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An intact membrane is essential for small extracellular vesicle-induced modulation of ?-synuclein fibrillization.

ABSTRACT: The misfolding and fibrillization of the protein, ?-synuclein (?syn), is associated with neurodegenerative disorders referred to as the synucleinopathies. Understanding the mechanisms of ?syn misfolding is an important area of interest given that ?syn misfolding contributes to disease pathogenesis. While many studies report the ability of synthetic lipid membranes to modulate ?syn folding, there is little data pertaining to the mechanism(s) of this interaction. ?Syn has previously been shown to associate with small lipid vesicles released by cells called extracellular vesicles (EVs) and it is postulated these interactions may assist in the spreading of pathological forms of this protein. Together, this presents the need for robust characterisation studies on ?syn fibrillization using biologically-derived vesicles. In this study, we comprehensively characterised the ability of lipid-rich small extracellular vesicles (sEVs) to alter the misfolding of ?syn induced using the Protein Misfolding Cyclic Amplification (PMCA) assay. The biochemical and biophysical properties of misfolded ?syn were examined using a range of techniques including: Thioflavin T fluorescence, transmission electron microscopy, analytical centrifugation and western immunoblot coupled with protease resistance assays and soluble/insoluble fractionation. We show that sEVs cause an acceleration in ?syn fibrillization and provide comprehensive evidence that this results in an increase in the abundance of mature insoluble fibrillar species. In order to elucidate the relevance of the lipid membrane to this interaction, sEV lipid membranes were modified by treatment with methanol, or a combination of methanol and sarkosyl. These treatments altered the ultrastructure of the sEVs without changing the protein cargo. Critically, these modified sEVs had a reduced ability to influence ?syn fibrillization compared to untreated counterparts. This study reports the first comprehensive examination of ?syn:EV interactions and demonstrates that sEVs are powerful modulators of ?syn fibrillization, which is mediated by the sEV membrane. In doing so, this work provides strong evidence for a role of sEVs in contributing directly to ?syn misfolding in the synucleinopathy disorders.

SUBMITTER: Ugalde CL

PROVIDER: S-EPMC7726797 | biostudies-literature | 2020 Dec

REPOSITORIES: biostudies-literature

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An intact membrane is essential for small extracellular vesicle-induced modulation of α-synuclein fibrillization.

Ugalde Cathryn L CL Gordon Shane E SE Shambrook Mitch M Nasiri Kenari Amirmohammad A Coleman Bradley M BM Perugini Matthew A MA Lawson Victoria A VA Finkelstein David I DI Hill Andrew F AF

Journal of extracellular vesicles 20201210 2

The misfolding and fibrillization of the protein, α-synuclein (αsyn), is associated with neurodegenerative disorders referred to as the synucleinopathies. Understanding the mechanisms of αsyn misfolding is an important area of interest given that αsyn misfolding contributes to disease pathogenesis. While many studies report the ability of synthetic lipid membranes to modulate αsyn folding, there is little data pertaining to the mechanism(s) of this interaction. αSyn has previously been shown to ...[more]

PMID: 33318779

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Project description:The most established pathognomonic protein of Parkinson's disease (PD), α-synuclein, is extensively investigated for disease diagnosis and prognosis; however, investigations into whether the free form of α-synuclein in the blood functions as a PD biomarker have not been fruitful. Extracellular vesicles (EVs) secreted from cells and present in blood transport molecules are novel platforms for biomarker identification. In blood EVs, α-synuclein originates predominantly from the brain without the interference of the blood-brain barrier. The present study investigated the role of plasma EV-borne α-synuclein as a biomarker of PD. Patients with mild to moderate stages of PD (n = 116) and individuals without PD (n = 46) were recruited to serve as the PD study group and the control group, respectively. Plasma EVs were isolated, and immunomagnetic reduction-based immunoassay was used to assess EV α-synuclein levels. Conventional statistical analysis was performed using SPSS 25.0, and p < 0.05 was considered significant. Compared with controls, we observed significantly lower plasma EV α-synuclein levels in the patients with PD (PD: 56.0 ± 3.7 fg/mL vs. control: 74.5 ± 4.3 fg/mL, p = 0.009), and the significance remained after adjustment for age and sex. Plasma EV α-synuclein levels in the patients with PD did not correlate with age, disease duration, Part I and II scores of the Unified Parkinson's Disease Rating Scale (UPDRS), or the Mini-Mental State Examination scores. However, such levels were significantly correlated with UPDRS Part III score, which assesses motor dysfunction. Furthermore, the severity of akinetic-rigidity symptoms, but not tremor, was inversely associated with plasma EV α-synuclein level. Plasma EV α-synuclein was significantly different between the control and PD group and was associated with akinetic-rigidity symptom severity in patients with PD. This study corroborates the possible diagnostic and subtyping roles of plasma EV α-synuclein in patients with PD, and it further provides a basis for this protein's clinical relevance and feasibility as a PD biomarker.

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An intact membrane is essential for small extracellular vesicle-induced modulation of ?-synuclein fibrillization.

Publications

An intact membrane is essential for small extracellular vesicle-induced modulation of α-synuclein fibrillization.

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