Search

Accumulation of filamentous aggregates of α-synuclein is a pathological hallmark of several neurodegenerative diseases, including Parkinson's disease (PD). The interaction between α-synuclein and phospholipids has been shown to play a critical role in the aggregation of α-synuclein. Most structural studies have, however, been focused on α-synuclein filaments formed in the absence of lipids. Here, we report the structural investigation of α-synuclein filaments assembled under the quiescent condition in the presence of anionic lipid vesicles using electron microscopy (EM), including cryogenic electron microscopy (cryo-EM). Our transmission electron microscopy (TEM) analyses reveal that α-synuclein forms curly protofilaments at an early stage of aggregation. The flexible protofilaments were then converted to long filaments after a longer incubation of 30 days. More detailed structural analyses using cryo-EM reveal that the long filaments adopt untwisted structures with different diameters, which have not been observed in previous α-synuclein fibrils formed in vitro. The untwisted filaments are rather similar to straight filaments with no observable twist that are extracted from patients with dementia with Lewy bodies. Our structural studies highlight the conformational diversity of α-synuclein filaments, requiring additional structural investigation of not only more ex vivo α-synuclein filaments but also in vitro α-synuclein filaments formed in the presence of diverse cofactors to better understand the molecular basis of diverse molecular conformations of α-synuclein filaments.

An increasing body of evidence suggests that aggregation-prone proteins associated with various neurodegenerative diseases synergistically promote their mutual aggregation, leading to the co-occurrence of multiple neurodegenerative diseases in the same patient. Here we investigated teh molecular basis of synergistic interactions between the two pathological proteins, tau and α-synuclein, using various biophysical techniques including transmission electron microscopy (TEM), circular dichroism (CD), and solution and solid-state NMR. Our biophysical analyses of α-synuclein aggregation in the absence and presence of tau reveal that tau monomers promote the formation of α-synuclein oligomers and subsequently fibril formation. Solution NMR results also indicate that monomeric forms of tau selectively interact with the C-terminal region of the α-synuclein monomer, accelerating α-synuclein aggregation. In addition, a combined use of TEM and solid-state NMR spectroscopy reveals that the synergistic interactions lead to the formation of toxic α-synuclein aggregates with a distinct morphology and molecular conformation. The filamentous α-synuclein aggregates as well as α-synuclein monomers were also able to induce tau aggregation.

Tau aggregation is a pathological hallmark of Alzheimer's disease, Parkinson's disease, and many other neurodegenerative disorders known as tauopathies. Tau aggregates take on many forms, and their formation is a multistage process with intermediate stages. Recently, tau oligomers have emerged as the pathogenic species in tauopathies and a possible mediator of amyloid-β toxicity in Alzheimer's disease. Here, we use a novel, physiologically relevant method (oligomer cross-seeding) to prepare homogeneous populations of tau oligomers and characterize these oligomers in vitro. We show that both Aβ and α-synuclein oligomers induce tau aggregation and the formation of β-sheet-rich neurotoxic tau oligomers.

Amyloid plaques are hallmark neuropathological lesions in Alzheimer's disease, which consist of abnormally aggregated Abeta protein. Multiple Abeta aggregated species have been identified, and neurotoxicity appears to be correlated with the amount of nonfibrillar oligomers. Therefore, selective inhibition of Abeta oligomer formation has emerged as an attractive means of therapeutic intervention. To investigate whether small molecules can modulate aggregation to achieve selective inhibition of neurotoxic amyloid oligomers, Abeta aggregation was assayed in vitro in the presence of methylene blue, using immunoreactivity with the prefibrillar oligomer-specific antibody A11, transmission electron microscopy, and turbidity assays. Methylene blue inhibited oligomerization when used at substoichiometric concentrations relative to that of the Abeta monomer. Inhibition of Abeta oligomerization was achieved concomitant with promotion of fibrillization, suggesting that oligomer and fibril formation are distinct and competing pathways. Methylene blue-mediated promotion of fiber formation occurred via a dose-dependent decrease in the lag time and an increase in the fibrillization rate, consistent with promotion of both filament nucleation and elongation. Addition of methylene blue to preformed oligomers resulted in oligomer loss and promotion of fibrillization. The data show that Abeta oligomer formation is inhibited by promoting fibril formation, which suggests that the relative pathological significance of oligomers and fibrils may be tested in vivo using methylene blue. If Abeta oligomers represent the primary pathogenic species, then inhibition of this highly toxic species via promotion of formation of less toxic aggregates may be therapeutically useful.