title: Cysteine 111 affects aggregation and cytotoxicity of mutant Cu,Zn-superoxide dismutase associated with familial amyotrophic lateral sclerosis.
abstract: Converging evidence indicates that aberrant aggregation of mutant Cu,Zn-superoxide dismutase (mutSOD1) is strongly implicated in familial amyotrophic lateral sclerosis (FALS). MutSOD1 forms high molecular weight oligomers, which disappear under reducing conditions, both in neural tissues of FALS transgenic mice and in transfected cultured cells, indicating a role for aberrant intermolecular disulfide cross-linking in the oligomerization and aggregation process. To study the contribution of specific cysteines in the mechanism of aggregation, we mutated human SOD1 in each of its four cysteine residues and, using a cell transfection assay, analyzed the solubility and aggregation of those SOD1s. Our results suggest that the formation of mutSOD1 aggregates are the consequence of covalent disulfide cross-linking and non-covalent interactions. In particular, we found that the removal of Cys-111 strongly reduces the ability of a range of different FALS-associated mutSOD1s to form aggregates and impair cell viability in cultured NSC-34 cells. Moreover, the removal of Cys-111 impairs the ability of mutSOD1s to form disulfide cross-linking. Treatments that deplete the cellular pool of GSH exacerbate mutSOD1s insolubility, whereas an overload of intracellular GSH or overexpression of glutaredoxin-1, which specifically catalyzes the reduction of protein-SSG-mixed disulfides, significantly rescues mutSOD1s solubility. These data are consistent with the view that the redox environment influences the oligomerization/aggregation pathway of mutSOD1 and point to Cys-111 as a key mediator of this process.
authors.affiliation: Laboratory of Neurochemistry, Fondazione S. Lucia Istituto di Ricovero e Cura a Carattere Scientifico, Rome 00179, Italy.
authors.forename: Mauro
authors.initials: M
authors.lastname: Cozzolino
authors.forename: Ilaria
authors.initials: I
authors.lastname: Amori
authors.forename: Maria Grazia
authors.initials: MG
authors.lastname: Pesaresi
authors.forename: Alberto
authors.initials: A
authors.lastname: Ferri
authors.forename: Monica
authors.initials: M
authors.lastname: Nencini
authors.forename: Maria Teresa
authors.initials: MT
authors.lastname: Carrì
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D001921:Brain / Q000201:enzymology; D003545:Cysteine; D017931:DNA Primers; D018076:DNA, Complementary / Q000235:genetics; D006801:Humans; D051379:Mice; D016472:Motor Neuron Disease / Q000201:enzymology / Q000235:genetics*; D009154:Mutation; D010641:Phenotype; D010957:Plasmids; D020641:Polymorphism, Single Nucleotide; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013482:Superoxide Dismutase / Q000737:chemistry / Q000235:genetics*; D000072105:Superoxide Dismutase-1
chemical_list: D017931:DNA Primers; D018076:DNA, Complementary; D011994:Recombinant Proteins; C000606290:SOD1 protein, human; C000606291:Sod1 protein, mouse; D013482:Superoxide Dismutase; D000072105:Superoxide Dismutase-1; D003545:Cysteine