Wild-type alpha-synuclein interacts with pro-apoptotic proteins PKCdelta and BAD to protect dopaminergic neuronal cells against MPP+-induced apoptotic cell death

Alpha-synuclein is a pre-synaptic protein of unknown function that has been implicated in the pathogenesis of Parkinson's disease (PD). Recently, we demonstrated that 1-methyl-4-phenylpyridinium (MPP+) induces caspase-3-dependent proteolytic activation of PKCdelta, which subsequently contributes to neuronal apoptotic cell death in mesencephalic dopaminergic neuronal cells. In the present study, we examined whether PKCdelta interacts with alpha-synuclein to modulate MPP+-induced dopaminergic degeneration. Over-expression of wild-type human alpha-synuclein in mesencephalic dopaminergic neuronal cells (N27 cells) attenuated MPP+-induced (300 microM) cytotoxicity, release of mitochondrial cytochrome c, and subsequent caspase-3 activation, without affecting reactive oxygen species (ROS) generation. Wild-type alpha-synuclein over-expression also dramatically reduced MPP+-induced caspase-3-mediated proteolytic cleavage of PKCdelta, whereas over-expression of the mutant human alpha-synucleinA53T did not alter the PKCdelta cleavage under similar conditions. Immunoprecipitation-kinase assay revealed reduced PKCdelta kinase activity in wild-type alpha-synuclein over-expressing cells in response to MPP+ treatment. Wild-type alpha-synuclein over-expression also rescued mesencephalic dopaminergic neuronal cells from MPP+-induced apoptotic cell death, while alpha-synucleinA53T exacerbated the MPP+-induced DNA fragmentation. Furthermore, co-immunoprecipitation studies revealed that alpha-synuclein interacts with the pro-apoptotic proteins PKCdelta and BAD, but not with the anti-apoptotic protein Bcl-2 following MPP+ treatment. We also observed that the interaction between PKCdelta and alpha-synuclein does not involve direct phosphorylation. Together, our results demonstrate that wild-type alpha-synuclein interacts with the pro-apoptotic molecules BAD and PKCdelta to protect dopaminergic neuronal cells against neurotoxic insults.