An evaluation of the molecular model of alpha-synuclein-mediated cytotoxicity

Parkinson's disease (PD) is a neurodegenerative disorder characterized by the loss of a specific subset of neurons in the midbrain and the intracellular deposition of amyloid fibrils composed of the presynaptic protein alpha-synuclein. Three point mutants of alpha-synuclein have been linked to rare cases of familial PD, thus confirming that alpha-synuclein plays an integral role in the pathogenic process that leads to neurodegeneration. Considerable evidence indicates that aggregation of alpha-synuclein leads to a toxic gain of function, but the precise molecular mechanisms by which alpha-synuclein mediates this toxicity within cells have not been determined. One proposed model of alpha-synuclein-induced cytotoxicity involves the transient formation of soluble oligomeric species that are possible intermediates in the process of amyloid fibril formation. These protofibrils possess membrane-permeabilizing activity, and their action could account for the alpha-synuclein-dependent toxicity observed in the absence of detectable fibril formation in certain cell culture models. The recent discovery of novel variants of alpha-synuclein that may play a role in the development of both familial and sporadic PD provides an opportunity to evaluate the predictive ability of the model in terms of alpha-synuclein cellular toxicity. E46K alpha-synuclein was identified as the most recent genetic link between alpha-synuclein and PD. S129E alpha-synuclein serves as a stable mimic of serine phosphorylation of alpha-synuclein, a modification that has been associated with amyloid deposits in human brain. The D98A/Q99A double mutant was constructed to interfere with the lysosomal degradation of alpha-synuclein, in order to better understand the role of the lysosome in the regulation of alpha-synuclein toxicity. Each variant was characterized in terms of fibrillization rate, protofibril formation, membrane permeabilization activity, and yeast cell toxicity. The general correlation between the in vitro predictors of toxicity and an initial toxicity assessment in yeast supports the hypothesis that a soluble aggregate species mediates alpha-synuclein toxicity. Certain inconsistencies with the protofibril-based toxicity model suggest refinements that could improve the utility of the model in directing further research in the field.