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Fig. 8. AJ disassembly induces a Rap1 activity-dependent assembly of focal adhesions. FRT cells transiently transfected with EGFP-zyxin, as a marker for focal adhesions, were plated on FN-coated coverslips and grown to confluence for 12 hours. Living cells were then analyzed by time-lapse microscopy during disruption of cell-cell junctions by EGTA treatment in serum-free medium. (A) The two selected frames of the video sequence show the subcellular distribution of EGFP-zyxin before cell treatments (Ctrl), and after 30 minutes of EGTA treatment (EGTA). Notice the assembly of EGFP-zyxin-containing focal adhesions upon EGTA treatment. (B) Cells expressing EGFP-zyxin were microinjected with GST-Rap1GAP (left cell) or with GST alone (right cell) 30 minutes before EGTA treatment. The two selected frames of the video sequence show the subcellular distribution of EGFP-zyxin in microinjected cells before (Ctrl) and after 30 minutes of EGTA treatment (EGTA). Notice that GST-Rap1GAP injection clearly affected the formation of focal adhesions upon disruption of cell-cell junctions. Phase contrast images of the treated cells are also shown. QuickTime movies are available in the supplementary material.





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