Transient Lipid-Protein Structures and Selective Ganglioside Uptake During α-Synuclein-Lipid Co-aggregation

Abstract

α-Synuclein is a membrane-interacting protein involved in Parkinson’s disease. Here we have investigated the co-association of α-synuclein and lipids from ganglioside-containing model membranes. Our study relies on the reported importance of ganglioside lipids, which are found in high amounts in neurons and exosomes, on cell-to-cell prion-like transmission of misfolded α-synuclein. Samples taken along various stages of the aggregation process were imaged using cryogenic transmission electron microscopy, and the composition of samples corresponding to the final state analyzed using NMR spectroscopy. The combined data shows that α-synuclein co-assembles with lipids from the ganglioside (GM1)-containing model membranes. The lipid-protein samples observed during the aggregation process contain non-vesicular objects not present at the final stage, thus capturing the co-existence of species under non-equilibrium conditions. A range of different lipid-protein co-assemblies are observed during the time course of the reaction and some of these appear to be transient assemblies that evolve into other co-aggregates over time. At the end of the aggregation reaction, the samples become more homogeneous, showing thin fibrillar structures heavily decorated with small vesicles. From the NMR analysis, we conclude that the ratio of GM1 to phosphatidyl choline (PC) in the supernatant of the co-aggregated samples is significantly reduced compared to the GM1/PC ratio of the lipid dispersion from which these samples were derived. Taken together, this indicates a selective uptake of GM1 into the fibrillar aggregates and removal of GM1-rich objects from the solution.

Dual Effects of Presynaptic Membrane Mimetics on α-Synuclein Amyloid Aggregation

“…For example, αSN 130CF amyloid fibrils formed in the presence of 3 mM of DOPC lipids were mostly composed of antiparallel β -strands, while amyloid fibrils generated with 3 mM of Mimic lipids contained both parallel and antiparallel β -strands ( Supplementary Figures S8,S9 ). Along the same lines, previous studies have reported that alterations in either lipid concentrations or liposome compositions can induce morphologically distinct amyloid fibrils ( Kinoshita et al, 2017 ; Gaspar et al, 2021 ). Amyloid fibrils with different structures showed type-specific fluorescence intensity due to different binding sites of ThT on the surfaces of the amyloid fibrils ( Sidhu et al, 2018 ).…”

Comparing α-Synuclein Fibrils Formed in the Absence and Presence of a Model Lipid Membrane: A Small and Wide-Angle X-Ray Scattering Study

“…There are also several other reports on the formation of lipid-protein coaggregates. Both in the case of the early stage of aggregation, involving membrane disruption and protein oligomer formation (Fusco et al, 2014;van Maarschalkerweerd et al, 2014;Fusco et al, 2016;Hannestad et al, 2020), and in the case of mature fibrils (van Rooijen et al, 2009;Grey et al, 2011;Gaspar et al, 2021). However, the very nature of co-aggregation is not fully clear in the case of mature fibrils.…”

“…In vivo, the fibril formation process occurs in a highly lipid-rich environment, and Lewy bodies are known to contain significant amounts of lipid (Shults, 2006;Stefanis, 2012;Araki et al, 2019;Fanning et al, 2020;Lashuel, 2020). These facts have motivated studies of αS-lipid interactions, involving both monomeric (Fusco et al, 2014;van Maarschalkerweerd et al, 2014;Fusco et al, 2016;Hannestad et al, 2020;Makasewicz et al, 2021), andfibrillar αS (van Rooijen et al, 2009;Hellstrand et al, 2013b;Galvagnion et al, 2019;Gaspar et al, 2021). It has been shown that monomeric αS adsorbs to anionic lipid membranes (Jao et al, 2004;Pfefferkorn et al, 2012;Hellstrand et al, 2013a;Fusco et al, 2014), and that the membrane under some conditions can have a catalytic effect on fibril formation (Galvagnion et al, 2015;Grey et al, 2015;Galvagnion, 2017;Gaspar et al, 2019).…”

“…Interestingly, this adsorption appears to be a strongly cooperative process (Makasewicz et al, 2021). Several studies, performed with different systems, have also indicated that αS and lipids may form coaggregates (Reynolds et al, 2011;Hellstrand et al, 2013b;van Maarschalkerweerd et al, 2014;Galvagnion et al, 2019;Gaspar et al, 2021). The exact nature of such aggregates still remains unclear.…”

On the Cluster Formation of α-Synuclein Fibrils

“…The demonstration that the main component of Lewy bodies is a β-sheet-rich, fibrillar form of αS ( Shults 2006 ; Araki, Yagi et al, 2019 ; Lashuel 2020 ), has motivated extensive studies of αS fibrils ( Waxman and Giasson 2009 ; Alam, Bousset et al, 2019 ; Guerrero-Ferreira, Kovacik et al, 2020 ). It has also been shown that Lewy bodies contain membrane lipids ( Lashuel 2020 ; Mahul-Mellier, Burtscher et al, 2020 ), which has motivated detailed studies on interaction between αS and lipids ( Pfefferkorn, Jiang et al, 2012 ; Andreasen, Lorenzen et al, 2015 ; Iyer and Claessens 2019 ; Lashuel 2020 ), covering systems where the protein is present in the monomeric state ( Jain, Bhasne et al, 2013 ; Fusco, De Simone et al, 2014 ; Fusco, Pape et al, 2016 ; Hannestad, Rocha et al, 2020 ), during the aggregation ( Jiang, de Messieres et al, 2013 ; Galvagnion, Brown et al, 2016 ; Gaspar, Pallbo et al, 2018 ) as well as in the final amyloid aggregates ( Hellstrand et al, 2013b ; Galvagnion, Topgaard et al, 2019 ; Gaspar, Idini et al, 2021 ).…”