Sphingolipid changes in Parkinson L444P GBA mutation fibroblasts promote alpha-synuclein aggregation

Intraneuronal accumulation of aggregated alpha-synudein is a pathological hallmark of Parkinson's disease. Therefore, mechanisms capable of promoting alpha-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 alpha-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 alpha-synuclein solutions. The kinetics of alpha-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 alpha-synuclein-lipid co-assembly. Lipids extracted from alpha-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 alpha-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 alpha-synuclein aggregate pathology.

Automated summary

This study found that mutations of the glucocerebrosidase 1 (GBA) gene, a prevalent risk factor for Parkinson's disease, led to altered lipid membrane composition in Parkinson fibroblasts. The altered lipid profile featured increased levels of sphingolipids and shorter-chain sphingolipid molecules. The extent of this alteration was correlated with the reduction of fibroblast glucocerebrosidase activity and accelerated alpha-synuclein aggregation. Treatment with a small molecule chaperone, ambroxol, restored the lipid profile and reversed the pro-aggregation effect. These findings suggest that the GBA mutation and consequent loss of enzymatic activity are associated with a distinct membrane lipid profile, which may increase the risk for alpha-synuclein aggregate pathology.