Expression of A53T mutant but not wild-type alpha-synuclein in PC12 cells induces alterations of the ubiquitin-dependent degradation system, loss of dopamine release, and autophagic cell death.
Alpha-synuclein mutations have been identified in certain families with Parkinson's disease (PD), and alpha-synuclein is a major component of Lewy bodies. Other genetic data indicate that the ubiquitin-dependent proteolytic system is involved in PD pathogenesis. We have generated stable PC12 cell lines expressing wild-type or A53T mutant human alpha-synuclein. Lines expressing mutant but not wild-type alpha-synuclein show: (1) disruption of the ubiquitin-dependent proteolytic system, manifested by small cytoplasmic ubiquitinated aggregates and by an increase in polyubiquitinated proteins; (2) enhanced baseline nonapoptotic death; (3) marked accumulation of autophagic-vesicular structures; (4) impairment of lysosomal hydrolysis and proteasomal function; and (5) loss of catecholamine-secreting dense core granules and an absence of depolarization-induced dopamine release. Such findings raise the possibility that the primary abnormality in these cells may involve one or more deficits in the lysosomal and/or proteasomal degradation pathways, which in turn lead to loss of dopaminergic capacity and, ultimately, to death. These cells may serve as a model to study the effects of aberrant alpha-synuclein on dopaminergic cell function and survival.
Stefanis
L
L
Department of Pathology, Columbia University, New York, New York 10032, USA. ls76@columbia.edu
Larsen
K E
KE
Rideout
H J
HJ
Sulzer
D
D
Greene
L A
LA
D019943:Amino Acid Substitution; D000818:Animals; D001343:Autophagy / Q000187:drug effects / Q000502:physiology*; D002402:Cathepsin D / Q000378:metabolism; D016923:Cell Death / Q000502:physiology; D002478:Cells, Cultured; D002999:Clone Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D003546:Cysteine Endopeptidases / Q000378:metabolism; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D003593:Cytoplasm / Q000378:metabolism / Q000648:ultrastructure; D004298:Dopamine / Q000378:metabolism*; D005456:Fluorescent Dyes; D006801:Humans; D008247:Lysosomes / Q000378:metabolism / Q000648:ultrastructure; D046911:Macromolecular Substances; D009097:Multienzyme Complexes / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D009419:Nerve Tissue Proteins / Q000096:biosynthesis* / Q000235:genetics; D016716:PC12 Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D010300:Parkinson Disease / Q000378:metabolism; D046988:Proteasome Endopeptidase Complex; D011506:Proteins / Q000378:metabolism; D051381:Rats; D051843:Synucleins; D014162:Transfection; D025801:Ubiquitin / Q000378:metabolism*; D051844:alpha-Synuclein
D015853:Cysteine Proteinase Inhibitors; D005456:Fluorescent Dyes; D046911:Macromolecular Substances; D009097:Multienzyme Complexes; D009419:Nerve Tissue Proteins; D011506:Proteins; C497604:SNCA protein, human; C497606:Snca protein, rat; D051843:Synucleins; D025801:Ubiquitin; D051844:alpha-Synuclein; D003546:Cysteine Endopeptidases; D002402:Cathepsin D; D046988:Proteasome Endopeptidase Complex; D004298:Dopamine