Fig. 3: Vacuolar pH oscillations require the SPS response, TORC1 signaling, v-ATPase activity, oscillatory production of PtdIns(3,5)P2 by the Fab1 complex and the nutrient-responsive CDK, Pho85.
From: Cell cycle-linked vacuolar pH dynamics regulate amino acid homeostasis and cell growth
a, Oscillatory vacuolar pH amplitude of wild-type cells and cells lacking the SPS components, Ptr3 (288 cells), Ssy1 (447 cells) and Ssy5 (399 cells), grown in SDC. Median vacuolar pH amplitude is displayed as box-and-whisker plots with whiskers calculated as 1.5 × the interquartile ranges, median values indicated by the horizontal red bar and P values are calculated by a two-sided Dunn’s test with Holm adjustment. Numbers inset below bar graphs are the median period of vacuolar pH oscillations. b, Cells grown in SDC imaged every 2 min for 1,000 min in microfluidics chambers (CellASIC) and treated in the device with rapamycin (vertical red bar). v-SEP intensity over time is shown for two representative cells. c, Cells grown in SDC imaged every 2 min for 1,000 min in microfluidics chambers (CellASIC) and treated in the device with ConcA (vertical red bar). v-SEP intensity over time is shown for two representative cells. Dashed lines indicate the timing of bud emergence. d, Oscillatory vacuolar pH amplitude of WT cells and cells lacking the Fab1 components, Vac7 (177 cells), Vac14 (287 cells), Atg18 (200 cells), Fab1 (21 cells) and Fig4 (206 cells), grown in SDC. Median vacuolar pH amplitude is displayed as box-and-whisker plots with whiskers calculated as 1.5 × the interquartile ranges, median values indicated by the horizontal red bar and P values are calculated by a two-sided Dunn’s test with Holm adjustment. Numbers inset below bar graphs are the median period of vacuolar pH oscillations. e, Oscillatory vacuolar pH amplitude of wild-type cells and cells lacking the CDK5 homolog Pho85 (201 cells) and associated cyclins Pho80 (240 cells), Pcl6 (315 cells), Pcl7 (277 cells) and the Pcl6 and Pcl7 double mutant (206 cells). Cells were grown in SDC. Median vacuolar pH amplitude is displayed as box-and-whisker plots with whiskers calculated as 1.5 × the interquartile ranges, median values indicated by the horizontal red bar and P values are calculated by a two-sided Dunn’s test with Holm adjustment. Numbers inset below bar graphs are the median period of vacuolar pH oscillations. f, Oscillatory vacuolar pH amplitude of WT cells and cells lacking seven putative Pho85 phospho-sites in Vac7 (vac7-7A, 228 cells) and 21 putative phospho-sites in Fab1 (fab1-21A, 168 cells) and the double mutant (vac7-7A fab1-21A, 257 cells). Cells were grown in SDC. Median vacuolar pH amplitude is displayed as box-and-whisker plots with whiskers calculated as 1.5 × the interquartile ranges, median values indicated by the horizontal red bar and P values are calculated by a two-sided Dunn’s test with Holm adjustment. Numbers inset below bar graphs are the median period of vacuolar pH oscillations. See Source Data for additional curves.
