title: Intercellular propagated misfolding of wild-type Cu/Zn superoxide dismutase occurs via exosome-dependent and -independent mechanisms.
abstract: Amyotrophic lateral sclerosis (ALS) is predominantly sporadic, but associated with heritable genetic mutations in 5-10% of cases, including those in Cu/Zn superoxide dismutase (SOD1). We previously showed that misfolding of SOD1 can be transmitted to endogenous human wild-type SOD1 (HuWtSOD1) in an intracellular compartment. Using NSC-34 motor neuron-like cells, we now demonstrate that misfolded mutant and HuWtSOD1 can traverse between cells via two nonexclusive mechanisms: protein aggregates released from dying cells and taken up by macropinocytosis, and exosomes secreted from living cells. Furthermore, once HuWtSOD1 propagation has been established, misfolding of HuWtSOD1 can be efficiently and repeatedly propagated between HEK293 cell cultures via conditioned media over multiple passages, and to cultured mouse primary spinal cord cells transgenically expressing HuWtSOD1, but not to cells derived from nontransgenic littermates. Conditioned media transmission of HuWtSOD1 misfolding in HEK293 cells is blocked by HuWtSOD1 siRNA knockdown, consistent with human SOD1 being a substrate for conversion, and attenuated by ultracentrifugation or incubation with SOD1 misfolding-specific antibodies, indicating a relatively massive transmission particle which possesses antibody-accessible SOD1. Finally, misfolded and protease-sensitive HuWtSOD1 comprises up to 4% of total SOD1 in spinal cords of patients with sporadic ALS (SALS). Propagation of HuWtSOD1 misfolding, and its subsequent cell-to-cell transmission, is thus a candidate process for the molecular pathogenesis of SALS, which may provide novel treatment and biomarker targets for this devastating disease.
authors.affiliation: Department of Medicine (Neurology), University of British Columbia and Vancouver Coastal Health Research Institute, Brain Research Centre, Vancouver, BC, Canada V6T 2B5.
authors.forename: Leslie I
authors.initials: LI
authors.lastname: Grad
authors.forename: Justin J
authors.initials: JJ
authors.lastname: Yerbury
authors.forename: Bradley J
authors.initials: BJ
authors.lastname: Turner
authors.forename: William C
authors.initials: WC
authors.lastname: Guest
authors.forename: Edward
authors.initials: E
authors.lastname: Pokrishevsky
authors.forename: Megan A
authors.initials: MA
authors.lastname: O'Neill
authors.forename: Anat
authors.initials: A
authors.lastname: Yanai
authors.forename: Judith M
authors.initials: JM
authors.lastname: Silverman
authors.forename: Rafaa
authors.initials: R
authors.lastname: Zeineddine
authors.forename: Lisa
authors.initials: L
authors.lastname: Corcoran
authors.forename: Janet R
authors.initials: JR
authors.lastname: Kumita
authors.forename: Leila M
authors.initials: LM
authors.lastname: Luheshi
authors.forename: Masoud
authors.initials: M
authors.lastname: Yousefi
authors.forename: Bradley M
authors.initials: BM
authors.lastname: Coleman
authors.forename: Andrew F
authors.initials: AF
authors.lastname: Hill
authors.forename: Steven S
authors.initials: SS
authors.lastname: Plotkin
authors.forename: Ian R
authors.initials: IR
authors.lastname: Mackenzie
authors.forename: Neil R
authors.initials: NR
authors.lastname: Cashman
mesh_terms: D000690:Amyotrophic Lateral Sclerosis / Q000378:metabolism / Q000503:physiopathology*; D000818:Animals; D002460:Cell Line; D004591:Electrophoresis, Polyacrylamide Gel; D055354:Exosomes / Q000378:metabolism*; D006801:Humans; D051379:Mice; D008854:Microscopy, Electron; D010873:Pinocytosis / Q000502:physiology; D017510:Protein Folding; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics; D013482:Superoxide Dismutase / Q000737:chemistry* / Q000378:metabolism
chemical_list: D034741:RNA, Small Interfering; D013482:Superoxide Dismutase
keywords: disease-specific epitope; intercellular transmission; prion-like; protein misfolding