Inhibition of protein ubiquitination by paraquat and 1-methyl-4-phenylpyridinium impairs ubiquitin-dependent protein degradation pathways

Authors

Juliana Navarro-Yepes, University of Nebraska-Lincoln
Anandhan Annadurai, University of Nebraska-LincolnFollow
Erin Bradley, University of Nebraska-Lincoln
Iryna Bohovych, University of Nebraska- LincolnFollow
Bo Yarabe, University of Nebraska-Lincoln
Annemieke de Jong, The Netherlands Cancer Institute
Huib Ovaa, The Netherlands Cancer Institute
You Zhou, University of Nebraska-LincolnFollow
Oleh Khalimonchuk, University of Nebraska-LincolnFollow
Betzabet Quintanilla-Vega, CINVESTAV-IPN, Mexico City, Mexico
Rodrigo Franco, University of Nebraska-LincolnFollow

Date of this Version

2016

Citation

Mol Neurobiol. 2016 October ; 53(8): 5229–5251.

Comments

Copyright 2015 Springer Verlag.

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 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 effect 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. Inhibition of proteasomal activity by PQ was found to be a late event in cell death progression, and had no effect on either the toxicity of MPP⁺ or PQ, or 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 autophagic. 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 inhibition of protein ubiquitination by PQ and MPP⁺ is involved in the dysfunction of Ub-dependent protein degradation pathways.