FRAP analyses reveal a role for vesicle trafficking in the asymmetrical distribution of ACA. (A) Cartoon depicting the potential effect of intracellular vesicles on the fluorescence recovery pattern after photobleaching. (B, inset) Confocal fluorescent images of cAR1-YFP/car1/3⁻ cells (left) and ACA-YFP/aca⁻ cells (right) showing the bleached area (white) and the side (blue) and middle (red) boxes where the fluorescence recovery is monitored. The graphs depict the recovery of ACA-YFP and cAR1-YFP from differentiated nonpolar cells. The side fluorescence recovery values are shown in blue and the middle recovery values are in red. Each graph is normalized by setting initial fluorescence to 100% and the fluorescence immediately after bleaching to 0. Data are presented as a mean of five cells ± SEM. (C, top) Confocal fluorescent images of vegetative, differentiated nonpolar, polar, and migrating cAR1-YFP/car1/3⁻ (left) and ACA-YFP/aca⁻ (right) cells showing the bleached area in white and a red box where the fluorescence recovery is monitored. (bottom) Graphs depicting the fluorescence recovery of cAR1-YFP (left) and ACA-YFP (right) in the middle boxes (red) in vegetative, differentiated nonpolar, polar, and migrating cells. Data are presented as a mean of five cells ± SEM. (D, inset) Confocal fluorescent images of a ACA-YFP/aca⁻ polar cell showing the bleached area as in B. The graphs depict the recovery of ACA-YFP at the front (left) and back (right) of a polarized cell. See B for details. The front versus back graph compares the recovery from the middle box (red) at the front and the back, respectively.