunoproteasomes. These results provide new insights on the regulatory mechanisms by which the ^•NO-mediated immunoproteasome signaling pathway affords cytoprotection in endothelial cells.