TORC1 is strongly inhibited by phosphate starvation in <i>C</i>. <i>glabrata</i>, likely contributing to the ASR via its proximal kinase, Sch9.

(A) A representative Western Blot for phosphorylated Rps6 (P-Rps6) and total Rps6 in both species under rich media, nitrogen starvation (-N) or phosphate starvation (-Pi) conditions. Red arrows point to the loss vs presence of the P-Rps6 band under -Pi in C. glabrata and S. cerevisiae, respectively. (B) Quantification of %P-Rps6 over total Rps6 (n = 3). *Bonferroni corrected P-values from Student’s t-tests were shown. (C) Inhibiting TORC1 by rapamycin induces Cta1-GFP in a dose-dependent manner. The dots, error bars and lines have the same meaning as before. (D) ASR for H2O2 with rapamycin as the primary treatment. Plotted are survival rates with or without rapamycin treatment (n = 12 for C. glabrata, n = 8 for S. cerevisiae). 60 mM and 6 mM of H2O2 were used as the secondary stress for the two species, resulting in a similar basal survival rate (P = 0.6). A Wilcoxon signed-rank test was used to compare the paired experiments with or without the primary treatment for each species. The raw, one-sided P-values were shown on the top. (E) Same plot as C, comparing Cta1-GFP induction in a phosphomimetic mutant of Sch9, a key proximal effector of TORC1, and a matching wild type Sch9 strain. (F) Same as D but with phosphate starvation (-Pi) as the primary stress, comparing the Sch9-3E mutant (n = 4) and the matching Sch9-wt control (n = 4). 100 mM and 40 mM of H2O2 were used as the secondary stress for the two genotypes, resulting in a similar basal survival rate (P = 0.7).