(A) Two-hybrid assay of the interaction between Kin2-C1 and Rho3. The assay was performed as described in Materials and Methods. pGBDU-C1 (DBD, vector), pGBDU-RHO3Q74L, ΔC, and pGBDU-RHO3T30N, ΔC were used to pair with pOAD (AD, vector) and pOAD-KIN2-C1. Cells were grown on SC-Leu-Ura-Ade plate at 30°C for 3 days. (B) BiFC assay between Kin2-C9 and Rho3. Cells of strain JGY3088 (kin1Δ kin2Δ) carrying pVN1-KIN2-C9/pVC1, pVN1-KIN2-C9/pVC1-PMT-RHO3Q74L, and pVN1/ pVC1-PMT-RHO3Q74L pairs were grown on SC-His-Ura plate at 30°C for 16 hr. Green fluorescence was examined. Bar, 5 μm. (C) Two-hybrid assay of the interaction between Kin2 segments and Bmh1. pGBDU-C1 (DBD, vector) and pGBDU-KIN2 segments were used to pair with pOAD (AD, vector) and pOAD-BMH15-267. Cells were grown on SC-Leu-Ura-His containing 3 mM 3-AT (-His+3-AT) and SC-Leu-Ura-Ade (-Ade) plates at 30°C for 3 days.