FIGURE 7.

The clo-3-0 (S558Y,N242K) suppressor reverses the drug-sensitive phenotype of S558Y. A, quantitative analysis of clo-3-0 growth in the presence of trityl was carried out as described under “Experimental Procedures.” Growth of WT (▪), S558Y (•), clo-1-0 (○), and clo-3-0 (▴) in the presence of increasing concentrations of tritylimidazole was performed. Growth was estimated on the basis of absorbance at 600 nm and plotted as percentage growth, where the absorbance in the absence of tritylimidazole was taken to be 100%. The mean values (±S.D.) of three independent experiments are plotted. B, R6G (2.5 μm) accumulation in yeast strains bearing WT and the mutants S558Y and S558Y,N242K (clo-3-0 strain) was analyzed with flow cytometry (21). Representative histograms from four independent experiments are shown. The strains used are labeled on the figure. C, clo3–0 phenotype was recreated by inserting pS558Y,N242K plasmid DNA into R-1 (pdr5::KANMX4) by lithium chloride transformation. Quantitative growth in liquid medium containing trityl was carried out as described in A with WT (R-1 + pSS607; •), S558Y (R-1 + pS558Y; ▪), transformants of clo-3-0 (S558Y,N242K; ▴), and R-1 + pS558Y,N242K; ▾). The mean values (±S.D.) of three independent experiments are plotted.