title: Impaired extracellular secretion of mutant superoxide dismutase 1 associates with neurotoxicity in familial amyotrophic lateral sclerosis.
abstract: Mutations in the intracellular metalloenzyme superoxide dismutase 1 (SOD1) are linked to neurotoxicity in familial amyotrophic lateral sclerosis (ALS) by an unclear mechanism. Golgi fragmentation and endoplasmic reticulum stress are early hallmarks of spinal motor neuron pathology in transgenic mice overexpressing mutant SOD1, suggesting that dysfunction of the neuronal secretory pathway may contribute to ALS pathogenesis. We therefore proposed that mutant SOD1 directly engages and modulates the secretory pathway based on recent evidence of SOD1 secretion in diverse human cell lines. Here, we demonstrate that a fraction of active endogenous SOD1 is secreted by NSC-34 motor neuron-like cells via a brefeldin-A (BFA)-sensitive pathway. Expression of enhanced green fluorescent protein-tagged mutant human SOD1 (hSOD1-EGFP) in NSC-34 cells induced frequent cytoplasmic inclusions and protein insolubility that correlated with toxicity. In contrast, transfection of non-neuronal COS-7 cells resulted in mutant hSOD1-EGFP cytoplasmic inclusions, oligomerization, and fragmentation without detectable toxicity. Importantly, impaired secretion of hSOD1-EGFP was common to all 10 SOD1 mutants tested relative to wild-type protein in NSC-34 cells. Treatment with BFA inhibited hSOD1-EGFP secretion with pronounced BFA-induced toxicity in mutant cells. Extracellular targeting of mutant hSOD1-EGFP via SOD3 signal peptide fusion attenuated cytoplasmic inclusion formation and toxicity. The effect of elevated extracellular SOD1 was then evaluated in a transgenic rat model of ALS. Chronic intraspinal infusion of exogenous wild-type hSOD1 significantly delayed disease progression and endpoint in transgenic SOD1(G93A) rats. Collectively, these results suggest novel extracellular roles for SOD1 in ALS and support a causal relationship between mutant SOD1 secretion and intraneuronal toxicity.
authors.affiliation: Motor Neuron Disease Research Laboratory, Brain Injury and Repair Group, Howard Florey Institute of Experimental Physiology and Medicine, University of Melbourne, Victoria 3010, Australia. b.turner@hfi.unimelb.edu.au
authors.forename: Bradley J
authors.initials: BJ
authors.lastname: Turner
authors.forename: Julie D
authors.initials: JD
authors.lastname: Atkin
authors.forename: Manal A
authors.initials: MA
authors.lastname: Farg
authors.forename: Da Wei
authors.initials: DW
authors.lastname: Zang
authors.forename: Alan
authors.initials: A
authors.lastname: Rembach
authors.forename: Elizabeth C
authors.initials: EC
authors.lastname: Lopes
authors.forename: Justin D
authors.initials: JD
authors.lastname: Patch
authors.forename: Andrew F
authors.initials: AF
authors.lastname: Hill
authors.forename: Surindar S
authors.initials: SS
authors.lastname: Cheema
mesh_terms: D000690:Amyotrophic Lateral Sclerosis / Q000201:enzymology* / Q000235:genetics / Q000473:pathology* / Q000503:physiopathology; D000818:Animals; D030801:Animals, Genetically Modified; D019556:COS Cells; D002478:Cells, Cultured; D002522:Chlorocebus aethiops; D004195:Disease Models, Animal; D006801:Humans; D002479:Inclusion Bodies / Q000473:pathology; D051379:Mice; D009046:Motor Neurons / Q000201:enzymology* / Q000473:pathology*; D009068:Movement / Q000502:physiology; D009154:Mutation; D051381:Rats; D017207:Rats, Sprague-Dawley; D011993:Recombinant Fusion Proteins; D013482:Superoxide Dismutase / Q000235:genetics / Q000378:metabolism*; D000072105:Superoxide Dismutase-1
chemical_list: D011993:Recombinant Fusion Proteins; C000606290:SOD1 protein, human; C000606291:Sod1 protein, mouse; C000606292:Sod1 protein, rat; D013482:Superoxide Dismutase; D000072105:Superoxide Dismutase-1