title: Changes in endoplasmic reticulum stress proteins and aldolase A in cells exposed to dopamine.
abstract: In Parkinson's disease, oxidative stress is implicated in protein misfolding and aggregation, which may activate the unfolded protein response by the endoplasmic reticulum (ER). Dopamine (DA) can initiate oxidative stress via H(2)O(2) formation by DA metabolism and by oxidation into DA quinone. We have previously shown that DA quinone induces oxidative protein modification, mitochondrial dysfunction in vitro, and dopaminergic cell toxicity in vivo and in vitro. In this study, we used cysteine- and lysine-reactive fluorescent dyes with 2D difference in-gel electrophoresis, mass spectrometry, and peptide mass fingerprint analysis to identify proteins in PC12 cell mitochondrial-enriched fractions that were altered in abundance following DA exposure (150 muM, 16 h). Quantitative changes in proteins labeled with fluorescent dyes indicated increases in a subset of proteins after DA exposure: calreticulin, ERp29, ERp99, Grp58, Grp78, Grp94 and Orp150 (149-260%), and decreased levels of aldolase A (39-42%). Changes in levels of several proteins detected by 2D difference in-gel electrophoresis were confirmed by western blot. Using this unbiased proteomics approach, our findings demonstrated that in PC12 cells, DA exposure leads to a cellular response indicative of ER stress prior to the onset of cell death, providing a potential link between DA and the unfolded protein response in the pathogenesis of Parkinson's disease.
authors.affiliation: Department of Neurology, University of Pittsburgh, Pittsburgh, Pennsylvania 15261, USA.
authors.forename: April A
authors.initials: AA
authors.lastname: Dukes
authors.forename: Victor S
authors.initials: VS
authors.lastname: Van Laar
authors.forename: Michael
authors.initials: M
authors.lastname: Cascio
authors.forename: Teresa G
authors.initials: TG
authors.lastname: Hastings
mesh_terms: D000818:Animals; D037282:Calreticulin / Q000187:drug effects / Q000378:metabolism; D004298:Dopamine / Q000378:metabolism* / Q000494:pharmacology; D015536:Down-Regulation / Q000187:drug effects / Q000502:physiology; D015180:Electrophoresis, Gel, Two-Dimensional; D004721:Endoplasmic Reticulum / Q000187:drug effects / Q000378:metabolism*; D005456:Fluorescent Dyes; D005634:Fructose-Bisphosphate Aldolase / Q000187:drug effects / Q000378:metabolism*; D018840:HSP70 Heat-Shock Proteins / Q000187:drug effects / Q000378:metabolism; D006360:Heat-Shock Proteins / Q000187:drug effects / Q000378:metabolism*; D013058:Mass Spectrometry; D008562:Membrane Glycoproteins / Q000187:drug effects / Q000378:metabolism; D008565:Membrane Proteins / Q000187:drug effects / Q000378:metabolism; D018832:Molecular Chaperones / Q000187:drug effects / Q000378:metabolism; D009474:Neurons / Q000187:drug effects / Q000378:metabolism*; D018384:Oxidative Stress / Q000187:drug effects / Q000502:physiology*; D016716:PC12 Cells; D010300:Parkinson Disease / Q000378:metabolism / Q000503:physiopathology; D019704:Protein Disulfide-Isomerases / Q000187:drug effects / Q000378:metabolism; D017510:Protein Folding; D011506:Proteins / Q000187:drug effects / Q000378:metabolism; D051381:Rats; D015854:Up-Regulation / Q000187:drug effects / Q000502:physiology; D000091342:Endoplasmic Reticulum Chaperone BiP
chemical_list: D037282:Calreticulin; D004298:Dopamine; D005456:Fluorescent Dyes; D005634:Fructose-Bisphosphate Aldolase; D018840:HSP70 Heat-Shock Proteins; D006360:Heat-Shock Proteins; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D018832:Molecular Chaperones; D019704:Protein Disulfide-Isomerases; D011506:Proteins; C110909:Erp29 protein, rat; C051684:GRP78 protein, rat; D000091342:Endoplasmic Reticulum Chaperone BiP; C018551:glucose-regulated proteins; C050087:oxygen-regulated proteins; C052768:endoplasmic reticulum glycoprotein p99; C513196:PDIA3 protein, rat