The Role of Nitric Oxide Synthase in the Regulation of Ultraviolet Light-induced Phosphorylation

Ultraviolet light (UV) induces phosphorylation of the alpha subunit of the eukaryotic initiation factor 2 (eIF2α) and inhibits global protein synthesis. Both eIF2 kinases, PERK and GCN2, have been shown to phosphorylate eIF2α in response to UV-irradiation. However, the roles of PERK and GCN2 in UV-induced eIF2α phosphorylation are controversial. The upstream signaling pathway(s) that leads to the activation of PERK or GCN2 remains unknown. In this report we provide data showing that both PERK and GCN2 contribute to UV-induced eIF2α phosphorylation in human keratinocyte (HaCaT) and mouse embryonic fibroblast (MEF) cells. Reduction of expression of PERK or GCN2 by small interfering RNA decreases phosphorylation of eIF2α after UV-irradiation. The data also show that nitric oxide synthase (NOS)-mediated oxidative stress plays a role in regulation of eIF2α phosphorylation upon UV-irradiation. Treating the cells with the broad NOS inhibitor N-methyl-L-arginine, the free radical scavenger N-acetyl-L-cysteine or the NOS substrate L-arginine partially inhibits UV-induced eIF2α phosphorylation. The results presented above led us to propose that NOS mediates UV-induced eIF2α phosphorylation by activation of both PERK and GCN2 via oxidative-stress and L-arginine starvation signaling pathways.