Proteasomal inhibition by misfolded mutant superoxide dismutase 1 induces selective motor neuron death in familial amyotrophic lateral sclerosis

Abstract

Accumulating evidence indicates that abnormal conformation of mutant superoxide dismutase 1 (SOD1) is an essential feature underlying the pathogenesis of mutant SOD1‐linked familial amyotrophic lateral sclerosis (ALS). Here we investigated the role of ubiquitin‐proteasome pathway in the mutant SOD1‐related cell death and the effect of oxidative stress on the misfolding of mutant SOD1. Transient overexpression of ubiquitin with human SOD1 (wild‐type, ala4val, gly85arg, gly93ala) in Neuro2A cells decreased the amount of mutant SOD1, but not of wild‐type, while only mutants were co‐immunoprecipitated with poly‐ubiquitin. Proteasome inhibition by lactacystin augmented accumulation of mutant SOD1 in the non‐ionic detergent‐insoluble fraction. The spinal cord lysates from mutant SOD1 transgenic mice showed multiple carbonylated proteins, including mutant SOD1 with SDS‐resistant dimer formation. Furthermore, the treatment of hSOD1‐expressing cells with hydrogen peroxide promoted the oligomerization, and detergent‐insolubility of mutant SOD1 alone, and the oxidized mutant SOD1 proteins were more heavily poly‐ubiquitinated. In Neuro2A cells stably expressing human SOD1 protein, the proteasome function measured by chymotrypsin‐like activity, was decreased over time without a quantitative alteration of the 20S proteasomal component. Finally, primary motor neurons from the mouse embryonic spinal cord were more vulnerable to lactacystin than non‐motor neurons. These results indicate that the sustained expression of mutant SOD1 leads to proteasomal inhibition and motor neuronal death, which in part explains the pathogenesis of mutant SOD1‐linked ALS.