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.