title: The yeast vacuolar Rab GTPase Ypt7p has an activity beyond membrane recruitment of the homotypic fusion and protein sorting-Class C Vps complex.
abstract: A previous report described lipid mixing of reconstituted proteoliposomes made using lipid mixtures that mimic the composition of yeast vacuoles. This lipid mixing required SNARE {SNAP [soluble NSF (N-ethylmaleimide-sensitive factor)-attachment protein] receptor} proteins, Sec18p and Sec17p (yeast NSF and α-SNAP) and the HOPS (homotypic fusion and protein sorting)-Class C Vps (vacuole protein sorting) complex, but not the vacuolar Rab GTPase Ypt7p. The present study investigates the activity of Ypt7p in proteoliposome lipid mixing. Ypt7p is required for the lipid mixing of proteoliposomes lacking cardiolipin [1,3-bis-(sn-3'-phosphatidyl)-sn-glycerol]. Omission of other lipids with negatively charged and/or small head groups does not cause Ypt7p dependence for lipid mixing. Yeast vacuoles made from strains disrupted for CRD1 (cardiolipin synthase) fuse to the same extent as vacuoles from strains with functional CRD1. Disruption of CRD1 does not alter dependence on Rab GTPases for vacuole fusion. It has been proposed that the recruitment of the HOPS complex to membranes is the main function of Ypt7p. However, Ypt7p is still required for lipid mixing even when the concentration of HOPS complex in lipid-mixing reactions is adjusted such that cardiolipin-free proteoliposomes with or without Ypt7p bind to equal amounts of HOPS. Ypt7p therefore must stimulate membrane fusion by a mechanism that is in addition to recruitment of HOPS to the membrane. This is the first demonstration of such a stimulatory activity--that is, beyond bulk effector recruitment--for a Rab GTPase.
authors.affiliation: Department of Molecular Physiology and Biological Physics and Center for Membrane Biology, University of Virginia, Charlottesville, VA 22908, USA. chris.stroupe@virginia.edu
authors.forename: Christopher
authors.initials: C
authors.lastname: Stroupe
mesh_terms: D002308:Cardiolipins / Q000737:chemistry; D020690:GTPase-Activating Proteins; D055786:Gene Knockout Techniques; D008081:Liposomes; D008561:Membrane Fusion; D050576:Membrane Fusion Proteins / Q000737:chemistry*; D008563:Membrane Lipids / Q000737:chemistry; D008565:Membrane Proteins / Q000235:genetics; D046912:Multiprotein Complexes / Q000737:chemistry; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry*; D017855:Transferases (Other Substituted Phosphate Groups) / Q000235:genetics; D014617:Vacuoles / Q000737:chemistry / Q000201:enzymology; D033921:Vesicular Transport Proteins / Q000737:chemistry*; D020691:rab GTP-Binding Proteins / Q000737:chemistry*
chemical_list: D002308:Cardiolipins; D020690:GTPase-Activating Proteins; C416529:GYP1 protein, S cerevisiae; D008081:Liposomes; D050576:Membrane Fusion Proteins; D008563:Membrane Lipids; D008565:Membrane Proteins; D046912:Multiprotein Complexes; D029701:Saccharomyces cerevisiae Proteins; D033921:Vesicular Transport Proteins; D017855:Transferases (Other Substituted Phosphate Groups); C100091:cardiolipin synthetase; C079005:YPT7 protein, S cerevisiae; D020691:rab GTP-Binding Proteins