title: Characterization of Vta1p, a class E Vps protein in Saccharomyces cerevisiae.
abstract: We identified VTA1 in a screen for mutations that result in altered vacuole morphology. Deletion of VTA1 resulted in delayed trafficking of the lipophilic dye FM4-64 to the vacuole and altered vacuolar morphology when cells were exposed to the dye 5-(and 6)-carboxy-2',7'-dichlorofluorescein diacetate (CDCFDA). Deletion of class E vacuolar protein sorting (VPS) genes, which encode proteins that affect multivesicular body formation, also showed altered vacuolar morphology upon exposure to high concentrations of CDCFDA. These results suggest a VPS defect for Deltavta1 cells. Deletion of VTA1 did not affect growth on raffinose and only mildly affected carboxypeptidase S sorting. Turnover of the surface protein Ste3p, the a-factor receptor, was affected in Deltavta1 cells with the protein accumulating on the vacuolar membrane. Likewise the alpha-factor receptor Ste2p accumulated on the vacuolar membrane in Deltavta1 cells. We demonstrated that many class E VPS deletion strains are hyper-resistant to the cell wall disruption agent calcofluor white. Deletion of VTA1 or VPS60, another putative class E gene, resulted in calcofluor white hypersensitivity. A Vta1p-green fluorescent protein fusion protein transiently associated with a Pep12p-positive compartment. This localization was altered by deletion of many of the class E VPS genes, indicating that Vta1p binds to endosomes in a manner dependent on the assembly of the endosomal sorting complexes required for transport. Membrane-associated Vta1p co-purified with Vps60p, suggesting that Vta1p is a class E Vps protein that interacts with Vps60p on a prevacuolar compartment.
authors.affiliation: Department of Pathology, Division of Cell Biology and Immunology, University of Utah Health Science Center, Salt Lake City, Utah 84132-2501, USA.
authors.forename: Shelly L
authors.initials: SL
authors.lastname: Shiflett
authors.forename: Diane McVey
authors.initials: DM
authors.lastname: Ward
authors.forename: Dinh
authors.initials: D
authors.lastname: Huynh
authors.forename: Michael B
authors.initials: MB
authors.lastname: Vaughn
authors.forename: Jennifer C
authors.initials: JC
authors.lastname: Simmons
authors.forename: Jerry
authors.initials: J
authors.lastname: Kaplan
mesh_terms: D010641:Phenotype; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*; D013347:Subcellular Fractions / Q000378:metabolism
chemical_list: D029701:Saccharomyces cerevisiae Proteins