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- Title
α-Synuclein Phosphorylation Enhances Eosinophilic Cytoplasmic Inclusion Formation in SH-SY5Y Cells.
- Authors
Smith, Wanli W.; Margolis, Russell L.; Xiaojie Li; Troncoso, Juan C.; Lee, Michael K.; Dawson, Valina L.; Dawson, Ted M.; Iwatsubo, Takashi; Ross, Christopher A.
- Abstract
Parkinson's disease (PD) is a neurodegenerative disorder characterized by selective loss of dopaminergic neurons and the presence of Lewy bodies. Previous reports have shown that α-synuclein deposited in brain tissue from individuals with synucleinopathy is extensively phosphorylated at Ser-129. Here, we investigate the role of phosphorylation of α-synuclein in the formation of inclusions involving synphilin-1 and parkin using site-directed mutagenesis to change Set-129 of α-synuclein to alanine (S129A) to abolish phosphorylation at this site. Coexpression of wild-type α-synuclein and synphilin-1 in human neuroblastoma SH-SY5Y cells yielded cytoplasmic eosinophilic inclusions with some features resembling Lewy bodies, whereas coexpression of S129A α-synuclein and synphlin-1 formed few or no inclusions. Moreover, coexpression of parkin with α-synuclein and synphilin-1 formed more ubiquitinated inclusions, but these inclusions decreased with expression of S129A α-synuclein instead of wild-type α-synuclein. Coimmunoprecipitation assays revealed a decreased interaction of S129A α-synuclein with synphilin-1 compared with wild-type α-synuclein. Expression of S129A α-synuclein instead of wild-type α-synuclein also decreased the association of synphilin-1 and parkin and subsequently reduced the parkin-mediated ubiquitination of synphilin-1 and the formation of ubiquitinated inclusions. Treatment of SH-SY5Y cells with H2O2 increased α-synuclein phosphorylation and enhanced the formation of inclusions formed by coexpression of α-synuclein, synphilin-1, and parkin, whereas treatment with the casein kinase 2 inhibitor 5,6-dichloro-1-β-D-ribofuranosylbenzimidazole had the opposite affect. These results indicate that phosphorylation of α-synuclein at S129 may be important for the formation of inclusions in PD and related α synucleinopathies.
- Subjects
PHOSPHORYLATION; PARKINSON'S disease; NEURODEGENERATION; DOPAMINERGIC neurons; LEWY body dementia; NEUROBLASTOMA
- Publication
Journal of Neuroscience, 2005, Vol 25, Issue 23, p5544
- ISSN
0270-6474
- Publication type
Article
- DOI
10.1523/JNEUROSCI.0482-05.2005