Four subtypes of disease-causing missense mutations underlie pathogenic protein interactions in neurodegenerative VPS13A disease

VPS13A is an intracellular lipid transfer protein comprising over 3,000 amino acids. Mutations in human VPS13A cause VPS13A disease, a neurodegenerative disorder that affects movement and cognition. VPS13A forms a complex with the membrane protein XK to mediate ATP-induced phospholipid scrambling in the plasma membrane. Here, we established a mouse cell system expressing full-length mouse VPS13A and examined its interaction with XK. Mutational analysis revealed that VPS13A binds to XK through a C-terminal β-strand that interacts with a β-hairpin in the central region of XK, an interaction essential for scramblase activity. The XK paralog XKR2, which contains a similar β-hairpin structure, also associates with VPS13A and supports phospholipid scrambling. We analyzed ten mouse VPS13A variants corresponding to patient mutations and classified them into four groups: (1) L67P, I90K, and W2453R, which showed reduced expression; (2) A1091P and M3080R, which were normally expressed but lacked scramblase activity; (3) S1446P, Q2689H, Y2713C, and R3084H, which modestly impaired expression or activity; and (4) I2763R, which altered cell size, and disrupted ER independently of XK. These findings define the VPS13A-XK interaction interface, clarify the functional impact of disease-causing mutations, and reveal an unexpected gain-of-function mutation of a VPS13A variant.