Lead-induced Neurotoxicity is Modulated by Phosphorylation of Neuronal Nitric Oxide Synthase by PKA and CaM-KII

초록

This study aimed to identify the neuroprotective mechanism by the phosphorylation of neuronal nitric oxide synthase (nNOS) against Pb-caused cytotoxicity. We have used a neuroblastoma dopaminergic cell line, SH-SY5Y that has constitutive Ca++-dependent nNOS activity and thus produces NO spontaneously. Treatment of these cells with high concentration (100 mM) of lead (Pb-acetate) led to the phosphorylation of nNOS at Ser847, as determined by Western-blot analysis with a selective anti-phospho-nNOS antibody. This phosphoryltion was accompanied by the decreased reductase activity and mediated by activation of protein kinase A (PKA), PKC and/or calcium-calmodulin-dependent kinase II (CaM-KII). Pretreatment of these cells with selective inhibitor of PKA (H-89) or CaM-KII (KN-93) restored NOS activity, while PKC inhibitor (Ro-31-8220) did not affect. The results suggested that CaM-KII and/or PKA might negatively regulate nNOS to modulate NO production as neuroprotective mechanism against the overproduced NO and/or oxidative stress in brain.