Cdc42p focus formation is dependent on both Fus1p and Fus2p. (A and B) Deletion of FUS1 or FUS2 cause defects in Cdc42p focus formation. (A) Representative images of prezygotes. WT (MY15747), fus1Δ (MY15752), fus2Δ (MY15717), and fus1Δfus2Δ (MY15748) strains all containing GFP–CDC42 were mated to a fus1Δfus2Δ strain (JY429) for 2.5 h at 30°, fixed with 2% formaldehyde, and imaged. (B) Quantification of Cdc42p ZCF localization for all prezygotes imaged. Fluorescence intensities across the ZCF were measured for all prezygotes, interpolated to account for differences in ZCF length, and plotted as a ratio of the center values to the mean of the two edges. n ≥ 75 prezygotes per strain. (C and D) Cdc42p localization in shmoos is independent of Fus1p and Fus2p. (C) Representative images of shmoos. The same strains as in A were imaged after treatment with pheromone for 1.5 h. (D) Quantification of the fluorescence across the shmoo tip for all shmoos imaged. Fluorescence intensities were measured across the shmoo tip for all shmoos, interpolated, and plotted as a ratio of the center values to the mean of the two edges. n ≥ 37 shmoos per strain. (E and F) Cdc42p138 does not form a ZCF focus. (E) Representative images of GFP–Cdc42p138 prezygotes. GFP–cdc42–138 (MY15789) was mated to a fus1Δfus2Δ strain (JY429) for 2.5 h at 30°, fixed with 2% formaldehyde, and imaged. (F) Quantification of Cdc42p138 ZCF localization in prezygotes. n ≥ 84 prezygotes per strain. (G and H) Fus2p localization to the ZCF is not strongly altered in cdc42–138. (G) Representative images of Fus2p–GFP in WT (MY15588) and Cdc42p138 (MY15590) cells, both mated to fus1Δfus2Δ (JY429). (H) Quantification of Fus2p across the ZCF in prezygotes. Fluorescence intensities across the ZCF were measured for all prezygotes, interpolated to account for differences in ZCF length, and plotted as a ratio of the center values to the mean of the two edges. n ≥ 22 for each strain. Bars, 2 µm.
