%0 Figure %A Sekino, Saki %A Kashiwagi, Yuriko %A Kanazawa, Hitoshi %A Takada, Kazuki %A Baba, Takashi %A Sato, Seiichi %A Inoue, Hiroki %A Kojima, Masaki %A Tani, Katsuko %D 2015 %T Additional file 1: Figure S1. of The NESH/Abi-3-based WAVE2 complex is functionally distinct from the Abi-1-based WAVE2 complex %U https://springernature.figshare.com/articles/figure/Additional_file_1_Figure_S1_of_The_NESH_Abi-3-based_WAVE2_complex_is_functionally_distinct_from_the_Abi-1-based_WAVE2_complex/4465646 %R 10.6084/m9.figshare.c.3643466_D1.v1 %2 https://springernature.figshare.com/ndownloader/files/7195070 %K c-Abl %K Abi-1 %K NESH/Abi-3 %K WAVE2 %K Imatinib mesylate %K Membrane protrusions %X (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) %I figshare