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Files in this Data Supplement:
Movie 1. Membrane recruitment of single Crac-GFP molecules during chemoattractant stimulation. A Dictyostelium cell expressing Crac-GFP was observed under TIR-FM. Individual white spots are fluorescence from single Crac-GFP molecules, which become detectable only while the molecule is bound to the membrane. After the stimulation with cAMP (indicated by the interruption with a white frame) the number of Crac-GFP molecules increased transiently. Images were collected every 33 milliseconds and are displayed at the rate of 30 frames per second. The width of the image is 32 mm.
Movie 2. Membrane recruitment of single Crac-GFP molecules in a Dictyostelium cell expressing a lesser amount of Crac-GFP molecules. The low expression level allowed us to trace individual Crac-GFP molecules. The stimulation with cAMP was performed and is indicated as in Movie 1. Images were collected every 33 milliseconds and are displayed at the rate of 30 frames per second. The width of the image is 26 mm.
Movie 3. Single Crac-GFP molecules in a chemotactically migrating cell. A Dictyostelium cell expressing Crac-GFP was observed under TIR-FM during chemotactic movement. The frequent repetition of Crac-GFP molecules binding to the membrane of was observed at the leading edge, whereas a small amount of molecules was bound to the membrane in the posterior region (indicated by triangles). Images were collected every 33 milliseconds and are displayed at the rate of 30 frames per second. The width of the image is 19 mm.
Movie 4. Single Crac-GFP molecules in a chemotactically migrating cell with a lesser expression of Crac-GFP. The behaviors of Crac-GFP molecules during the association with the membrane at the leading edge was analyzed at single-molecule level. In the movie, two cells are migrating towards the lower left corner of the image, and their anterior halves are shown. Images were collected every 33 milliseconds and are displayed at the rate of 30 frames per second. The height of the image is 19 mm.
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