Aggregation and fibrillation of SODl have been implicated in disease mechanisms of Amyotrophic Lateral Sclerosis (ALS), and it is a major new goal of this Program Project renewal to develop better biological assays to study the toxicity of these multimeric forms of SODl in systems that will be more relevant to the disease in humans. In Project 2 we further develop and use a human cell culture system that closely models important cell biological aspects of motor neuron degeneration?our recently developed human embryonic stem cell-derived motor neuron (HESC-MN) system. The cells have distinct advantages over other model systems as they represent the major cell type that degenerates in ALS and they are fully human. The cells express identifying neuronal markers, exhibit electrophysiological function typical for mature motor neurons, and can be co-cultured with other neuronal and non-neuronal cells. Transfection of these cells to express ALS-SODl proteins causes deleterious effeds on cell survival and morphology. Importantly for this project, we have recently shown that exogenously added ALS-SODl protein multimers are taken up quite well. We will utilize these cells to study the toxicity of SODl protein multimers and aggregates at different stages of their formation and relate it to the progression of motor neuron degeneration. This research plan outlines a highly collaborative, step-by-step approach to evaluate spontaneous and induced mutant and WT SODl aggregate formation in motor neurons, followed by an investigation of the consequences of SODl aggregates on neurodegenerative mechanisms and, finally, by using pharmacological inhibitors of SODl aggregation to investigate whether reduced SODl aggregation can prevent motor neuron death.
Functional HESC-MNs expressing ALS-SODl mutant proteins provide an unparalleled opportunity to explore why various multimeric forms of mutant SODl are toxic to motor neurons. The project will move from the urgently needed improved understanding of intracellular SODl multimerization as a disease pathway of ALS to defining a new disease model system for designing and testing new ALS therapeutics.
|Ayers, Jacob I; McMahon, Benjamin; Gill, Sabrina et al. (2016) Relationship between mutant SOD1 maturation and inclusion formation in cell models. J Neurochem :|
|Xu, Guilian; Fromholt, Susan; Ayers, Jacob I et al. (2015) Substantially elevating the levels of Î±B-crystallin in spinal motor neurons of mutant SOD1 mice does not significantly delay paralysis or attenuate mutant protein aggregation. J Neurochem 133:452-64|
|Gelfand, Paul; Smith, Randy J; Stavitski, Eli et al. (2015) Characterization of Protein Structural Changes in Living Cells Using Time-Lapsed FTIR Imaging. Anal Chem 87:6025-31|
|Chattopadhyay, Madhuri; Nwadibia, Ekeoma; Strong, Cynthia D et al. (2015) The Disulfide Bond, but Not Zinc or Dimerization, Controls Initiation and Seeded Growth in Amyotrophic Lateral Sclerosis-linked Cu,Zn Superoxide Dismutase (SOD1) Fibrillation. J Biol Chem 290:30624-36|
|Xu, Guilian; Ayers, Jacob I; Roberts, Brittany L et al. (2015) Direct and indirect mechanisms for wild-type SOD1 to enhance the toxicity of mutant SOD1 in bigenic transgenic mice. Hum Mol Genet 24:1019-35|
|Saelices, Lorena; Johnson, Lisa M; Liang, Wilson Y et al. (2015) Uncovering the Mechanism of Aggregation of Human Transthyretin. J Biol Chem 290:28932-43|
|Bourassa, Megan W; Brown, Hilda H; Borchelt, David R et al. (2014) Metal-deficient aggregates and diminished copper found in cells expressing SOD1 mutations that cause ALS. Front Aging Neurosci 6:110|
|Ivanova, Magdalena I; Sievers, Stuart A; Guenther, Elizabeth L et al. (2014) Aggregation-triggering segments of SOD1 fibril formation support a common pathway for familial and sporadic ALS. Proc Natl Acad Sci U S A 111:197-201|
|Ming, Li-June; Valentine, Joan Selverstone (2014) Insights into SOD1-linked amyotrophic lateral sclerosis from NMR studies of Ni(2+)- and other metal-ion-substituted wild-type copper-zinc superoxide dismutases. J Biol Inorg Chem 19:647-57|
|Brown, Hilda H; Borchelt, David R (2014) Analysis of mutant SOD1 electrophoretic mobility by Blue Native gel electrophoresis; evidence for soluble multimeric assemblies. PLoS One 9:e104583|
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