Cell-produced alpha-synuclein oligomers are targeted to, and impair, the 26S proteasome.
Proteasomal dysfunction may play a role in neurodegenerative conditions and protein aggregation. Overexpression in neuronal cells of alpha-synuclein, a molecule linked to Parkinson's Disease, may lead to proteasomal dysfunction. Using PC12 cells stably expressing wild-type or mutant alpha-synuclein and gel filtration, we demonstrate that soluble, intermediate size oligomers of alpha-synuclein co-elute with the 26S proteasome. These soluble oligomers associate with the 26S proteasome and are significantly increased following treatment with proteasomal, but not lysosomal, inhibitors, indicating specific degradation of these particular species by the 26S proteasome. Importantly, expression of alpha-synuclein resulted in a significant inhibition of all proteasomal activities without affecting the levels or assembly of the 26S proteasome. Pharmacological dissociation of alpha-synuclein oligomers restored proteasomal function and reduced polyubiquitinated protein load in intact cells. Our findings suggest a model where only a subset of specific soluble cell-derived alpha-synuclein oligomers is targeted to the 26S proteasome for degradation, and simultaneously inhibit its function, likely by impeding access of other proteasomal substrates.
Emmanouilidou
Evangelia
E
Division of Basic Neurosciences, Biomedical Research Foundation of the Academy of Athens (BRFAA), Athens, Greece.
Stefanis
Leonidas
L
Vekrellis
Kostas
K
D000409:Alanine / Q000235:genetics; D000643:Ammonium Chloride / Q000494:pharmacology; D000818:Animals; D016227:Benzoquinones / Q000494:pharmacology; D002540:Cerebral Cortex / Q000187:drug effects / Q000201:enzymology; D002850:Chromatography, Gel / Q000379:methods; D004396:Coloring Agents / Q000494:pharmacology; D003224:Congo Red / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D049452:Green Fluorescent Proteins / Q000235:genetics; D006801:Humans; D047468:Immunoprecipitation / Q000379:methods; D047029:Lactams, Macrocyclic / Q000494:pharmacology; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D009154:Mutation / Q000235:genetics; D016716:PC12 Cells / Q000378:metabolism; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D017510:Protein Folding; D051381:Rats; D014162:Transfection / Q000379:methods; D014443:Tyrosine / Q000235:genetics; D051844:alpha-Synuclein / Q000235:genetics / Q000378:metabolism*
D016227:Benzoquinones; D004396:Coloring Agents; D004791:Enzyme Inhibitors; D047029:Lactams, Macrocyclic; D051844:alpha-Synuclein; C485184:enhanced green fluorescent protein; D000643:Ammonium Chloride; D049452:Green Fluorescent Proteins; D003224:Congo Red; D014443:Tyrosine; D046988:Proteasome Endopeptidase Complex; C061553:ATP dependent 26S protease; D000409:Alanine; C001277:geldanamycin