Journal
PLOS ONE
Volume 7, Issue 12, Pages -Publisher
PUBLIC LIBRARY SCIENCE
DOI: 10.1371/journal.pone.0052868
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Funding
- Research Committee of CNS Degenerative Diseases, the Ministry of Health, Labour and Welfare of Japan
- Ministry of Education, Culture, Sports, Science, and Technology, Japan [24591272, 23591252, 22591595, 21500329]
- NICHD, National Institutes of Health
- Grants-in-Aid for Scientific Research [21500329, 23591252, 22591595, 24591272] Funding Source: KAKEN
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alpha-Synuclein is the main component of Lewy bodies, the intraneuronal inclusion bodies characteristic of Parkinson's disease. Although alpha-synuclein accumulation is caused by inhibition of proteasome and autophagy-lysosome, the degradation of alpha-synuclein inclusions is still unknown. Formation of Lewy body-like inclusions can be replicated in cultured cells by introducing alpha-synuclein fibrils generated in vitro. We used this cell culture model to investigate the autophagy of alpha-synuclein inclusions and impaired mitochondria. The intracellular alpha-synuclein inclusions immediately underwent phosphorylation and ubiquitination. Simultaneously they were encircled by an adaptor protein p62/SQSTM1 and directed to the autophagy-lysosome pathway in HEK293 cell line. Most phospho-alpha-synuclein-positive inclusions were degraded in 24 h, however, lysosomal dysfunction with bafilomycin A1 significantly affected their clearance. Moreover, inhibition of autophagy by Atg-5 siRNA treatment reduced the incorporation of alpha-synuclein inclusions into LC3-positive autophagosomes. Knockdown experiments demonstrated the requirement of p62 for alpha-synuclein autophagy. These results demonstrate that alpha-synuclein inclusions are preferred targets for p62-dependent autophagy. Next, we investigated the autophagic clearance of impaired mitochondria in alpha-synuclein inclusion-containing cells. Impaired mitochondria were almost completely eliminated after mitochondrial uncoupling even in the presence of alpha-synuclein inclusions, suggesting that mitochondrial clearance is not prevented by alpha-synuclein inclusions in HEK293 cells.
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