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Assay Development for High Content Quantification of Sod1 Mutant Protein Aggregate Formation in Living Cells
Published on: October 4, 2017
In vivo pathogenic role of mutant SOD1 localized in the mitochondrial intermembrane space
Anissa Igoudjil1, Jordi Magrané, Lindsey R Fischer
1Department of Neurology and Neuroscience, Weill Medical College of Cornell University, New York, New York 10065, USA.
Abstract:
Mutations in Cu,Zn superoxide dismutase (SOD1) are associated with familial amyotrophic lateral sclerosis (ALS). Mutant SOD1 causes a complex array of pathological events, through toxic gain of function mechanisms, leading to selective motor neuron degeneration. Mitochondrial dysfunction is among the well established toxic effects of mutant SOD1, but its mechanisms are just starting to be elucidated. A portion of mutant SOD1 is localized in mitochondria, where it accumulates mostly on the outer membrane and inside the intermembrane space (IMS). Evidence in cultured cells suggests that mutant SOD1 in the IMS causes mitochondrial dysfunction and compromises cell viability. Therefore, to test its pathogenic role in vivo we generated transgenic mice expressing G93A mutant or wild-type (WT) human SOD1 targeted selectively to the mitochondrial IMS (mito-SOD1). We show that mito-SOD1 is correctly localized in the IMS, where it oligomerizes and acquires enzymatic activity. Mito-G93ASOD1 mice, but not mito-WTSOD1 mice, develop a progressive disease characterized by body weight loss, muscle weakness, brain atrophy, and motor impairment, which is more severe in females. These symptoms are associated with reduced spinal motor neuron counts and impaired mitochondrial bioenergetics, characterized by decreased cytochrome oxidase activity and defective calcium handling. However, there is no evidence of muscle denervation, a cardinal pathological feature of ALS. Together, our findings indicate that mutant SOD1 in the mitochondrial IMS causes mitochondrial dysfunction and neurodegeneration, but per se it is not sufficient to cause a full-fledged ALS phenotype, which requires the participation of mutant SOD1 localized in other cellular compartments.
Insights
Mutant copper-zinc superoxide dismutase (SOD1) in mitochondria causes neurodegeneration and mitochondrial dysfunction in mice. However, this mitochondrial localization alone is insufficient to cause the full pathology of amyotrophic lateral sclerosis (ALS).
Area of Science:
- Neuroscience
- Molecular Biology
- Genetics
Background:
- Mutations in Cu,Zn superoxide dismutase (SOD1) are a known cause of familial amyotrophic lateral sclerosis (ALS).
- Mitochondrial dysfunction is a key pathological event in ALS, linked to toxic gain-of-function mechanisms of mutant SOD1.
- Mutant SOD1 accumulates in mitochondria, particularly in the intermembrane space (IMS), where it is hypothesized to drive cellular damage.
Purpose of the Study:
- To investigate the specific role of mutant SOD1 localized to the mitochondrial IMS in the pathogenesis of ALS.
- To determine if mitochondrial IMS-localized mutant SOD1 is sufficient to cause a complete ALS phenotype in vivo.
Main Methods:
- Generation of transgenic mice expressing human G93A mutant or wild-type SOD1 targeted exclusively to the mitochondrial IMS (mito-SOD1).
- Assessment of mito-SOD1 localization, oligomerization, and enzymatic activity within the IMS.
- Phenotypic analysis of mito-SOD1 mice, including body weight, motor function, brain morphology, motor neuron counts, and mitochondrial bioenergetics (cytochrome oxidase activity, calcium handling).
Main Results:
- Mito-SOD1 was correctly targeted to the mitochondrial IMS and exhibited oligomerization and activity.
- Mice expressing mito-G93ASOD1 developed progressive neurodegenerative symptoms including weight loss, muscle weakness, brain atrophy, and motor impairment, with females being more severely affected.
- These symptoms correlated with reduced spinal motor neuron counts and impaired mitochondrial function, but notably lacked muscle denervation, a hallmark of ALS.
Conclusions:
- Mutant SOD1 within the mitochondrial IMS is sufficient to induce mitochondrial dysfunction and neurodegeneration.
- Mitochondrial IMS localization of mutant SOD1 alone does not recapitulate the full spectrum of ALS pathology.
- The complete ALS phenotype likely requires the contribution of mutant SOD1 in other cellular compartments besides the mitochondrial IMS.
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