A novel ?-synuclein missense mutation in Parkinson disease.
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ABSTRACT:
SUBMITTER: Proukakis C
PROVIDER: S-EPMC3653201 | biostudies-literature | 2013 Mar
REPOSITORIES: biostudies-literature
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Project description:?-Sheet-rich ?-synuclein (?S) aggregates characterize Parkinson's disease (PD). ?S was long believed to be a natively unfolded monomer, but recent work suggests it also occurs in ?-helix-rich tetramers. Crosslinking traps principally tetrameric ?S in intact normal neurons, but not after cell lysis, suggesting a dynamic equilibrium. Here we show that freshly biopsied normal human brain contains abundant ?S tetramers. The PD-causing mutation A53T decreases tetramers in mouse brain. Neurons derived from an A53T patient have decreased tetramers. Neurons expressing E46K do also, and adding 1-2 E46K-like mutations into the canonical ?S repeat motifs (KTKEGV) further reduces tetramers, decreases ?S solubility and induces neurotoxicity and round inclusions. The other three fPD missense mutations likewise decrease tetramer:monomer ratios. The destabilization of physiological tetramers by PD-causing missense mutations and the neurotoxicity and inclusions induced by markedly decreasing tetramers suggest that decreased ?-helical tetramers and increased unfolded monomers initiate pathogenesis. Tetramer-stabilizing compounds should prevent this.
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Project description:Deposition of the pathological α-synuclein (αSynP) in the brain is the hallmark of synucleinopathies, including Parkinson disease (PD), Lewy body dementia (LBD), and multiple system atrophy (MSA). Whether real-time quaking-induced conversion (RT-QuIC) and protein misfolding cyclic amplification (PMCA) assays can sensitively detect skin biomarkers for PD and non-PD synucleinopathies remains unknown. To develop sensitive and specific skin biomarkers for antemortem diagnosis of PD and other synucleinopathies. This retrospective and prospective diagnostic study evaluated autopsy and biopsy skin samples from neuropathologically and clinically diagnosed patients with PD and controls without PD. Autopsy skin samples were obtained at 3 medical centers from August 2016 to September 2019, and biopsy samples were collected from 3 institutions from August 2018 to November 2019. Based on neuropathological and clinical diagnoses, 57 cadavers with synucleinopathies and 73 cadavers with nonsynucleinopathies as well as 20 living patients with PD and 21 living controls without PD were included. Specifically, cadavers and participants had PD, LBD, MSA, Alzheimer disease, progressive supranuclear palsy, or corticobasal degeneration or were nonneurodegenerative controls (NNCs). A total of 8 approached biopsy participants either refused to participate in or were excluded from this study due to uncertain clinical diagnosis. Data were analyzed from September 2019 to April 2020. Skin αSynP seeding activity was analyzed by RT-QuIC and PMCA assays. A total of 160 autopsied skin specimens from 140 cadavers (85 male cadavers [60.7%]; mean [SD] age at death, 76.8 [10.1] years) and 41 antemortem skin biopsies (27 male participants [66%]; mean [SD] age at time of biopsy, 65.3 [9.2] years) were analyzed. RT-QuIC analysis of αSynP seeding activity in autopsy abdominal skin samples from 47 PD cadavers and 43 NNCs revealed 94% sensitivity (95% CI, 85-99) and 98% specificity (95% CI, 89-100). As groups, RT-QuIC also yielded 93% sensitivity (95% CI, 85-97) and 93% specificity (95% CI, 83-97) among 57 cadavers with synucleinopathies (PD, LBD, and MSA) and 73 cadavers without synucleinopathies (Alzheimer disease, progressive supranuclear palsy, corticobasal degeneration, and NNCs). PMCA showed 82% sensitivity (95% CI, 76-88) and 96% specificity (95% CI, 85-100) with autopsy abdominal skin samples from PD cadavers. From posterior cervical and leg skin biopsy tissues from patients with PD and controls without PD, the sensitivity and specificity were 95% (95% CI, 77-100) and 100% (95% CI, 84-100), respectively, for RT-QuIC and 80% (95% CI, 49-96) and 90% (95% CI, 60-100) for PMCA. This study provides proof-of-concept that skin αSynP seeding activity may serve as a novel biomarker for antemortem diagnoses of PD and other synucleinopathies.
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Project description:Endosomal sorting is a highly orchestrated cellular process. Retromer is a heterotrimeric complex that associates with endosomal membranes and facilitates the retrograde sorting of multiple receptors, including the cation-independent mannose 6-phosphate receptor for lysosomal enzymes. The cycling of retromer on and off the endosomal membrane is regulated by a network of retromer-interacting proteins. Here, we find that Parkinson disease-associated Vps35 variant, R524W, but not P316S, is a loss-of-function mutation as marked by a reduced association with this regulatory network and dysregulation of endosomal receptor sorting. Expression of Vps35 R524W-containing retromer results in the accumulation of intracellular α-synuclein-positive aggregates, a hallmark of Parkinson disease. Overall, the Vps35 R524W-containing retromer has a decreased endosomal association, which can be partially rescued by R55, a small molecule previously shown to stabilize the retromer complex, supporting the potential for future targeting of the retromer complex in the treatment of Parkinson disease.
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Project description:Protein aggregation within the central nervous system has been recognized as a defining feature of neurodegenerative diseases since the early 20th century. Since that time, there has been a growing list of neurodegenerative disorders, including Parkinson disease, which are characterized by inclusions of specific pathogenic proteins. This has led to the long-held dogma that these characteristic protein inclusions, which are composed of large insoluble fibrillar protein aggregates and visible by light microscopy, are responsible for cell death in these diseases. However, the correlation between protein inclusion formation and cytotoxicity is inconsistent, suggesting that another form of the pathogenic proteins may be contributing to neurodegeneration. There is emerging evidence implicating soluble oligomers, smaller protein aggregates not detectable by conventional microscopy, as potential culprits in the pathogenesis of neurodegenerative diseases. The protein ?-synuclein is well recognized to contribute to the pathogenesis of Parkinson disease and is the major component of Lewy bodies and Lewy neurites. However, ?-synuclein also forms oligomeric species, with certain conformations being toxic to cells. The mechanisms by which these ?-synuclein oligomers cause cell death are being actively investigated, as they may provide new strategies for diagnosis and treatment of Parkinson disease and related disorders. Here we review the possible role of ?-synuclein oligomers in cell death in Parkinson disease and discuss the potential clinical implications.
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Project description:Intraneuronal accumulation of aggregated α-synuclein is a pathological hallmark of Parkinson's disease. Therefore, mechanisms capable of promoting α-synuclein deposition bear important pathogenetic implications. Mutations of the glucocerebrosidase 1 (GBA) gene represent a prevalent Parkinson's disease risk factor. They are associated with loss of activity of a key enzyme involved in lipid metabolism, glucocerebrosidase, supporting a mechanistic relationship between abnormal α-synuclein-lipid interactions and the development of Parkinson pathology. In this study, the lipid membrane composition of fibroblasts isolated from control subjects, patients with idiopathic Parkinson's disease and Parkinson's disease patients carrying the L444P GBA mutation (PD-GBA) was assayed using shotgun lipidomics. The lipid profile of PD-GBA fibroblasts differed significantly from that of control and idiopathic Parkinson's disease cells. It was characterized by an overall increase in sphingolipid levels. It also featured a significant increase in the proportion of ceramide, sphingomyelin and hexosylceramide molecules with shorter chain length and a decrease in the percentage of longer-chain sphingolipids. The extent of this shift was correlated to the degree of reduction of fibroblast glucocerebrosidase activity. Lipid extracts from control and PD-GBA fibroblasts were added to recombinant α-synuclein solutions. The kinetics of α-synuclein aggregation were significantly accelerated after addition of PD-GBA extracts as compared to control samples. Amyloid fibrils collected at the end of these incubations contained lipids, indicating α-synuclein-lipid co-assembly. Lipids extracted from α-synuclein fibrils were also analysed by shotgun lipidomics. Data revealed that the lipid content of these fibrils was significantly enriched by shorter-chain sphingolipids. In a final set of experiments, control and PD-GBA fibroblasts were incubated in the presence of the small molecule chaperone ambroxol. This treatment restored glucocerebrosidase activity and sphingolipid levels and composition of PD-GBA cells. It also reversed the pro-aggregation effect that lipid extracts from PD-GBA fibroblasts had on α-synuclein. Taken together, the findings of this study indicate that the L444P GBA mutation and consequent enzymatic loss are associated with a distinctly altered membrane lipid profile that provides a biological fingerprint of this mutation in Parkinson fibroblasts. This altered lipid profile could also be an indicator of increased risk for α-synuclein aggregate pathology.
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Project description:Duplications and triplications of the ?-synuclein (SNCA) gene increase risk for PD, suggesting increased expression levels of the gene to be associated with increased PD risk. However, past SNCA expression studies in brain tissue report inconsistent results. We examined expression of the full-length SNCA transcript (140 amino acid protein isoform), as well as total SNCA mRNA levels in 165 frontal cortex samples (101 PD, 64 control) using quantitative real-time polymerase chain reaction. Additionally, we evaluated the relationship of eight SNPs in both 5' and 3' regions of SNCA with the gene expression levels. The association between postmortem interval (PMI) and SNCA expression was different for PD and control samples: SNCA expression decreased with increasing PMI in cases, while staying relatively constant in controls. For short PMI, SNCA expression was increased in PD relative to control samples, whereas for long PMI, SNCA expression in PD was decreased relative to control samples.
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Project description:BackgroundGaucher's disease (GD) is an autosomal recessive disorder caused by a deficiency of acid β-glucosidase (glucocerebrosidase [GBA]) that results in the accumulation of glucocerebroside within macrophages. Many mutations have been reported to be associated with this disorder. This study aimed to discover more mutations and provide data for the genetic pattern of the gene, which will help the development of quick and accurate genetic diagnostic tools for this disease.MethodsGenomic DNA was obtained from peripheral blood leukocytes of the patient and Sanger sequencing is used to sequence GBA gene. Sequence alignments of mammalian β-GBA (GCase) and three-dimensional protein structure prediction of the mutation were made. A construct of this mutant and its compound heterozygous counterpart were used to measure GCase in vitro.ResultsGCase is relatively conserved at p.T219A. This novel mutation differs from its wild-type in structure. Moreover, it also causes a reduction in GCase enzyme activity.ConclusionThis novel mutation (c.655A>G, p.T219A) is a pathogenic missense mutation, which contributes to GD.
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