role of disulfide cross-linking of mutant sod1 in the formation of inclusion-body-like structures二硫交联作用的突变sod1 inclusion-body-like结构的形成.pdfVIP

Role of Disulfide Cross-Linking of Mutant SOD1 in the Formation of Inclusion-Body-Like Structures . . Brittany L. T. Roberts , Kinaree Patel , Hilda H. Brown, David R. Borchelt* Department of Neuroscience, Center for Translational Research in Neurodegenerative Disease, SantaFe HealthCare Alzheimer’s Disease Research Center, McKnight Brain Institute, University of Florida, Gainesville, Florida, United States of America Abstract Background: Pathologic aggregates of superoxide dismutase 1 (SOD1) harboring mutations linked to familial amyotrophic lateral sclerosis (fALS) have been shown to contain aberrant intermolecular disulfide cross-links. In prior studies, we observed that intermolecular bonding was not necessary in the formation of detergent- insoluble SOD1 complexes by mutant SOD1, but we were unable to assess whether this type of bonding may be important for pathologic inclusion formation. In the present study, we visually assess the formation of large inclusions by fusing mutant SOD1 to yellow fluorescent protein (YFP). Methodology/Principal Findings: Experimental constructs possessing mutations at all cysteine residues in SOD1 (sites 6, 57, 111, and 146 to F,S,Y,R or G,S,Y,R, respectively) were shown to maintain a high propensity of inclusion formation despite the inability to form disulfide cross-links. Interestingly, although aggregates form when all cysteines were mutated, double mutants of the ALS mutation C6G with an experimental mutation C111S exhibited low aggregation propensity. Conclusions/Significance: Overall, this study is an extension of previous work demonstrating that cysteine residues in mutant SOD1 play a role in modulating aggregation and that intermolecular disulfi