Pathological structural conversion of alpha-synuclein at the mitochondria induces neuronal toxicity

Aggregation of alpha-synuclein (alpha-Syn) drives Parkinson's disease (PD), although the initial stages of self-assembly and structural conversion have not been directly observed inside neurons. In this study, we tracked the intracellular conformational states of alpha-Syn using a single-molecule Forster resonance energy transfer (smFRET) biosensor, and we show here that alpha-Syn converts from a monomeric state into two distinct oligomeric states in neurons in a concentration-dependent and sequence-specific manner. Three-dimensional FRET-correlative light and electron microscopy (FRET-CLEM) revealed that intracellular seeding events occur preferentially on membrane surfaces, especially at mitochondrial membranes. The mitochondrial lipid cardiolipin triggers rapid oligomerization of A53T alpha-Syn, and cardiolipin is sequestered within aggregating lipid-protein complexes. Mitochondrial aggregates impair complex I activity and increase mitochondrial reactive oxygen species (ROS) generation, which accelerates the oligomerization of A53T alpha-Syn and causes permeabilization of mitochondrial membranes and cell death. These processes were also observed in induced pluripotent stem cell (iPSC)-derived neurons harboring A53T mutations from patients with PD. Our study highlights a mechanism of de novo alpha-Syn oligomerization at mitochondrial membranes and subsequent neuronal toxicity. This study tracked the initial self-assembly, oligomerization and structural conversion of alpha-synuclein inside neurons. Early seeding events occur on mitochondrial membranes, where oligomerization induces mitochondrial dysfunction and neuronal loss.