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Figure 4


Fig. 4. Microscopy of live chemotaxing cells expressing GFP-ABD. (A-F) Chemotaxing GFP-ABD-expressing cells developed for 8 hours in submerged conditions. (A,B) Fluorescence (top panel) and the corresponding phase-contrast (bottom panel) images at the indicated times. (C,D) Kymographs corresponding to A and B, respectively. Vertical streaks correspond to intercalated phase-contrast images taken every fifth second. The kymographs were aligned and overlaid with plots of the temporal change of centroid velocity calculated from the fluorescent images. Note that the peaks of centroid velocity coincide with bleb formation (arrowheads). A and C were imaged in SB, and B and D in high osmolarity buffer (HI). (C) Spans from -10 to 40 seconds in the sequence shown in A. (D) Spans from 15 to 65 seconds in the sequence shown in B. The arrowheads in A and C indicate a bleb in formation. The arrowheads in B and D indicate a filopodium in formation. (E,F) Higher magnifications of the leading edge of the cells presented in A and B at indicated times. Bars, 10 µm (A,B); 1 µm (C,D); 0.125 µm/second for the centroid velocity plots (C,D); 5 µm in (E,F).





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