Figure 7. Ub-Fur4-GFP is correctly sorted at the MVB in ubp2Δ mutant cells.

pFUR4-GFP or pUb-FUR4-GFP expression plasmids were transformed into WT and ubp2Δ cells. Strains were grown in sucrose, diluted (OD₆₀₀ = 0.5) in media containing galactose and Fur4 synthesis induced for 4 hours. Transcription was stopped by adding glucose for 1 hour to chase Fur4 fusion reporters to the plasma membrane. Uracil (40 µg/ml) was then added to trigger internalization. GFP signal was viewed by fluorescence confocal microscopy both before (T0) and 60 min after uracil addition. (Note: cells in the left panel were also transformed with a control vector (YEp46Δ) and incubated with 0.1 mM copper sulphate, as the experiment was done in parallel to that shown in Figure 5.)