Proteasome activation and nNOS down-regulation in neuroblastoma cells expressing a Cu,Zn superoxide dismutase mutant involved in familial ALS.
Reactive oxygen and nitrogen species have emerged as predominant effectors of neurodegeneration. We demonstrated that expression of the fully active G93A Cu,Zn superoxide dismutase mutant in neuroblastoma cells is associated with an increased level of oxidatively modified proteins, in terms of carbonylated residues. A parallel increase in proteasome activity was detected and this was mandatory in order to assure cell viability. In fact, proteasome inhibition by lactacystin or MG132 resulted in programmed cell death. Nitrosative stress was not involved in the oxidative unbalance, as a decrease in neuronal nitric oxide production and down-regulation of neuronal nitric oxide synthase (nNOS) level were detected. The nNOS down-regulation was correlated to increased proteolytic degradation by proteasome, because comparable levels of nNOS were detected in G93A and parental cells upon treatment with lactacystin. The altered rate of proteolysis observed in G93A cells was specific for nNOS as Cu,Zn superoxide dismutase (Cu,Zn SOD) degradation by proteasome was influenced neither by its mutation nor by increased proteasome activity. Treatment with the antioxidant 5,5'-dimethyl-1-pyrroline N-oxide resulted in inhibition of protein oxidation and decrease in proteasome activity to the basal levels. Overall these results confirm the pro-oxidant activity of G93A Cu,Zn SOD mutant and, at the same time, suggest a cross-talk between reactive oxygen and nitrogen species via the proteasome pathway.
Aquilano
Katia
K
Department of Biology, University of Rome Tor Vergata, Rome, Italy.
Rotilio
Giuseppe
G
Ciriolo
Maria Rosa
MR
D000690:Amyotrophic Lateral Sclerosis / Q000201:enzymology* / Q000235:genetics; D002470:Cell Survival / Q000502:physiology; D003497:Cyclic N-Oxides / Q000494:pharmacology; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D015536:Down-Regulation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D009097:Multienzyme Complexes / Q000037:antagonists & inhibitors / Q000378:metabolism*; D009154:Mutation; D009447:Neuroblastoma / Q000188:drug therapy / Q000201:enzymology*; D009569:Nitric Oxide / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000378:metabolism*; D052248:Nitric Oxide Synthase Type I; D010084:Oxidation-Reduction / Q000187:drug effects; D046988:Proteasome Endopeptidase Complex; D011506:Proteins / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D013482:Superoxide Dismutase / Q000096:biosynthesis* / Q000235:genetics; D014407:Tumor Cells, Cultured
D003497:Cyclic N-Oxides; D004791:Enzyme Inhibitors; D009097:Multienzyme Complexes; D011506:Proteins; D017382:Reactive Oxygen Species; D009569:Nitric Oxide; C017245:5,5-dimethyl-1-pyrroline-1-oxide; C496320:NOS1 protein, human; D019001:Nitric Oxide Synthase; D052248:Nitric Oxide Synthase Type I; C109415:SOD1 G93A protein; D013482:Superoxide Dismutase; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex