Gap1 is recruited to sites of Ras1 activity. (A) Gap1-GFP during vegetative growth in WT and mutant strains. (B) Colocalization of Scd2-mCherry (red) and Gap1-GFP (green) during early mating in h90 WT cells. Right: Kymographs of the cell periphery. Arrowheads highlight dynamic zones of colocalization. (C) Colocalization of Scd2-mCherry and Gap1-GFP in a dynamic polarity patch of h-sxa2Δ cells treated with 0.01 µg/ml P-factor. Red arrowhead highlights one Scd2-mCherry patch that appears and disappears in the course of the time lapse. For more examples, see Fig. S4. (D) Cortical profiles of Scd2-mCherry and Gap1-GFP normalized fluorescence intensity at the exploratory patch, showing that Gap1-GFP signal persists after Scd2-mCherry signal has disappeared. n = 10 patches from 10 different cells. Thick line, mean; shaded area, SD. Two-sample^{t test} was calculated for each time point and resulted in *, P ≤ 1.3 × 10⁻² between 224 and 365 s. (E) Cortical profiles of Gap1-GFP fluorescence in h90 WT strains during early (exploration) or late (fusion site) mating; n = 20. Thick line, mean; shaded area, SD. (F) Normalized value of Gap1-GFP cortical fluorescence over time at the fusion site in h90 WT cells. Fluorescence profiles were aligned to the fusion time (t = 0) and normalized to maximal value; n = 22. Error bars, SD. (G and H) Myo52-tdTomato (red) and Gap1-GFP (green; G), and RasAct^mCherry (red) and Gap1-GFP (green; H) in h90 WT strains during fusion. Gap1-GFP signal is broader than Myo52-tdTomato or RasAct^mCherry (arrowheads). (I) Cortical profiles of Gap1-GFP, RasAct^GFP, and Ste6-sfGFP at the fusion site in h90 WT strains as in G and Fig. 3 (B and F). The profiles were aligned to the Myo52-tdTomato signal coimaged in the same cells (dashed line); n = 20. Bars, 2 µm.