FIGURE 6:

PH^Ste5-domain mutants defective primarily in the binding of Ste11 MAPKKK. (A) Yeast cells (ste5Δ) carrying PH^Ste5-domain mutants that are defective in binding to RBL^Ste11 are defective in mating. (B) Reestablishing the interaction of Ste5 mutants in A with Ste11 through another protein–protein interaction alleviates the mating defect. Yeast cells in A were transformed with Ste5 alleles as indicated and carrying the SAM domain of Ste50 (SAM₅₀), which interacts with the SAM domain of Ste11, were assayed for the ability to mate. (C) The PH^Ste5-domain mutants have normal cellular localization. Yeast strain YCW338 (MATa sst1) carrying GFP-tagged Ste5 alleles as indicated were induced in galactose media, treated with α-factor mating pheromone (αF) (1 μM) for 1 h, and photographed using a Leitz photomicroscope equipped with a 100× objective and a MicroMax camera (bottom); GFP fluorescence photographs were acquired and processed as described in Materials and Methods.