FIG 4 .
Toremifene binds the EF hand protein Cml1 and modulates its ability to activate calcineurin. (A) Serial dilutions (10-fold) of suspensions (1 OD600 unit/ml) of cells for wild-type (CM018) and cml1Δ strains were spotted on yeast-peptone-dextrose (YPD) plates containing DMSO solvent (1%) or toremifene (TOR; 8 µg/ml). The plates were incubated at 30°C for 3 days and photographed. (B) TOR (blue trace) shifts the Tm for denaturation of Cml1 compared to solvent control (DMSO, red trace). Differential scanning fluorimetry was performed as described in the legend to Fig. 2 and in Materials and Methods. Arrows indicate the Tm for the two conditions, which were determined as described in Materials and Methods. (C) TOR increases the activity of Cml1-mediated calcineurin activity. The specific activity (absorbance [abs]/mol calcineurin/min) of human calcineurin is plotted on the y axis, and the no-drug (DMSO-only) control indicates activation.