Figure 5.

Identification of an additional part of Cobl-like that, besides the WH2 domain, is critical for Cobl-like–mediated promotion of F-actin–rich ruffles. (A) Scheme of Cobl-like and mutants thereof used to identify the Cobl-like regions required for the formation of F-actin–rich 3D ruffles in COS-7 cells (green, inducing 3D ruffles; red, not inducing 3D ruffles; graphical summary on the right side). (B–J) Functional analyses in the context of the C-terminal half (B–G) and in full-length (H–J). (B–E and H) Representative examples of phalloidin-stained cells overexpressing the Cobl-like deletion mutants indicated. Bars, 10 µm. (F and I) Anti-Flag immunoblottings of Cobl-like and mutants thereof ensuring that all proteins are intact and that proteins negative in functional assays were expressed at least as high as the active proteins. (G and J) Quantitative analyses of ruffle formation in Cobl-like mutants-overexpressing COS-7 cells in comparison to positive and negative controls (datasets from Fig. 1). Cobl-like^1016-1104 is critical for 3-D ruffle formation (yet, not sufficient); all Cobl-like mutants lacking this sequence are not sufficient to induce ruffles and are not statistically different from mCherry and untransfected cells (controls). Data are mean ± SEM, one-way ANOVA plus Tukey’s post test (G and J). ***, P < 0.001.