Tks5 regulates Dyn2 assembly around actin. (A–C) Effects of Amphiphysin II and Tks5 on Dyn2-actin bundle. SH3 domain from Tks5 (PX3A) or Amphiphysin II (AmphII-SH3) were added into Dyn2 and actin filament reactions. Bundled actin was sedimented, and the ratio of bundled actin (pellet/total) was analyzed and quantified with ImageJ (B) or imaged with negative stain TEM (C). (D) Immunogold staining for Tks5 on Dyn2-actin bundle. Tks5/GST-PX3A was labeled with anti-GST antibody and subsequently with 6 nm gold particle–conjugated anti-rabbit antibody. A magnified image was shown with red arrows indicating the gold particles. Scale bars, 100 nm. (E) Distribution of endogenous Dyn2 in invadosome. Differentiated myoblasts were stained with indicated antibodies and imaged with confocal, confocal with Airyscan detector, or STED microscopy. Single focal images are shown. Scale bars, 2 µm. *, P < 0.05.