E46K Mutant alpha-Synuclein Is Degraded by Both Proteasome and Macroautophagy Pathway

Genetic studies have revealed that rare mutations and multiplications of the gene locus in alpha-synuclein (alpha-syn) are implicated in the pathogenesis of Parkinson's disease (PD). However, the pathological effects of alpha-syn are still obscure. The neurotoxicity of alpha-syn is mainly determined by its protein levels, which depend on a balance between synthesis and degradation. Therefore, verifying the possible routes contributing to the clearance of alpha-syn is important for PD therapy. In this study, we established stable lines overexpressing human wild-type (WT) and E46K mutant alpha-syn in rat PC12 cells and investigated the degradation pathways of alpha-syn by using a panel of inhibitors and inducers of lysosome and proteasome function. We also monitored the degradation kinetics of alpha-syn by using cycloheximide to block protein synthesis. Our data showed that both proteasome and chaperon-mediated autophagy (CMA) are responsible for the degradation of the WT alpha-syn. Meanwhile, E46K mutant alpha-syn is mainly degraded by the proteasome and macroautophagy pathway. Compared with the WT protein, E46K mutant alpha-syn turned over more slowly in PC12 cells. In addition, overexpression of E46K mutant alpha-syn increased vulnerability of PC12 cells to apoptosis insults when compared with WT alpha-syn. Our findings may verify the possible routes contributing to the degradation of the E46K mutant alpha-syn.