Tiam1 accumulates at adhesions with talin. (A) Migrating U251 cells were stained with talin (green) and Tiam1 (red). The images were taken under TIRFM. Insets (a, front region; b, rest) in the left panel are magnified in the right panels. Bars, 10 µm. (B) Size distribution of adhesions and quantification of Tiam1/talin colocalization in migrating U251 cells. Each migrating cell is divided into two regions as indicated diagramatically on the left: front region (1) and rest (2). Small and large adhesions were defined by the distribution of adhesion sizes (histogram, middle). (C) U251 cells transfected with siRNAs indicated in the leftmost images were stained with talin and Tiam1 antibodies. Insets in the leftmost images (merged) are magnified in the panels to the right. Bars, 10 µm. Bar graph on right represents the number of talin- or Tiam1-containing adhesion per 100 µm². Data represent the means ± SD of five independent experiments. n > 50. *, P < 0.05 versus respective control (Tukey’s HSD). Results are representative of more than three experiments.