RhoA and cofilin localization and dynamics. (A) RhoA/ZO-1 double staining in MDCK II cells. (B) RhoA/ZO-1 double staining in claudin/JAM-A KO cells. Apical junction localization of RhoA was increased. (C and D) Magnified views of RhoA/ZO-1 double staining in claudin/JAM-A KO cells. RhoA was often localized to the edge of broken junctions (yellow arrows), although RhoA did not localize to some junction breakage sites (yellow arrowheads). (E) Cofilin/ZO-1 double staining in MDCK II cells. (F) Cofilin/ZO-1 double staining in claudin/JAM-A KO cells. Cofilin concentration along the cell junctions was increased. (A–F) are z-stacked images of the apical confocal sections. (G) Rho activation during cytokinesis in MDCK II cells revealed by tdTomato-2×rGBD probe. Rho was activated at the cleavage furrow (white arrow). (H) Rho activation during cytokinesis in claudin/JAM-A KO cells revealed by tdTomato-2×rGBD probe. Rho was activated at the cleavage furrow (white arrow). (I) Transient focal activation of Rho in MDCK II cells around the remodeling junctions (white arrowhead). This junction underwent a T1 transition, followed by focal activation of RhoA adjacent to the newly forming junction. (J) Transient focal activation of Rho was associated with the junction repair in claudin/JAM-A KO cells (yellow arrows). (K) An example of junction repair with prominent Rho activation at the edge of a broken junction in claudin/JAM-A KO cells (yellow arrows). Scale bars: 10 μm in A–H; 2 μm in I–K.