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Stabilization of ?-Synuclein Fibril Clusters Prevents Fragmentation and Reduces Seeding Activity and Toxicity.

ABSTRACT: Protein misfolding results in the accumulation of aggregated ?-sheet-rich structures in Parkinson's disease (PD) and Alzheimer's disease. The toxic oligomer hypothesis stipulates that prefibrillar assemblies, such as soluble oligomers or protofibrils, are responsible for the poor prognosis of these diseases. Previous studies demonstrated that a small molecule related to the natural compound orcein, O4, directly binds to amyloid-? fibrils and stabilizes them, accelerating the formation of end-stage mature fibrils. Here we demonstrate a similar phenomenon during O4 treatment of ?-synuclein (?syn) aggregates, the protein responsible for PD pathology. While the drug did not change the kinetics of aggregate formation as measured by the amyloidophilic dye thioflavin T, O4 depleted ?syn oligomers and promoted the formation of sodium dodecyl sulfate and proteinase K resistant aggregates consisting of large fibril clusters. These fibril clusters exhibited reduced toxicity to human neuronal model cells and reduced seeding activity in vitro. The effectiveness of O4 decreased when it was added at later points in the ?syn aggregation pathway, which suggests that the incorporation of O4 into fibril assemblies stabilizes them against chemical, enzymatic, and mechanic degradation. These findings suggest that small molecules, which stabilize amyloid fibrils, can prevent fibril fragmentation and seeding and consequently prevent prion-like replication of misfolded ?syn. Inhibiting prion replication by fibril stabilization could thus be a therapeutic strategy for PD.

SUBMITTER: Lam HT

PROVIDER: S-EPMC5706464 | biostudies-literature | 2016 Feb

REPOSITORIES: biostudies-literature

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Stabilization of α-Synuclein Fibril Clusters Prevents Fragmentation and Reduces Seeding Activity and Toxicity.

Lam Huy T HT Graber Michael C MC Gentry Katherine A KA Bieschke Jan J

Biochemistry 20160122 4

Protein misfolding results in the accumulation of aggregated β-sheet-rich structures in Parkinson's disease (PD) and Alzheimer's disease. The toxic oligomer hypothesis stipulates that prefibrillar assemblies, such as soluble oligomers or protofibrils, are responsible for the poor prognosis of these diseases. Previous studies demonstrated that a small molecule related to the natural compound orcein, O4, directly binds to amyloid-β fibrils and stabilizes them, accelerating the formation of end-sta ...[more]

PMID: 26799377

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Project description:The role of extracellular ?-synuclein (?-syn) in the initiation and the spreading of neurodegeneration in Parkinson's disease (PD) has been studied extensively over the past 10 years. However, the nature of the ?-syn toxic species and the molecular mechanisms by which they may contribute to neuronal cell loss remain controversial. In this study, we show that fully characterized recombinant monomeric, fibrillar or stabilized forms of oligomeric ?-syn do not trigger significant cell death when added individually to neuroblastoma cell lines. However, a mixture of preformed fibrils (PFFs) with monomeric ?-syn becomes toxic under conditions that promote their growth and amyloid formation. In hippocampal primary neurons and ex vivo hippocampal slice cultures, ?-syn PFFs are capable of inducing a moderate toxicity over time that is greatly exacerbated upon promoting fibril growth by addition of monomeric ?-syn. The causal relationship between ?-syn aggregation and cellular toxicity was further investigated by assessing the effect of inhibiting fibrillization on ?-syn-induced cell death. Remarkably, our data show that blocking fibril growth by treatment with known pharmacological inhibitor of ?-syn fibrillization (Tolcapone) or replacing monomeric ?-syn by monomeric ?-synuclein in ?-syn mixture composition prevent ?-syn-induced toxicity in both neuroblastoma cell lines and hippocampal primary neurons. We demonstrate that exogenously added ?-syn fibrils bind to the plasma membrane and serve as nucleation sites for the formation of ?-syn fibrils and promote the accumulation and internalization of these aggregates that in turn activate both the extrinsic and intrinsic apoptotic cell death pathways in our cellular models. Our results support the hypothesis that ongoing aggregation and fibrillization of extracellular ?-syn play central roles in ?-syn extracellular toxicity, and suggest that inhibiting fibril growth and seeding capacity constitute a viable strategy for protecting against ?-syn-induced toxicity and slowing the progression of neurodegeneration in PD and other synucleinopathies.

Dataset Information

Stabilization of ?-Synuclein Fibril Clusters Prevents Fragmentation and Reduces Seeding Activity and Toxicity.

Publications

Stabilization of α-Synuclein Fibril Clusters Prevents Fragmentation and Reduces Seeding Activity and Toxicity.

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