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AMBRA1, a novel α-synuclein-binding protein, is implicated in the pathogenesis of multiple system atrophy.

ABSTRACT: The accumulation of abnormal α-synuclein is the major histopathological feature of Lewy body disease and multiple system atrophy (MSA), which are referred to as synucleinopathies. Cytoplasmic degradation systems, such as the autophagy-lysosome and proteasome pathways, are involved in their pathogenesis. Autophagy is tightly regulated by several upstream proteins including UNC-51-like kinase 1/2, beclin1, vacuolar protein sorting-associated protein 34 and autophagy/beclin1 regulator 1 (AMBRA1). Recently, we revealed that both cortical and brainstem-type Lewy bodies were immunopositive for several upstream proteins of autophagy. Therefore, we conducted the present study to elucidate the role of upstream proteins of autophagy in the pathogenesis of MSA. Pathological and biochemical analyses using human brain samples revealed that AMBRA1 is a component of the pathological hallmarks of MSA and upstream proteins of autophagy are impaired in the MSA brain. In vitro and in vivo analyses revealed a ninefold stronger affinity of AMBRA1 with α-synuclein phosphorylated at serine 129 compared with non-phosphorylated α-synuclein. Furthermore, a weak but significant correlation between AMBRA1 overexpression and reduction of abnormal α-synuclein was observed. Silencing AMBRA1 function caused aggregates of α-synuclein in the cytoplasm of mouse primary cultured neurons, which was simulated by the treatment of Bafilomycin, an autophagy inhibitor. Our results demonstrated for the first time that AMBRA1 is a novel hub binding protein of α-synuclein and plays a central role in the pathogenesis of MSA through the degradative dynamics of α-synuclein. These results raise the possibility that molecular modulation targeting AMBRA1 can be a promising candidate for the treatment of synucleinopathies.

SUBMITTER: Miki Y

PROVIDER: S-EPMC8028368 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Synucleinopathies are neurodegenerative diseases that are associated with the misfolding and aggregation of α-synuclein, including Parkinson's disease, dementia with Lewy bodies and multiple system atrophy1. Clinically, it is challenging to differentiate Parkinson's disease and multiple system atrophy, especially at the early stages of disease2. Aggregates of α-synuclein in distinct synucleinopathies have been proposed to represent different conformational strains of α-synuclein that can self-propagate and spread from cell to cell3-6. Protein misfolding cyclic amplification (PMCA) is a technique that has previously been used to detect α-synuclein aggregates in samples of cerebrospinal fluid with high sensitivity and specificity7,8. Here we show that the α-synuclein-PMCA assay can discriminate between samples of cerebrospinal fluid from patients diagnosed with Parkinson's disease and samples from patients with multiple system atrophy, with an overall sensitivity of 95.4%. We used a combination of biochemical, biophysical and biological methods to analyse the product of α-synuclein-PMCA, and found that the characteristics of the α-synuclein aggregates in the cerebrospinal fluid could be used to readily distinguish between Parkinson's disease and multiple system atrophy. We also found that the properties of aggregates that were amplified from the cerebrospinal fluid were similar to those of aggregates that were amplified from the brain. These findings suggest that α-synuclein aggregates that are associated with Parkinson's disease and multiple system atrophy correspond to different conformational strains of α-synuclein, which can be amplified and detected by α-synuclein-PMCA. Our results may help to improve our understanding of the mechanism of α-synuclein misfolding and the structures of the aggregates that are implicated in different synucleinopathies, and may also enable the development of a biochemical assay to discriminate between Parkinson's disease and multiple system atrophy.

Project description:Multiple system atrophy (MSA) is characterized by the presence of distinctive glial cytoplasmic inclusions (GCIs) within oligodendrocytes that contain the neuronal protein alpha-synuclein (aSyn) and the oligodendroglia-specific phosphoprotein TPPP/p25α. However, the role of oligodendroglial aSyn and p25α in the formation of aSyn-rich GCIs remains unclear. To address this conundrum, we have applied human aSyn (haSyn) pre-formed fibrils (PFFs) to rat wild-type (WT)-, haSyn-, or p25α-overexpressing oligodendroglial cells and to primary differentiated oligodendrocytes derived from WT, knockout (KO)-aSyn, and PLP-haSyn-transgenic mice. HaSyn PFFs are readily taken up by oligodendroglial cells and can recruit minute amounts of endogenous aSyn into the formation of insoluble, highly aggregated, pathological assemblies. The overexpression of haSyn or p25α accelerates the recruitment of endogenous protein and the generation of such aberrant species. In haSyn PFF-treated primary oligodendrocytes, the microtubule and myelin networks are disrupted, thus recapitulating a pathological hallmark of MSA, in a manner totally dependent upon the seeding of endogenous aSyn. Furthermore, using oligodendroglial and primary cortical cultures, we demonstrated that pathology-related S129 aSyn phosphorylation depends on aSyn and p25α protein load and may involve different aSyn "strains" present in oligodendroglial and neuronal synucleinopathies. Importantly, this hypothesis was further supported by data obtained from human post-mortem brain material derived from patients with MSA and dementia with Lewy bodies. Finally, delivery of haSyn PFFs into the mouse brain led to the formation of aberrant aSyn forms, including the endogenous protein, within oligodendroglia and evoked myelin decompaction in WT mice, but not in KO-aSyn mice. This line of research highlights the role of endogenous aSyn and p25α in the formation of pathological aSyn assemblies in oligodendrocytes and provides in vivo evidence of the contribution of oligodendroglial aSyn in the establishment of aSyn pathology in MSA.

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AMBRA1, a novel α-synuclein-binding protein, is implicated in the pathogenesis of multiple system atrophy.

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