Fig. 1. Translocation of Crac-GFP to membranes in D. discoideum cells after stimulation with cAMP. Images in A and C show one cell. (A) D. discoideum cells were stimulated at t=0 seconds with a uniform concentration of cAMP. Standard epifluorescence images were taken at 30 images per second. Before stimulation with cAMP Crac-GFP was distributed in the cytosol. After stimulation it translocated uniformly and almost entirely to the cell surface. (B) Time course of the Crac-GFP recruitment shown in A. Fluorescence intensities of Crac-GFP on membranes were measured and plotted over time. Fluorescence intensities were calibrated by using the photobleaching rates of GFP. (C) Localization of Crac-GFP at the leading-edge pseudopod of cells in gradients of cAMP.