Metal-deficient aggregates and diminished copper found in cells expressing SOD1 mutations that cause ALS

Disruptions in metal ion homeostasis have been described in association with amyotrophic lateral sclerosis (ALS) for a number of years but the precise mechanism of involvement is poorly understood. Metal ions are especially important to familial ALS cases caused by mutations in the metalloenzyme copper-zinc superoxide dismutase (SOD1). To investigate the role of metals in aggregation of mutant SOD1, we have examined the localization of metal ions in a cell culture model of overexpression. Chinese hamster ovary cells (CHO-K1) were transfected to overexpress SOD1 fused to yellow fluorescent protein (YFP) to readily identify the transfected cells and the intracellular aggregates that develop in the cells expressing mutant or wild-type (WT) SOD1. The concentration and distribution of iron, copper, and zinc were determined for four SOD1 mutants (A4V, G37R, H80R, and D125H) as well as a WT SOD1 using X-ray fluorescence microscopy (XFM). Results demonstrated that the SOD1 aggregates were metal-deficient within the cells, which is consistent with recent in vitro studies. In addition, all SOD1 mutants showed significantly decreased copper content compared to the WT SOD1 cells, regardless of the mutant’s ability to bind copper. These results suggest that SOD1 overexpression creates an unmet demand on the cell for copper. This is particularly true for the SOD1 mutants where copper delivery may also be impaired. Hence, the SOD1 mutants are less stable than WT SOD1 and if copper is limited, aggregate formation of the metal-deficient, mutant SOD1 protein occurs.

[1]  Jacob I. Ayers,et al.  Distinctive features of the D101N and D101G variants of superoxide dismutase 1; two mutations that produce rapidly progressing motor neuron disease , 2014, Journal of neurochemistry.

[2]  D. Borchelt,et al.  An Analysis of Interactions between Fluorescently-Tagged Mutant and Wild-Type SOD1 in Intracellular Inclusions , 2013, PloS one.

[3]  D. Borchelt,et al.  Features of wild-type human SOD1 limit interactions with misfolded aggregates of mouse G86R Sod1 , 2013, Molecular Neurodegeneration.

[4]  Olubunmi Abel,et al.  Development of a Smartphone App for a Genetics Website: The Amyotrophic Lateral Sclerosis Online Genetics Database (ALSoD) , 2013, JMIR mHealth and uHealth.

[5]  D. Borchelt,et al.  A novel variant of human superoxide dismutase 1 harboring amyotrophic lateral sclerosis‐associated and experimental mutations in metal‐binding residues and free cysteines lacks toxicity in vivo , 2012, Journal of neurochemistry.

[6]  D. Borchelt,et al.  Superoxide dismutase 1 encoding mutations linked to ALS adopts a spectrum of misfolded states , 2011, Molecular Neurodegeneration.

[7]  A. Al-Chalabi,et al.  Keeping up with genetic discoveries in amyotrophic lateral sclerosis: The ALSoD and ALSGene databases , 2011, Amyotrophic lateral sclerosis : official publication of the World Federation of Neurology Research Group on Motor Neuron Diseases.

[8]  D. Borchelt,et al.  Copper and Zinc Metallation Status of Copper-Zinc Superoxide Dismutase from Amyotrophic Lateral Sclerosis Transgenic Mice* , 2010, The Journal of Biological Chemistry.

[9]  A. Hubbard,et al.  Copper handling machinery of the brain. , 2010, Metallomics : integrated biometal science.

[10]  J. Valentine,et al.  Disrupted zinc-binding sites in structures of pathogenic SOD1 variants D124V and H80R. , 2010, Biochemistry.

[11]  Lisa M. Miller,et al.  Amyloid plaques in PSAPP mice bind less metal than plaques in human Alzheimer's disease , 2009, NeuroImage.

[12]  D. Borchelt,et al.  Variation in aggregation propensities among ALS-associated variants of SOD1: Correlation to human disease , 2009, Human molecular genetics.

[13]  D. Borchelt,et al.  Role of mutant SOD1 disulfide oxidation and aggregation in the pathogenesis of familial ALS , 2009, Proceedings of the National Academy of Sciences.

[14]  L. Klomp,et al.  Role of transition metals in the pathogenesis of amyotrophic lateral sclerosis. , 2008, Biochemical Society transactions.

[15]  Bryan F. Shaw,et al.  Detergent-insoluble Aggregates Associated with Amyotrophic Lateral Sclerosis in Transgenic Mice Contain Primarily Full-length, Unmodified Superoxide Dismutase-1* , 2008, Journal of Biological Chemistry.

[16]  Bryan F. Shaw,et al.  How do ALS-associated mutations in superoxide dismutase 1 promote aggregation of the protein? , 2007, Trends in biochemical sciences.

[17]  N. Lee,et al.  Role of zinc in ALS , 2007, Amyotrophic lateral sclerosis : official publication of the World Federation of Neurology Research Group on Motor Neuron Diseases.

[18]  Antonio Lanzirotti,et al.  Synchrotron-based infrared and X-ray imaging shows focalized accumulation of Cu and Zn co-localized with beta-amyloid deposits in Alzheimer's disease. , 2006, Journal of structural biology.

[19]  H. Kozłowski,et al.  Copper homeostasis and neurodegenerative disorders (Alzheimer's, prion, and Parkinson's diseases and amyotrophic lateral sclerosis). , 2006, Chemical reviews.

[20]  Bryan F. Shaw,et al.  Destabilization of apoprotein is insufficient to explain Cu,Zn-superoxide dismutase-linked ALS pathogenesis. , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[21]  J. Valentine,et al.  Copper-zinc superoxide dismutase and amyotrophic lateral sclerosis. , 2005, Annual review of biochemistry.

[22]  Roberto Cappai,et al.  Metals and amyloid‐β in Alzheimer's disease , 2005, International journal of experimental pathology.

[23]  C. Griesinger,et al.  Release of long-range tertiary interactions potentiates aggregation of natively unstructured alpha-synuclein. , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[24]  D. Selkoe,et al.  Cell biology of protein misfolding: The examples of Alzheimer's and Parkinson's diseases , 2004, Nature Cell Biology.

[25]  L. Bruijn,et al.  Unraveling the mechanisms involved in motor neuron degeneration in ALS. , 2004, Annual review of neuroscience.

[26]  D. Selkoe Folding proteins in fatal ways , 2003, Nature.

[27]  Ashley I. Bush,et al.  The metallobiology of Alzheimer's disease , 2003, Trends in Neurosciences.

[28]  V. Bindokas,et al.  No correlation between aggregates of Cu/Zn superoxide dismutase and cell death in familial amyotrophic lateral sclerosis , 2002, Journal of neurochemistry.

[29]  Robert H. Brown,et al.  Familial Amyotrophic Lateral Sclerosis-associated Mutations Decrease the Thermal Stability of Distinctly Metallated Species of Human Copper/Zinc Superoxide Dismutase* , 2002, The Journal of Biological Chemistry.

[30]  Robert H. Brown,et al.  Decreased Metallation and Activity in Subsets of Mutant Superoxide Dismutases Associated with Familial Amyotrophic Lateral Sclerosis* 210 , 2002, The Journal of Biological Chemistry.

[31]  D. Selkoe Alzheimer's disease: genes, proteins, and therapy. , 2001, Physiological reviews.

[32]  T. O’Halloran,et al.  Multiple Protein Domains Contribute to the Action of the Copper Chaperone for Superoxide Dismutase* , 1999, The Journal of Biological Chemistry.

[33]  J. D. Robertson,et al.  Copper, iron and zinc in Alzheimer's disease senile plaques , 1998, Journal of the Neurological Sciences.

[34]  Xudong Huang,et al.  Zinc-induced Alzheimer’s Aβ1–40 Aggregation Is Mediated by Conformational Factors* , 1997, The Journal of Biological Chemistry.

[35]  A. Doble The pharmacology and mechanism of action of riluzole , 1996, Neurology.

[36]  L. Ellerby,et al.  Copper−Zinc Superoxide Dismutase: Why Not pH-Dependent? , 1996 .

[37]  D. Borchelt,et al.  Superoxide dismutase 1 with mutations linked to familial amyotrophic lateral sclerosis possesses significant activity. , 1994, Proceedings of the National Academy of Sciences of the United States of America.

[38]  知弘 松山,et al.  虚血負荷後の海馬Copper-Zinc superoxide dismutase発現の検討 , 1994 .

[39]  M. Pericak-Vance,et al.  Amyotrophic lateral sclerosis and structural defects in Cu,Zn superoxide dismutase. , 1993, Science.

[40]  C. Soto,et al.  Protein Misfolding in Neurodegenerative Diseases: The Key Pending Questions , 2013 .

[41]  Claudio Soto,et al.  Unfolding the role of protein misfolding in neurodegenerative diseases , 2003, Nature Reviews Neuroscience.

[42]  R. Miller,et al.  Riluzole for amyotrophic lateral sclerosis (ALS)/motor neuron disease (MND). , 2003, Amyotrophic lateral sclerosis and other motor neuron disorders : official publication of the World Federation of Neurology, Research Group on Motor Neuron Diseases.

[43]  Patrik Brundin,et al.  Pathogenesis of Parkinson's disease: dopamine, vesicles and alpha-synuclein. , 2002, Nature reviews. Neuroscience.