Figure 6.

VPS21 is required for the delivery of Vph1p but not for the delivery of ALP to the vacuole. The production of Vph1p was under the control of the galactose-inducible GAL1 promoter in both pep4-3 (NBY79) and vps21-T39K pep4-3 (SGY46) strains. Cells were grown to midlog phase at 25°C, galactose was added, and the cells were incubated for 30 min at either 25°C (A) or 37°C (B). Protein synthesis was terminated by the addition of cycloheximide, followed by incubation for an additional 45 min at 25°C (A) or for 15 min at 37°C (B) before fixation. Vph1p was labeled with rabbit anti-Vph1p antibodies and Texas Red–conjugated secondary antibody. ALP was labeled with mouse anti-ALP antibodies, followed by biotinylated secondary antibody and FITC-conjugated streptavidin.