ZO/afadin KD does not fragment monolayers or prevent tissue contractility. (A–D) 3D rendering of the whole tissue section (A–C, 10 µm; D, 15 µm) or cross sections (E–G) of control, ZO KD, afadin single KD, or ZO/afadin KD monolayers. Glycoprotein 135 (gp135) marks the apical surface. Control (A and E) and afadin single KD (C) monolayers have flat surfaces and uniform cell height (E, double arrow). ZO KD cells (B and F) are domed and slightly taller (F, double arrow) and have ZA actomyosin (arrowheads). ZO/afadin KD cells (D and G) vary from very tall and domed (arrowheads and right double arrow) to very short and not columnar (arrows and right double arrow). (H) Laser cutting of long (>6 µm) or short (<6 µm) borders. Error bars are SDs of >30 contacts (n = 3). ns, not significant. (I, left) Schematic of monolayer detachment by dispase. (right) DMSO (control) or 0.1 mM blebbistatin treatment. Control monolayer remains flat, whereas ZO KD and ZO/afadin KD monolayers are constricted. Blebbistatin relaxed constriction. (J and K) Higher magnification image of I.