Parkinson’s disease (PD) and dementia with Lewy bodies (DLB) belong to a family of neurodegenerative disorders called synucleinopathies, as they are caused by the pathological accumulation of the alpha-synuclein (aSyn) protein into structures called Lewy bodies (LBs) and Lewy neurites (LNs) in the brain. Focusing on an alpha-synuclein mutation that was recently identified in a patient with Lewy body dementia, scientists at Ecole Polytechnique Fédérale de Lausanne (EPFL) and colleagues carried out studies that have provided new insights into the mechanisms linking alpha-synuclein to neurodegeneration and pathology formation in PD.

Commenting on the findings from their in vitro work, research lead Hilal A. Lashuel, PhD, at EPFL’s School of Life Sciences, said, “Our findings support a central role of alpha-synuclein in the development of PD and other synucleinopathies and demonstrate that variations in the structural properties of alpha-synuclein aggregates could contribute to the neuropathological and clinical heterogeneity of synucleinopathies.”

Lashuel and colleagues reported on their findings in Science Advances, in a paper titled, “A NAC domain mutation (E83Q) unlocks the pathogenicity of human alpha-synuclein and recapitulates its pathological diversity.”

The accumulation of fibrillar and aggregated forms of alpha-synuclein within LBs and LNs is a characteristic hallmark of many synucleinopathies, including PD and DLB, the authors noted. In a healthy brain, alpha-synuclein is found in synapses as distinct monomers. But various mutations of the SNCA gene that encodes alpha-synuclein can cause the protein to clump together and form larger oligomers and even larger fibrils.

Scientists have identified and mapped out many mutations of the alpha-synuclein gene that are associated with synucleinopathies. Various studies, including research by the Lashuel lab, have shown that the mutations may act through distinct mechanisms, leading to the same pathology. Studying these rare mutations has led to important insights and helped to unmask different mechanisms that contribute to neurodegeneration and the development of Parkinson’s disease. “Although aSyn mutation carriers are rare, studies on the biochemical, cellular, aggregation, and toxic properties of these mutants have provided valuable insights into the mechanisms of aSyn aggregation and PD pathology,” the authors wrote. “These studies have also suggested that the various mutations may act via distinct mechanisms.”

A study reported in 2020 described a new mutation of the alpha-synuclein gene in a patient with LBD, associated with an atypical degeneration of the frontal and temporal lobes. The mutation substitutes the amino acid glutamate (E) with a glutamine (Q) at the 83rd position of the protein’s amino acid sequence—which is why the mutation is called E83Q. What distinguishes this mutation from all previously identified mutations is that it lies in the middle of the domain that regulates alpha-synuclein normal functions (interaction with membranes) and drives aggregation and pathology formation initiation. “Unlike all previously reported synucleinopathy-related mutations, which invariably occur in the N-terminal region spanning residues 30 to 53, and with most clustering between amino acids 46 and 53, the E83Q mutation is within the nonamyloid component (NAC) domain (residues 61 to 95), which plays a critical role in catalyzing aSyn oligomerization and fibril formation,” the team noted.

Lashuel further commented, “I was intrigued by the unique position of this mutation and the fact that the E83Q mutation carrier showed severe Lewy body pathology in the cortical and hippocampal regions of the brain than the usual substantia nigra which tends to be majorly affected in Parkinson’s disease. These observations suggested that the new mutation may influence alpha-synuclein’s structure, aggregation, and pathogenicity through mechanisms distinct from those of other mutations and could help us uncover novel mechanisms linking alpha-synuclein to neurodegeneration and pathology formation in Parkinson’s disease.”

Lashuel’s team collaborated with the groups of Markus Zweckstetter, PhD, at DZNE in Germany, and Frank Sobott at the University of Leeds. They applied a battery of biochemical, structural, and imaging approaches to dissect how this mutation modifies the structure of the different forms of alpha-synuclein and its aggregation properties in vitro. Next, they used a combination of cellular models of Lewy body formation to determine how the E83Q mutation influences various aspects of alpha-synuclein associated with its normal function and pathology.

The in vitro studies showed that the E83Q mutation not only increased dramatically the rate of alpha-synuclein aggregation but also formed aggregates with structural and morphological signatures that were distinct from those seen with the normal protein. “This was exciting since recent studies have shown that aggregates of different structures exhibit differences in their ability to induce pathology and spreading in mouse models of PD and could possibly explain the clinical heterogeneity of PD and other neurodegenerative diseases,” said Senthil T. Kumar, PhD, one of the study’s first authors.

To determine if these structural differences were sufficient to translate into differences in pathology formation and toxicity, the researchers compared the ability of E83Q and the normal alpha-synuclein protein to induce pathology formation in a neuronal model of Lew body formation and neurodegeneration that was developed in the Lashuel lab and is widely used to identify novel targets and test new alpha-synuclein targeting therapies.

“In a neuronal seeding model of LB formation, the E83Q mutation markedly increased the seeding activity of human preformed fibrils (PFFs) and promoted the formation of LB-like inclusions with diverse morphological features, resembling the diversity of aSyn pathology in PD brains,” the investigators reported. “Unlike mouse PFFs, which induce the formation of diffuse LB-like inclusions, the E83Q PFFs induced the formation of LB-like inclusions with a ring-like organization that recapitulates the immunohistochemical, structural, and morphological features of bona fide brainstem LBs observed in patient’s brains affected by late-stage PD.”

“In the neuronal seeding model of Lewy body formation, the E83Q mutation not only dramatically increased the seeding activity and the formation of Lewy body-like inclusions, but it also led to the formation of multiple aggregates with diverse morphological features— very similar to the diversity of alpha-synuclein pathology seen in the brains of patients with Parkinson’s disease,” pointed out Anne-Laure Mahul-Mellier, PhD, co-first author of the team’s study. “We were thrilled to see that we can achieve this in our Lewy-body in a dish model.”

The results, Lashuel noted, emphasized “… the critical importance of using disease models that reproduce to the extent possible the diversity of the human pathology and therapies capable of targeting the diversity of pathological alpha-synuclein species.”

As a next step, Lashuel’s group will validate these findings in animal models using material isolated from the affected patient, and will further investigate whether this mutation also influences the normal functions of alpha-synuclein.

“Together, our findings suggest that (E83Q) unlocks the pathogenicity of human aSyn fibrils and recapitulates some of its pathological diversity by promoting its oligomerization, increasing its toxicity, and inducing the formation of more pathogenic fibrillar structures,” the authors concluded. “Therefore, exploring the effects of this mutation in various models of synucleinopathies could yield novel insights into the molecular mechanisms underpinning aSyn-induced neurodegeneration and aSyn pathology formation and spreading within the brain.”