Single Particle Characterization of Iron-induced Pore-forming -Synuclein Oligomers

CNS Research, Boehringer Ingelheim Pharma GmbH & Co. KG, CNS Research, Birkendorferstrasse 65, 88397 Biberach, Germany.
Journal of Biological Chemistry (Impact Factor: 4.57). 05/2008; 283(16):10992-1003. DOI: 10.1074/jbc.M709634200
Source: PubMed


Aggregation of alpha-synuclein is a key event in several neurodegenerative diseases, including Parkinson disease. Recent findings suggest that oligomers represent the principal toxic aggregate species. Using confocal single-molecule fluorescence techniques, such as scanning for intensely fluorescent targets (SIFT) and atomic force microscopy, we monitored alpha-synuclein oligomer formation at the single particle level. Organic solvents were used to trigger aggregation, which resulted in small oligomers ("intermediate I"). Under these conditions, Fe(3+) at low micromolar concentrations dramatically increased aggregation and induced formation of larger oligomers ("intermediate II"). Both oligomer species were on-pathway to amyloid fibrils and could seed amyloid formation. Notably, only Fe(3+)-induced oligomers were SDS-resistant and could form ion-permeable pores in a planar lipid bilayer, which were inhibited by the oligomer-specific A11 antibody. Moreover, baicalein and N'-benzylidene-benzohydrazide derivatives inhibited oligomer formation. Baicalein also inhibited alpha-synuclein-dependent toxicity in neuronal cells. Our results may provide a potential disease mechanism regarding the role of ferric iron and of toxic oligomer species in Parkinson diseases. Moreover, scanning for intensely fluorescent targets allows high throughput screening for aggregation inhibitors and may provide new approaches for drug development and therapy.

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Available from: Karin Danzer, Nov 09, 2015
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    • "In addition, their progress can be followed during aggregation. Based on the variation in the brightness of the fluorescence events, growing species can be sorted according to size [19]. If the fluorescence signals are time-correlated with themselves, so-called fluorescence correlation spectroscopy (FCS), the output curve provides information on the diffusion velocity of the labeled species and can be used to follow the oligomer growth and size distributions [20,21]. "
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    • "Transition metals, such as manganese and iron, have been suggested to be important pathogenetic factors for a number of neurodegenerative diseases, including the most frequent neurodegenerative movement disorder, PD. In particular, metal ions have not only been implicated in mediating oxidative stress, e.g., by catalyzing the Fenton reaction, but have recently also been shown to increase aggregation of asynuclein (Kostka 2008; Uversky et al. 2001). Although PD is now increasingly recognized as a system disorder with a spreading pathology (Braak et al. 2003), which finally affects most regions of the brain, the special vulnerability of the dopaminergic nigrostriatal projections is particularly intriguing. "
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    • "One possible mechanism of oligomer toxicity is the formation of lipid bilayer permeabilizing pores [14] resulting in cytotoxicity [10]–[12]. Indeed, oligomers dissociated from preformed α-syn fibrils were able to increase the conductance of lipid bilayers [15]. Formation of pores by oligomeric intermediates may be a fundamental mechanism of cell death in a large range of neurodegenerative diseases. "
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