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Halo Assays and Doubling Time Measurements. For halo assays, a paper disk was placed on a plate spread with » 10⁶ YRP1 (snq2D pdr5D erg6D ) cells. We spotted 10 m l of a 10 mM stock of GW400426 in DMSO on the disk and the plate was placed at 30° for 48 h. For doubling time measurements, YRP1 cells growing in log phase were diluted to an OD₆₀₀ of 0.1 and treated with DMSO or various concentrations of GW400426. Aliquots of yeast were harvested every hour, fixed in 3.7% formaldehyde, and counted by using a Coulter counter. Doubling times were calculated by conducting linear regression with EXCEL (Microsoft).

Fig. 5. GW400426 elicits cellular responses in drug-sensitized but not WT yeast. (a) Scatter plots of gene expression ratios from WT (Left) or YRP1 (Right) cells treated with 20 m M GW400426. (b) Filter discs spotted with 10 m l of a 10 mM solution of GW400426 were applied to lawns of WT (Left) or YRP1 (Right) cells. (c) Doubling times of YRP1 cells treated with 10-fold dilutions of GW400426 ranging from 5 nM to 50 m M. The EC₅₀ value was » 20 m M. The orange dashed line represents doubling time measured after treatment with the equivalent amount of DMSO.

Fig. 6. (a) Scatter plot of expression ratios (log₂) from YRP1 cells treated with 20 m M GW400426 (x axis) and WT cells treated with 5 m M 1-NA-PP1 (y axis). All treatments were for 10 min. Blue dots correspond to genes in Fig. 2a (cluster C). Red dots correspond to genes in Fig. 2a (cluster B). Green dots correspond to genes in Fig. 3c. Correlation coefficient is noted in upper left hand corner. (b) Scatter plot of YRP1 cells treated with 20 m M GW400426 (x axis) and Pho85-as1 cells treated with 5 m M 1-NA-PP1 (y axis). (c) Scatter plot of YRP1 cells treated with 20 m M GW400426 (x axis) and Cdk1-as1 cells treated with 5 m M 1-NA-PP1 (y axis). (d) Scatter plot of YRP1 cells treated with 20 m M GW400426 (x axis) and Cdk1-as1/Pho85-as1 cells treated with 5 m M 1-NA-PP1 (y axis).