Additional file 1: Figure S1. of The NESH/Abi-3-based WAVE2 complex is functionally distinct from the Abi-1-based WAVE2 complex SekinoSaki KashiwagiYuriko KanazawaHitoshi TakadaKazuki BabaTakashi SatoSeiichi InoueHiroki KojimaMasaki TaniKatsuko 2015 (A) One hundred μg of lysates prepared from NIH 3T3 cells and mouse brain were analyzed by Western blotting with the indicated antibodies. For Abi-2, two different dilutions of the antibody were used to obtain the results. Several Abi-1 or Abi-2 bands observed for the mouse brain lysates may reflect splicing variants, or the phosphorylation or partial degradation of these proteins. (B) Control NIH3T3 cells and NIH3T3 cells expressing Abi-1 or NESH were plated onto FN-coated coverslips, and analyzed as described in Fig. 1a. (C) Quantitative analysis of the cells in (B). Cells with lamellipodial structures were counted under a fluorescence microscope. At least 100 cells were analyzed for each sample. Data represent the means ± S.D. for three independent experiments. (D) The wound-healing assay was performed using the control NIH3T3 cells and NIH3T3 cells expressing Abi-1 or NESH, and analyzed as described in Fig. 1e. (E) Mouse brain lysates were immunoprecipitated with a mouse IgG, the anti-Abi-1 monoclonal antibody (1G9), or the anti-NESH monoclonal antibody (2H7). The precipitated proteins were analyzed by Western blotting with the indicated antibodies. (F) Rac activity assay. Cell lysates of the control and FLAG-NESH expressing NIH3T3 cells were incubated with GST-CRIB immobilized on glutathione beads, and then the precipitated proteins were analyzed by Western blotting with an anti-Rac antibody. To determine the total amount of Rac, 4 % of each lysate was analyzed (Input). (TIFF 19164 kb)