Abstract
Yeast VPS13 is the founding member of a eukaryotic gene family of growing interest in cell biology and medicine. Mutations in three of four human VPS13 genes cause autosomal recessive neurodegenerative or neurodevelopmental disease, making yeast Vps13p an important structural and functional model. Using cell-free reconstitution with purified Vps13p, we show that Vps13p is directly required both for transport from the trans-Golgi network (TGN) to the late endosome/prevacuolar compartment (PVC) and for TGN homotypic fusion. Vps13p must be in complex with the small calcium-binding protein Cdc31p to be active. Single-particle electron microscopic analysis of negatively stained Vps13p indicates that this 358-kD protein is folded into a compact rod-shaped density (20 x 4 nm) with a loop structure at one end with a circular opening ~6 nm in diameter. Vps13p exhibits ATP-stimulated binding to yeast membranes and specific interactions with phosphatidic acid and phosphorylated forms of phosphatidyl inositol at least in part through the binding affinities of conserved N-and C-terminal domains.
Original language | English (US) |
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Pages (from-to) | 425-439 |
Number of pages | 15 |
Journal | Journal of Cell Biology |
Volume | 216 |
Issue number | 2 |
DOIs | |
State | Published - 2017 |
Funding
The authors thank John Kilmartin for providing the ZCdc31 plasmid and for helpful discussions, Mark Rose for providing cdc31 mutant strains and anti-Cdc31p antibody, and Akira Ono and Balaji Olety for helpful suggestions on liposome preparation. This work was supported by the Elyse (Lakritz) Weinbaum and Gaven Lakritz Research Fund, the Protein Folding Disease Initiative of the University of Michigan Medical School, the Advocacy for Chorea Acanthocytosis Patients, and National Institutes of Health grants RO1 GM50915 (to R.S. Fuller) and R01 DK090165 (to G. Skiniotis), Medical Scientist Training Program grant NIH GM0786, and Genetics Training Program grant NIH GM07544 (to M.E. Abazeed). The authors declare no competing financial interests.
ASJC Scopus subject areas
- Cell Biology