FIGURE 3.

CD2AP and Rac1 complex with actin-regulating proteins and co-localize in membrane ruffles. A, HeLa cells were either transfected with control siRNA or siRNA for CD2AP, followed by the isolation of Rac1 C terminus binding proteins. Bound proteins were separated by SDS-PAGE and blotted for CD2AP, cortactin, CAPZ, and c-Cbl. B and C, HeLa cells were seeded on fibronectin-coated glass coverslides and either transfected with CD2AP-GFP or co-transfected with either Myc-Rac1-Q61L or Myc-Rac1-T17N (C). D, untransfected HeLa cells were immunostained for endogenous CD2AP and endogenous Rac1. E, HeLa cells were co-transfected with CD2AP-GFP and Rac1-Q61L, and stained for paxillin to visualize peripheral adhesion structures. F, HeLa cells were transfected with CD2AP-GFP, N-CD2AP-GFP, or C-CD2AP-GFP and immunostained for the focal adhesion marker paxillin. Scale bars, 10 μm. WB, Western blot.