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. 2013 Jul 8;202(1):35–44. doi: 10.1083/jcb.201301039

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Figure 4. — Stresses that induce domain formation in vacuolar membranes. (A) Growing cultures of cells expressing Vph1-GFP were washed and resuspended in SC, H₂O, SC lacking nitrogen (SD-N), or SC lacking glucose (SC-glc), and the indicated compounds were added (Rapa, rapamycin; 2DG, 2-deoxyglucose; CHX, cycloheximide). The percentage of cells with vacuolar domains was measured after 3 h. (B–D) The cultures in A were grown to Stat-phase (18 h), and the percentage of cells with vacuolar domains (B), the final OD_600nm (C), and pH (D) of the cultures were determined (n = 100–300 cells from at least two independent experiments; means ± SD). (E) pH-dependent dissipation of vacuolar domains; cells expressing Vph1-GFP were grown to Stat-phase, and the percentage of cells with vacuolar domains was determined. A portion of the cultures was pelleted, and the medium was replaced with 100 mM Tris, pH 9.3. After 75 min of incubation at 30°C, the half of the culture in Tris was pelleted and resuspended in used SC medium. Means ± SD; two independent experiments.