Abstract
Mutations in SOD1 cause amyotrophic lateral sclerosis (ALS) through gain-of-function effects, yet the mechanisms by which misfolded mutant SOD1 (mutSOD1) protein impairs human motor neurons (MNs) remain unclear. Here, we use induced-pluripotent-stem-cell-derived MNs coupled to metabolic stable isotope labeling and mass spectrometry to investigate proteome-wide degradation dynamics. We find several proteins, including the ALS-causal valosin-containing protein (VCP), which predominantly acts in proteasome degradation and autophagy, that degrade slower in mutSOD1 relative to isogenic control MNs. The interactome of VCP is altered in mutSOD1 MNs in vitro, while VCP selectively accumulates in the affected motor cortex of ALS-SOD1 patients. Overexpression of VCP rescues mutSOD1 toxicity in MNs in vitro and in a C. elegans model in vivo, in part due to its ability to modulate the degradation of insoluble mutSOD1. Our results demonstrate that VCP contributes to mutSOD1-dependent degeneration, link two distinct ALS-causal genes, and highlight selective protein degradation impairment in ALS pathophysiology.
| Original language | English (US) |
|---|---|
| Article number | 113160 |
| Journal | Cell reports |
| Volume | 42 |
| Issue number | 10 |
| DOIs | |
| State | Published - Oct 31 2023 |
Funding
We are grateful to the following funding sources: National Institutes of Health (NIH), National Institute of Neurological Disorders and Stroke (NINDS), National Institute on Aging (NIA) R01NS104219 (E.K.) and R01AG078796 (J.N.S.), NIH/ NINDS grants R21NS107761 (E.K. and J.N.S.), the Les Turner ALS Foundation (E.K.), and the New York Stem Cell Foundation (E.K. and J.K.I.); Tau Consortium/Rainwater Charitable Foundation , NIH 2R01NS097850 ; Department of Defense grants W81XWH-21-1-0168 , W81XWH-20-1-0424 , and W81XWH-21-1-0131 ; the John Douglas French Alzheimer's Foundation (J.K.I.); NIH R01NS122908 , R01NS124802 and R01NS096746 and DOD W81XWH-21-1-0236 (R.G.K.); and the Swedish Research Council ( VR-MH 2019-01634 ) and Fondation Thierry Latran ( 00109319 ) (J.D.G.). We would like to thank Nandkishore Belur and Joe Mazzulli for assistance with GCase assays; Agneta Öberg for assistance with SOD1 ELISAs; Kathleen Wilsbach and Kathryn Gallo for providing paraffin-embedded tissue sections from the Johns Hopkins ALS Postmortem Core; Kunitoshi Yamanaka, Thorsten Hoppe, and Andre Franz for sharing the cdc48.1 overexpression worm strain; Jiou Wang for sharing the worms expressing SOD1; The National BioResource Project: C. elegans for the cdc48.1 and cdc48.2 mutant strains; and Joseph R. Klim and Kevin Eggan for sharing the HUES3 stem cell lines. Y.L. was supported by an ALS Association Milton-Safenowitz postdoctoral fellowship. J.K.I. is a New York Stem Cell Foundation – Robertson Investigator. E.K. is a Les Turner ALS Center Investigator and a New York Stem Cell Foundation – Robertson Investigator. We are grateful to the following funding sources: National Institutes of Health (NIH), National Institute of Neurological Disorders and Stroke (NINDS), National Institute on Aging (NIA) R01NS104219 (E.K.) and R01AG078796 (J.N.S.), NIH/NINDS grants R21NS107761 (E.K. and J.N.S.), the Les Turner ALS Foundation (E.K.), and the New York Stem Cell Foundation (E.K. and J.K.I.); Tau Consortium/Rainwater Charitable Foundation, NIH 2R01NS097850; Department of Defense grants W81XWH-21-1-0168, W81XWH-20-1-0424, and W81XWH-21-1-0131; the John Douglas French Alzheimer's Foundation (J.K.I.); NIH R01NS122908, R01NS124802 and R01NS096746 and DOD W81XWH-21-1-0236 (R.G.K.); and the Swedish Research Council (VR-MH 2019-01634) and Fondation Thierry Latran (00109319) (J.D.G.). We would like to thank Nandkishore Belur and Joe Mazzulli for assistance with GCase assays; Agneta Öberg for assistance with SOD1 ELISAs; Kathleen Wilsbach and Kathryn Gallo for providing paraffin-embedded tissue sections from the Johns Hopkins ALS Postmortem Core; Kunitoshi Yamanaka, Thorsten Hoppe, and Andre Franz for sharing the cdc48.1 overexpression worm strain; Jiou Wang for sharing the worms expressing SOD1; The National BioResource Project: C. elegans for the cdc48.1 and cdc48.2 mutant strains; and Joseph R. Klim and Kevin Eggan for sharing the HUES3 stem cell lines. Y.L. was supported by an ALS Association Milton-Safenowitz postdoctoral fellowship. J.K.I. is a New York Stem Cell Foundation – Robertson Investigator. E.K. is a Les Turner ALS Center Investigator and a New York Stem Cell Foundation – Robertson Investigator. Conceptualization, K.T. and E.K.; methodology, K.T. K.C.S. S.L.E. M.G. Y.L. C.W. J.D.G. J.K.I. R.G.K. J.N.S. and E.K.; investigation, K.T. K.C.S. S.L.E. M.G. Y.L. C.W. E.D.M. W.G. A.P.W. and T.J.H.; writing – original draft, E.K. and K.T.; writing – review & editing, K.T. K.C.S. S.L.E. M.G. Y.L. C.W. E.D.M. W.G. A.P.W. T.J.H. S.L.M. L.W.O. J.D.G. J.K.I. R.G.K. J.N.S. and E.K.; funding acquisition, J.K.I. R.G.K. J.N.S. and E.K.; resources, S.L.M. L.W.O. J.D.G. J.K.I. R.G.K. J.N.S. and E.K.; supervision, J.D.G. J.K.I. R.G.K. J.N.S. and E.K. J.K.I. is a co-founder of AcuraStem and Modulo Bio, SAB member of Spinogenix, Vesalius, and Synapticure, and employee of BioMarin Pharmaceutical. E.K. is a co-founder of NeuronGrow, SAB member of Axion Biosystems, ResQ Biotech, and Synapticure, and a consultant for Confluence Therapeutics; named companies were not involved in this project.
Keywords
- ALS
- CP: Neuroscience
- CP: Stem cell research
- SILAC-based mass spectrometry
- SOD1
- VCP/p97
- amyotrophic lateral sclerosis
- iPSCs
- motor neurons
- protein degradation
- ubiquitin
ASJC Scopus subject areas
- General Biochemistry, Genetics and Molecular Biology