Figure 3. Effect of nutrient deprivation and temperature on translation and lifespan. (A) Translation rate of WT and eIF2α.S52A (S52A) cells measured by 35S-methionine/cysteine metabolic labelling. Coomassie staining shows the total protein levels loaded in each lane (bottom panel). Cells were grown in standard conditions (3% glucose YES medium at 30° C, Control) or low glucose (0.1% Glc) YES medium and at 20° C or 35° C prior to labelling. Cells were metabolically labelled, as previously described. (B) Longevity assay of WT and eIF2α.S52A (S52A) cells cultivated under conditions described in A). Serial dilutions (1/5) of cells were plated onto YES-Agar plates at the exponential phase (Control) or several days after reaching stationary phase of growth, as indicated, and incubated at 30° C for 2-3 days. (C) Longevity assay of cells shifted from 3% to 0.1% glucose YES medium during exponential growth phase. Left panel: eIF2α.S52A cells growing in 3% glucose (black boxes) were shifted to 0.1% glucose YES medium (white boxes) at indicated times. Right panel: eIF2α.S52A cells growing in 0.1% glucose YES medium were shifted to 3% glucose by addition of glucose at the indicated times. WT and eIF2α.S52A cells grown in 3% or 0.1% glucose YES medium during the whole experiment were used as control. Serial dilutions of cells were plated as described in panel (B). Data information: (A–C) Representative results from three independent experiments.