Familial ATP10B R303W variant links autophagy-lysosome dysfunction to α-synuclein accumulation in Parkinson's disease

ATP10B gene was implicated in Parkinson's disease (PD) through lysosomal dysfunction, however, its direct association with α-synuclein (α-syn) pathology remained unclear. Here, a potential pathogenic ATP10B R303W variant was identified from 45-year-old Korean man and investigated in a familial case of early-onset PD. Genetic analyses confirmed a familial heterozygous ATP10B R303W mutation in PD within the family. Structural modeling predicted the destabilized local interactions, which altered physicochemical properties. To characterize the functional impact of the variant, ATP10B R303W (heterozygous) and SNCA A53T (positive control, homozygous) cells were generated using CRISPR/Cas9. Cells were evaluated for their mitochondrial functions, oxidative stress, autophagy, lysosomal activity, and α-syn accumulations. ATP10B R303W cells exhibited the reduced mitochondrial membrane potentials and the elevated reactive oxidative oxygen species in comparison to wild type (WT). The decreased LC3-II/I ratio indicated the impaired autophagosome formation in ATP10B R303W cells. Compromised Lysosomal function was confirmed by the increased pH, reduced membrane permeability, decreased β-glucocerebrosidase activity, and diminished Lysotracker staining. Live-cell imaging of pHrodo-labeled α-syn revealed the increased fluorescence progressively in ATP10B R303W cells. Significantly, the accumulated α-syn was noticed in the cytosolic and membrane fractions of ATP10B R303W cells, where the reduced α-syn levels were found in the supernatant. Phosphorylated α-syn displayed an increasing trend, exceeding the levels in SNCA A53T cells. Our findings provided additional evidence from a previous study on lysosomal impairment of ATP10B variants by demonstrating the first evidence of promoted pathological α-syn accumulations by ATP10B R303W. This study suggested that ATP10B R303W mutation was a potential pathogenic variant in contributing to PD pathogenesis, emphasizing the critical role in autophagy-lysosome dysfunction of α-syn pathology.