title: Progressive aggregation despite chaperone associations of a mutant SOD1-YFP in transgenic mice that develop ALS.
abstract: Recent studies suggest that superoxide dismutase 1 (SOD1)-linked amyotrophic lateral sclerosis results from destabilization and misfolding of mutant forms of this abundant cytosolic enzyme. Here, we have tracked the expression and fate of a misfolding-prone human SOD1, G85R, fused to YFP, in a line of transgenic G85R SOD1-YFP mice. These mice, but not wild-type human SOD1-YFP transgenics, developed lethal paralyzing motor symptoms at 9 months. In situ RNA hybridization of spinal cords revealed predominant expression in motor neurons in spinal cord gray matter in all transgenic animals. Concordantly, G85R SOD-YFP was diffusely fluorescent in motor neurons of animals at 1 and 6 months of age, but at the time of symptoms, punctate aggregates were observed in cell bodies and processes. Biochemical analyses of spinal cord soluble extracts indicated that G85R SOD-YFP behaved as a misfolded monomer at all ages. It became progressively insoluble at 6 and 9 months of age, associated with presence of soluble oligomers observable by gel filtration. Immunoaffinity capture and mass spectrometry revealed association of G85R SOD-YFP, but not WT SOD-YFP, with the cytosolic chaperone Hsc70 at all ages. In addition, 3 Hsp110's, nucleotide exchange factors for Hsp70s, were captured at 6 and 9 months. Despite such chaperone interactions, G85R SOD-YFP formed insoluble inclusions at late times, containing predominantly intermediate filament proteins. We conclude that motor neurons, initially "compensated" to maintain the misfolded protein in a soluble state, become progressively unable to do so.
authors.affiliation: Howard Hughes Medical Institute, Department of Genetics, Section of Comparative Medicine, and Department of Neurosurgery, Yale University School of Medicine, New Haven, CT 06510, USA.
authors.forename: Jiou
authors.initials: J
authors.lastname: Wang
authors.forename: George W
authors.initials: GW
authors.lastname: Farr
authors.forename: Caroline J
authors.initials: CJ
authors.lastname: Zeiss
authors.forename: Diego J
authors.initials: DJ
authors.lastname: Rodriguez-Gil
authors.forename: Jean H
authors.initials: JH
authors.lastname: Wilson
authors.forename: Krystyna
authors.initials: K
authors.lastname: Furtak
authors.forename: D Thomas
authors.initials: DT
authors.lastname: Rutkowski
authors.forename: Randal J
authors.initials: RJ
authors.lastname: Kaufman
authors.forename: Cristian I
authors.initials: CI
authors.lastname: Ruse
authors.forename: John R
authors.initials: JR
authors.lastname: Yates
authors.forename: Steve
authors.initials: S
authors.lastname: Perrin
authors.forename: Mel B
authors.initials: MB
authors.lastname: Feany
authors.forename: Arthur L
authors.initials: AL
authors.lastname: Horwich
mesh_terms: D000690:Amyotrophic Lateral Sclerosis / Q000235:genetics*; D000818:Animals; D005904:Glial Fibrillary Acidic Protein / Q000378:metabolism; D008164:Luminescent Proteins / Q000235:genetics*; D051379:Mice; D008822:Mice, Transgenic; D018832:Molecular Chaperones / Q000378:metabolism*; D009046:Motor Neurons / Q000378:metabolism; D009693:Nucleic Acid Hybridization; D012995:Solubility; D013116:Spinal Cord / Q000166:cytology / Q000378:metabolism; D013482:Superoxide Dismutase / Q000235:genetics*; D025801:Ubiquitin / Q000378:metabolism
chemical_list: D005904:Glial Fibrillary Acidic Protein; D008164:Luminescent Proteins; D018832:Molecular Chaperones; D025801:Ubiquitin; D013482:Superoxide Dismutase