title: Inhibition of Protein Ubiquitination by Paraquat and 1-Methyl-4-Phenylpyridinium Impairs Ubiquitin-Dependent Protein Degradation Pathways.
abstract: Intracytoplasmic inclusions of protein aggregates in dopaminergic cells (Lewy bodies) are the pathological hallmark of Parkinson's disease (PD). Ubiquitin (Ub), alpha (α)-synuclein, p62/sequestosome 1, and oxidized proteins are the major components of Lewy bodies. However, the mechanisms involved in the impairment of misfolded/oxidized protein degradation pathways in PD are still unclear. PD is linked to mitochondrial dysfunction and environmental pesticide exposure. In this work, we evaluated the effects of the pesticide paraquat (PQ) and the mitochondrial toxin 1-methyl-4-phenylpyridinium (MPP(+)) on Ub-dependent protein degradation pathways. No increase in the accumulation of Ub-bound proteins or aggregates was observed in dopaminergic cells (SK-N-SH) treated with PQ or MPP(+), or in mice chronically exposed to PQ. PQ decreased Ub protein content, but not its mRNA transcription. Protein synthesis inhibition with cycloheximide depleted Ub levels and potentiated PQ-induced cell death. The inhibition of proteasomal activity by PQ was found to be a late event in cell death progression and had neither effect on the toxicity of either MPP(+) or PQ, nor on the accumulation of oxidized sulfenylated, sulfonylated (DJ-1/PARK7 and peroxiredoxins), and carbonylated proteins induced by PQ. PQ- and MPP(+)-induced Ub protein depletion prompted the dimerization/inactivation of the Ub-binding protein p62 that regulates the clearance of ubiquitinated proteins by autophagy. We confirmed that PQ and MPP(+) impaired autophagy flux and that the blockage of autophagy by the overexpression of a dominant-negative form of the autophagy protein 5 (dnAtg5) stimulated their toxicity, but there was no additional effect upon inhibition of the proteasome. PQ induced an increase in the accumulation of α-synuclein in dopaminergic cells and membrane-associated foci in yeast cells. Our results demonstrate that the inhibition of protein ubiquitination by PQ and MPP(+) is involved in the dysfunction of Ub-dependent 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, 114 VBS 0905, Lincoln, NE, 68583, USA.|Department of Toxicology, CINVESTAV-IPN, IPN No. 2508, Colonia Zacatenco, Mexico City, D.F., 07360, Mexico.
authors.forename: Juliana
authors.initials: J
authors.lastname: Navarro-Yepes
authors.affiliation: Redox Biology Center, University of Nebraska-Lincoln, Lincoln, NE, USA.|School of Veterinary Medicine and Biomedical Sciences, University of Nebraska-Lincoln, 114 VBS 0905, Lincoln, NE, 68583, USA.
authors.forename: Annadurai
authors.initials: A
authors.lastname: Anandhan
authors.affiliation: Redox Biology Center, University of Nebraska-Lincoln, Lincoln, NE, USA.
authors.forename: Erin
authors.initials: E
authors.lastname: Bradley
authors.affiliation: Redox Biology Center, University of Nebraska-Lincoln, Lincoln, NE, USA.|Department of Biochemistry, University of Nebraska-Lincoln, Lincoln, NE, USA.
authors.forename: Iryna
authors.initials: I
authors.lastname: Bohovych
authors.affiliation: Redox Biology Center, University of Nebraska-Lincoln, Lincoln, NE, USA.
authors.forename: Bo
authors.initials: B
authors.lastname: Yarabe
authors.affiliation: Division of Cell Biology II, The Netherlands Cancer Institute, Amsterdam, The Netherlands.
authors.forename: Annemieke
authors.initials: A
authors.lastname: de Jong
authors.affiliation: Division of Cell Biology II, The Netherlands Cancer Institute, Amsterdam, The Netherlands.
authors.forename: Huib
authors.initials: H
authors.lastname: Ovaa
authors.affiliation: Department of Biochemistry, University of Nebraska-Lincoln, Lincoln, NE, USA.
authors.forename: You
authors.initials: Y
authors.lastname: Zhou
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: Department of Toxicology, CINVESTAV-IPN, IPN No. 2508, Colonia Zacatenco, Mexico City, D.F., 07360, Mexico. mquintan@cinvestav.mx.
authors.forename: Betzabet
authors.initials: B
authors.lastname: Quintanilla-Vega
authors.affiliation: Redox Biology Center, University of Nebraska-Lincoln, Lincoln, NE, USA. rfrancocruz2@unl.edu.|School of Veterinary Medicine and Biomedical Sciences, University of Nebraska-Lincoln, 114 VBS 0905, Lincoln, NE, 68583, USA. rfrancocruz2@unl.edu.
authors.forename: Rodrigo
authors.initials: R
authors.lastname: Franco
mesh_terms: D015655:1-Methyl-4-phenylpyridinium / Q000633:toxicity*; D000818:Animals; D001343:Autophagy / Q000187:drug effects; D016923:Cell Death / Q000187:drug effects; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D019281:Dimerization; D059290:Dopaminergic Neurons / Q000187:drug effects / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006801:Humans; D008810:Mice, Inbred C57BL; D008954:Models, Biological; D010084:Oxidation-Reduction; D010269:Paraquat / Q000633:toxicity*; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D059748:Proteolysis / Q000187:drug effects*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D000071456:Sequestosome-1 Protein / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects; D025801:Ubiquitin / Q000235:genetics / Q000378:metabolism*; D054875:Ubiquitination / Q000187:drug effects*; D051844:alpha-Synuclein / Q000378:metabolism
chemical_list: D012333:RNA, Messenger; C099718:SQSTM1 protein, human; D000071456:Sequestosome-1 Protein; D025801:Ubiquitin; D051844:alpha-Synuclein; D049452:Green Fluorescent Proteins; D046988:Proteasome Endopeptidase Complex; D010269:Paraquat; D015655:1-Methyl-4-phenylpyridinium
keywords: Autophagy; MPP+; Parkinson’s disease; Pesticides; SQSTM1; Sequestosome 1; Ubiquitin-proteasome system; Ubiquitylation