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Temperature-dependent structural changes of Parkinson's alpha-synuclein reveal the role of pre-existing oligomers in alpha-synuclein fibrillization.

ABSTRACT: Amyloid fibrils of ?-synuclein are the main constituent of Lewy bodies deposited in substantial nigra of Parkinson's disease brains. ?-Synuclein is an intrinsically disordered protein lacking compact secondary and tertiary structures. To enhance the understanding of its structure and function relationship, we utilized temperature treatment to study ?-synuclein conformational changes and the subsequent effects. We found that after 1 hr of high temperature pretreatment, >80°C, ?-synuclein fibrillization was significantly inhibited. However, the temperature melting coupled with circular dichroism spectra showed that ?-synuclein was fully reversible and the NMR studies showed no observable structural changes of ?-synuclein after 95°C treatment. By using cross-linking and analytical ultracentrifugation, rare amount of pre-existing ?-synuclein oligomers were found to decrease after the high temperature treatment. In addition, a small portion of C-terminal truncation of ?-synuclein also occurred. The reduction of pre-existing oligomers of ?-synuclein may contribute to less seeding effect that retards the kinetics of amyloid fibrillization. Overall, our results showed that the pre-existing oligomeric species is a key factor contributing to ?-synuclein fibrillization. Our results facilitate the understanding of ?-synuclein fibrillization.

SUBMITTER: Ariesandi W

PROVIDER: S-EPMC3551866 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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Temperature-dependent structural changes of Parkinson's alpha-synuclein reveal the role of pre-existing oligomers in alpha-synuclein fibrillization.

Ariesandi Winny W Chang Chi-Fon CF Chen Tseng-Erh TE Chen Yun-Ru YR

PloS one 20130122 1

Amyloid fibrils of α-synuclein are the main constituent of Lewy bodies deposited in substantial nigra of Parkinson's disease brains. α-Synuclein is an intrinsically disordered protein lacking compact secondary and tertiary structures. To enhance the understanding of its structure and function relationship, we utilized temperature treatment to study α-synuclein conformational changes and the subsequent effects. We found that after 1 hr of high temperature pretreatment, >80°C, α-synuclein fibrilli ...[more]

PMID: 23349712

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Project description:Oligomeric forms of alpha-synuclein are emerging as key mediators of pathogenesis in Parkinson's disease. Our understanding of the exact contribution of alpha-synuclein oligomers to disease is limited by the lack of a technique for their specific detection. We describe a novel method, the alpha-synuclein proximity ligation assay, which specifically recognizes alpha-synuclein oligomers. In a blinded study with post-mortem brain tissue from patients with Parkinson's disease (n = 8, age range 73-92 years, four males and four females) and age- and sex-matched controls (n = 8), we show that the alpha-synuclein proximity ligation assay reveals previously unrecognized pathology in the form of extensive diffuse deposition of alpha-synuclein oligomers. These oligomers are often localized, in the absence of Lewy bodies, to neuroanatomical regions mildly affected in Parkinson's disease. Diffuse alpha-synuclein proximity ligation assay signal is significantly more abundant in patients compared to controls in regions including the cingulate cortex (1.6-fold increase) and the reticular formation of the medulla (6.5-fold increase). In addition, the alpha-synuclein proximity ligation assay labels very early perikaryal aggregates in morphologically intact neurons that may precede the development of classical Parkinson's disease lesions, such as pale bodies or Lewy bodies. Furthermore, the alpha-synuclein proximity ligation assay preferentially detects early-stage, loosely compacted lesions such as pale bodies in patient tissue, whereas Lewy bodies, considered heavily compacted late lesions are only very exceptionally stained. The alpha-synuclein proximity ligation assay preferentially labels alpha-synuclein oligomers produced in vitro compared to monomers and fibrils, while stained oligomers in human brain display a distinct intermediate proteinase K resistance, suggesting the detection of a conformer that is different from both physiological, presynaptic alpha-synuclein (proteinase K-sensitive) and highly aggregated alpha-synuclein within Lewy bodies (proteinase K-resistant). These disease-associated conformers represent previously undetected Parkinson's disease pathology uncovered by the alpha-synuclein proximity ligation assay.

Project description:A variety of cellular processes, including vesicle clustering in the presynaptic compartment, are impaired in Parkinson's disease and have been closely associated with α-synuclein oligomerization. Emerging evidence proves the existence of α-synuclein-related pathology in the peripheral nervous system, even though the presence of α-synuclein oligomers in situ in living patients remains poorly investigated. In this case-control study, we show previously undetected α-synuclein oligomers within synaptic terminals of autonomic fibres in skin biopsies by means of the proximity ligation assay and propose a procedure for their quantification (proximity ligation assay score). Our study revealed a significant increase in α-synuclein oligomers in consecutive patients with Parkinson's disease compared to consecutive healthy controls (P < 0.001). Proximity ligation assay score (threshold value > 96 using receiver operating characteristic) was found to have good sensitivity, specificity and positive predictive value (82%, 86% and 89%, respectively). Furthermore, to disclose the role of putative genetic predisposition in Parkinson's disease aetiology, we evaluated the differential accumulation of oligomers in a unique cohort of 19 monozygotic twins discordant for Parkinson's disease. The significant difference between patients and healthy subjects was confirmed in twins. Intriguingly, although no difference in median values was detected between consecutive healthy controls and healthy twins, the prevalence of healthy subjects positive for proximity ligation assay score was significantly greater in twins than in the consecutive cohort (47% versus 14%, P = 0.019). This suggests that genetic predisposition is important, but not sufficient, in the aetiology of the disease and strengthens the contribution of environmental factors. In conclusion, our data provide evidence that α-synuclein oligomers accumulate within synaptic terminals of autonomic fibres of the skin in Parkinson's disease for the first time. This finding endorses the hypothesis that α-synuclein oligomers could be used as a reliable diagnostic biomarker for Parkinson's disease. It also offers novel insights into the physiological and pathological roles of α-synuclein in the peripheral nervous system.

Dataset Information

Temperature-dependent structural changes of Parkinson's alpha-synuclein reveal the role of pre-existing oligomers in alpha-synuclein fibrillization.

Publications

Temperature-dependent structural changes of Parkinson's alpha-synuclein reveal the role of pre-existing oligomers in alpha-synuclein fibrillization.

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