Figure 2.

Endocytic trafficking is defective in sec14-1^ts mutants. (A) GFP-Snc1 accumulates in intracellular compartments in sec14-1^ts mutants. Wild-type and isogenic sec14-1^ts yeast harboring YCp(GFP-SNC1) were grown to mid-logarithmic growth phase in uracil-free minimal medium at 30°C. Cells were incubated with FM4-64 (10 μM) for 10 min after which cells were intoxicated with NaN₃/NaF (10 mM final, each). Cells were washed twice with NaN₃/NaF (10 mM final, each), and FM4-64 and GFP-Snc1 profiles were imaged using a Nikon E600 microscope. GFP-Snc1, FM4-64 and merge profiles are shown, as indicated. (B) GFP-Snc1 and FYVE-dsRed localization profiles are shown. Wild-type and isogenic sec14-1^ts yeast harboring YCp(GFP-SNC1) were grown to mid-logarithmic growth phase in uracil-free minimal medium at 30°C. GFP-Snc1 and FYVE-dsRed profiles were imaged using a Nikon E600 microscope. GFP-Snc1, FYVE-dsRed, and merge profiles are shown, as indicated. Arrows highlight examples of where GFP-Snc1 and FYVE-dsRed compartments are juxtaposed. (C) GFP-Snc1 localization in bypass Sec14 mutant yeast. Yeast strains with the indicated genotypes harboring YCp(GFP-SNC1) were grown to mid-logarithmic growth phase in uracil-free minimal medium at 30°C and then either maintained at 30°C or shifted to 37°C for 60 min. The GFP-Snc1 profile was imaged using a Nikon E600 microscope.