title: The HOPS/class C Vps complex tethers membranes by binding to one Rab GTPase in each apposed membrane.
abstract: Many Rab GTPase effectors are membrane-tethering factors, that is, they physically link two apposed membranes before intracellular membrane fusion. In this study, we investigate the distinct binding factors needed on apposed membranes for Rab effector-dependent tethering. We show that the homotypic fusion and protein-sorting/class C vacuole protein-sorting (HOPS/class C Vps) complex can tether low-curvature membranes, that is, liposomes with a diameter of ∼100 nm, only when the yeast vacuolar Rab GTPase Ypt7p is present in both tethered membranes. When HOPS is phosphorylated by the vacuolar casein kinase I, Yck3p, tethering only takes place when GTP-bound Ypt7p is present in both tethered membranes. When HOPS is not phosphorylated, however, its tethering activity shows little specificity for the nucleotide-binding state of Ypt7p. These results suggest a model for HOPS-mediated tethering in which HOPS tethers membranes by binding to Ypt7p in each of the two tethered membranes. Moreover, because vacuole-associated HOPS is presumably phosphorylated by Yck3p, our results suggest that nucleotide exchange of Ypt7p on multivesicular bodies (MVBs)/late endosomes must take place before HOPS can mediate tethering at vacuoles.
authors.affiliation: Department of Molecular Physiology and Biological Physics and Center for Membrane Biology, University of Virginia School of Medicine, Charlottesville, VA 22908.
authors.forename: Ruoya
authors.initials: R
authors.lastname: Ho
authors.affiliation: Department of Molecular Physiology and Biological Physics and Center for Membrane Biology, University of Virginia School of Medicine, Charlottesville, VA 22908 chris.stroupe@virginia.edu.
authors.forename: Christopher
authors.initials: C
authors.lastname: Stroupe
mesh_terms: D033942:Adaptor Proteins, Vesicular Transport / Q000378:metabolism; D047389:Casein Kinase I / Q000378:metabolism; D058546:Class III Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D005656:Fungal Proteins / Q000378:metabolism; D007425:Intracellular Membranes / Q000378:metabolism; D008561:Membrane Fusion / Q000502:physiology*; D057146:Multivesicular Bodies / Q000378:metabolism; D010766:Phosphorylation; D011485:Protein Binding; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*; D014617:Vacuoles / Q000378:metabolism*; D033921:Vesicular Transport Proteins / Q000378:metabolism; D020691:rab GTP-Binding Proteins / Q000378:metabolism*
chemical_list: D033942:Adaptor Proteins, Vesicular Transport; D005656:Fungal Proteins; D029701:Saccharomyces cerevisiae Proteins; C406817:VAM6 protein, S cerevisiae; C065454:VPS33 protein, S cerevisiae; C083878:VPS41 protein, S cerevisiae; D033921:Vesicular Transport Proteins; D058546:Class III Phosphatidylinositol 3-Kinases; D047389:Casein Kinase I; C479667:YCK3 protein, S cerevisiae; C079005:YPT7 protein, S cerevisiae; D020691:rab GTP-Binding Proteins