Abstract
Misfolding and aggregation of α-synuclein into toxic soluble oligomeric α-synuclein aggregates has been strongly correlated with the pathogenesis of Parkinson’s disease (PD). Here, we show that two different morphologically distinct oligomeric α-synuclein aggregates are present in human post-mortem PD brain tissue and are responsible for the bulk of α-synuclein induced toxicity in brain homogenates from PD samples. Two antibody fragments that selectively bind the different oligomeric α-synuclein variants block this α-synuclein induced toxicity and are useful tools to probe how various cell models replicate the α-synuclein aggregation pattern of human PD brain. Using these reagents, we show that mammalian cell type strongly influences α-synuclein aggregation, where neuronal cells best replicate the PD brain α-synuclein aggregation profile. Overexpression of α-synuclein in the different cell lines increased protein aggregation but did not alter the morphology of the oligomeric aggregates generated. Differentiation of the neuronal cells into a cholinergic-like or dopaminergic-like phenotype increased the levels of oligomeric α-synuclein where the aggregates were localized in cell neurites and cell bodies.
Highlights
Parkinson’s disease (PD) is a progressive neurodegenerative disorder that affects approximately 1%of people aged 65 and older [1]
The -synuclein protein occurs in vivo in various forms and morphologies [8,38,39], and can interact with membranes [40,41,42], other proteins such as tau, p25alpha, tubulin, and transcription factor ELK-1 [43,44,45], metal ions including aluminum, copper, calcium, and iron [19], and catecholamines such as dopamine [15,46]
While ample evidence indicates that -synuclein plays an important role in the pathogenesis of PD, the impact of the various conformations of -synuclein in the progression of PD is much debated and poorly understood
Summary
Parkinson’s disease (PD) is a progressive neurodegenerative disorder that affects approximately 1%. While several morphologies of -synuclein can be generated in vitro, increasing evidence suggest that different soluble oligomeric species rather than mature fibril forms are responsible for neuronal dysfunction and toxicity in PD disease [20,21,22,23,24]. The increased cytotoxicity could be largely blocked in a concentration dependent manner by addition of D5 and/or 10H, indicating that most of the increased neuronal toxicity in human PD brain tissue samples compared to age matched control samples is attributable to the presence of specific oligomeric -synuclein species. We probed for the presence of both the D5 and 10H reactive toxic -synuclein aggregates in different mammalian cell lines expressing endogenous levels of -synuclein including non-differentiated and differentiated human neuroblastoma cells (SH-SY5Y), Chinese hamster ovary (CHO) cells and human embryonic kidney (HEK) cells. We show that the choice of cell model and differentiation state can quite dramatically impact the -synuclein aggregation process
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