title: Overexpression of alpha-synuclein at non-toxic levels increases dopaminergic cell death induced by copper exposure via modulation of protein degradation pathways.
abstract: Gene multiplications or point mutations in alpha (α)-synuclein are associated with familial and sporadic Parkinson's disease (PD). An increase in copper (Cu) levels has been reported in the cerebrospinal fluid and blood of PD patients, while occupational exposure to Cu has been suggested to augment the risk to develop PD. We aimed to elucidate the mechanisms by which α-synuclein and Cu regulate dopaminergic cell death. Short-term overexpression of wild type (WT) or mutant A53T α-synuclein had no toxic effect in human dopaminergic cells and primary midbrain cultures, but it exerted a synergistic effect on Cu-induced cell death. Cell death induced by Cu was potentiated by overexpression of the Cu transporter protein 1 (Ctr1) and depletion of intracellular glutathione (GSH) indicating that the toxic effects of Cu are linked to alterations in its intracellular homeostasis. Using the redox sensor roGFP, we demonstrated that Cu-induced oxidative stress was primarily localized in the cytosol and not in the mitochondria. However, α-synuclein overexpression had no effect on Cu-induced oxidative stress. WT or A53T α-synuclein overexpression exacerbated Cu toxicity in dopaminergic and yeast cells in the absence of α-synuclein aggregation. Cu increased autophagic flux and protein ubiquitination. Impairment of autophagy by overexpression of a dominant negative Atg5 form or inhibition of the ubiquitin/proteasome system (UPS) with MG132 enhanced Cu-induced cell death. However, only inhibition of the UPS stimulated the synergistic toxic effects of Cu and α-synuclein overexpression. Our results demonstrate that α-synuclein stimulates Cu toxicity in dopaminergic cells independent from its aggregation via modulation of protein degradation pathways.
authors.affiliation: Redox Biology Center, University of Nebraska-Lincoln, Lincoln, NE, USA; School of Veterinary Medicine and Biomedical Sciences, University of Nebraska-Lincoln, Lincoln, NE, USA.
authors.forename: Annadurai
authors.initials: A
authors.lastname: Anandhan
authors.affiliation: Redox Biology Center, University of Nebraska-Lincoln, Lincoln, NE, USA; School of Veterinary Medicine and Biomedical Sciences, University of Nebraska-Lincoln, Lincoln, NE, USA.
authors.forename: Humberto
authors.initials: H
authors.lastname: Rodriguez-Rocha
authors.affiliation: Department of Biochemistry, University of Nebraska-Lincoln, Lincoln, NE, USA.
authors.forename: Iryna
authors.initials: I
authors.lastname: Bohovych
authors.affiliation: Department of Medicinal Chemistry and Molecular Pharmacology, Purdue University, West Lafayette, IN, USA.
authors.forename: Amy M
authors.initials: AM
authors.lastname: Griggs
authors.affiliation: Redox Biology Center, University of Nebraska-Lincoln, Lincoln, NE, USA; School of Veterinary Medicine and Biomedical Sciences, University of Nebraska-Lincoln, Lincoln, NE, USA.
authors.forename: Laura
authors.initials: L
authors.lastname: Zavala-Flores
authors.affiliation: College of Medicine, The University of Arizona, Tucson, AZ, USA.
authors.forename: Elsa M
authors.initials: EM
authors.lastname: Reyes-Reyes
authors.affiliation: Redox Biology Center, University of Nebraska-Lincoln, Lincoln, NE, USA; Department of Biochemistry, University of Nebraska-Lincoln, Lincoln, NE, USA.
authors.forename: Javier
authors.initials: J
authors.lastname: Seravalli
authors.affiliation: Weldon School of Biomedical Engineering and School of Materials Engineering, Purdue University, West Lafayette, IN, USA.
authors.forename: Lia A
authors.initials: LA
authors.lastname: Stanciu
authors.affiliation: Redox Biology Center, University of Nebraska-Lincoln, Lincoln, NE, USA; Department of Biochemistry, University of Nebraska-Lincoln, Lincoln, NE, USA.
authors.forename: Jaekwon
authors.initials: J
authors.lastname: Lee
authors.affiliation: Department of Medicinal Chemistry and Molecular Pharmacology, Purdue University, West Lafayette, IN, USA.
authors.forename: Jean-Christophe
authors.initials: JC
authors.lastname: Rochet
authors.affiliation: Redox Biology Center, University of Nebraska-Lincoln, Lincoln, NE, USA; Department of Biochemistry, University of Nebraska-Lincoln, Lincoln, NE, USA.
authors.forename: Oleh
authors.initials: O
authors.lastname: Khalimonchuk
authors.affiliation: Redox Biology Center, University of Nebraska-Lincoln, Lincoln, NE, USA; School of Veterinary Medicine and Biomedical Sciences, University of Nebraska-Lincoln, Lincoln, NE, USA. Electronic address: rfrancocruz2@unl.edu.
authors.forename: Rodrigo
authors.initials: R
authors.lastname: Franco
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D020169:Caspases / Q000378:metabolism; D002478:Cells, Cultured; D003300:Copper / Q000494:pharmacology*; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D059290:Dopaminergic Neurons / Q000187:drug effects* / Q000378:metabolism; D004622:Embryo, Mammalian; D005786:Gene Expression Regulation / Q000187:drug effects*; D006801:Humans; D007976:Leupeptins / Q000494:pharmacology; D008636:Mesencephalon / Q000166:cytology; D009154:Mutation / Q000235:genetics; D009447:Neuroblastoma / Q000473:pathology; D059748:Proteolysis / Q000187:drug effects*; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D013997:Time Factors; D014446:Tyrosine 3-Monooxygenase / Q000378:metabolism; D051844:alpha-Synuclein / Q000235:genetics / Q000378:metabolism*
chemical_list: D015853:Cysteine Proteinase Inhibitors; D007976:Leupeptins; D051844:alpha-Synuclein; D003300:Copper; D014446:Tyrosine 3-Monooxygenase; D020169:Caspases; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde; C029892:cupric chloride
keywords: A53T; AMPK; Autophagy; Copper and Parkinson's disease; Oxidative stress; Proteasome; Ubiquitination; α-Synuclein