FIGURE 7: Activation of the Slt2 pathway is determined through phosphorylation in acetic acid-treated cells. (A) Protein extracts prepared from the indicated strains untreated or treated with acetic acid (40 mM) for 12'. Upper panel, immunoblots of wild-type (H3 WT) and mutant extracts with anti-Slt2 antibodies (control). Bottom panel, immunoblots of H3 WT and mutant extracts with antiphospho-p44/42 Slt2 antibodies. (B) Same as in A but with a treatment duration of 180'. (C) H3 WT and the mutants were spotted in YPD only (untreated) or different concentrations of Congo red (CR) (60, 100 μg/mL) or (D) Caffeine (Caf) (10, 15 mM). The spotted cells were grown at 30°C for 48h (CR) or 72h (Caf) and scanned.