Fig. 5. Localization of Rgf3p but not Rgf1p or Rho1p is disrupted in lad1-1 cells. (A,B) rgf1-GFP lad1-1 cells (KGY1116) were grown at 25°C (A) and then shifted to 36°C for 4 hours (B). (C,D) rgf3-GFP lad1-1 (KGY1117) (C) and rgf3-GFP cells (KGY4407) (D) cells were grown at 25°C and then shifted to 36°C for 4 hours. Images of live cells are shown in each panel. (D) The same strains grown in parts A-D were grown at 25°C and shifted to 36°C for 0 or 2 hours and equal amounts of cell lysates were probed for Cdc2p levels using anti-PSTAIRE, or immunoprecipitated and immunoblotted with antibodies to GFP. (F) lad1-1 cells containing pREP1 (vector) or pREP1rgf3⁺ were isolated at 25°C and struck to YE plates at 36°C. Colonies were allowed to form for 3 days. (G) rgf3-GFP lad1-1 cells (KGY1117) were transformed with pREP1gyp10 and colonies were allowed to form at 25°C. Transformants were grown in liquid medium lacking thiamine for 18 hours and then shifted to 36°C for 3 hours. Live cell images were captured. The arrowhead indicates the Rgf3-GFP medial ring.