Polymorphic alpha-Synuclein Aggregates Mediate Differential Cellular Toxicity Profiles

Aggregate of α-Synuclein is a major component of Lewy body, a pathological hallmark of Parkinson’s disease. Studies show different α-Synuclein’s polymorphs exhibited differential neuronal toxicities. To investigate detailed structural architecture of the polymorphs and its relationship with neuronal cell damage and underlying mechanism(s), we have produced distinct polymorphic forms of α-synuclein aggregates in vitro conditions by incubating with varied temperature and salt treatment. Aggregation kinetics and morphological characteristics were thoroughly characterized using Thioflavin T (ThT) fluorescence assay, ANS binding assays, and Atomic Force Microscopy (AFM), to understand the aggregation patterns and distinct aggregate morphologies respectively, followed by their impact on cellular survivability. Varied thermal stabilities of polymorphs as reflected by different melting temperature (Tm) values of 57.51°C and 63.33°C, investigated by Differential Scanning Calorimetry (DSC) suggests differences in aggregate core compactness and internal architecture. Thioflavin T (ThT), 8-Anilinonaphthalene-1-sulfonic acid (ANS) assay showed dissimilar aggregation patterns resulting in different morphologies of aggregates, confirmed by AFM data. Cellular toxicity assays (MTT assay) demonstrated variable degrees of cytotoxicity across polymorphs, implying that structural compactness and molecular arrangements may be key determinants of toxic potential. Our findings show how structural polymorphism in α-synuclein aggregates underlies differential neuronal damaging mechanisms and contributes to disease heterogeneity.

Shubham Kundu acknowledges Council of Scientific and Industrial Research (CSIR), Govt. of India for providing fellowship.