Uechi, H.; Sun, D.; Saeki, Y.; Hiraiwa, T.; Honigmann, A.; Hyman, A. A.; Kuranaga, E.

Epithelial morphogenesis relies on coordinated multicellular movements regulated by adhesion and force-generating molecules. It remains elusive how cells ensure organization of these molecules in space and time. Combining in vitro reconstitution and fly genetics, we demonstrate that condensation of an adhesive transmembrane protein shapes molecular dynamics at tricellular junctions to facilitate cell-cell junction remodeling. Specifically, we identify that the intracellular domain of Sidekick, known to localize to tricellular junctions and recruit mechanical force-generating myosin II, undergoes phase separation into condensates. This condensation controls the physiological localization and concentration of Sidekick on lipid membranes. Altering the molecular dynamics of these condensates disrupts Sidekick's localization against cellular movements, misdirecting myosin II and leading to delayed junction formation. These findings suggest that cells exploit the mechano-resistant properties of tricellular junction-anchored condensates to spatially allocate force-regulating molecules. This study underscores the importance of condensation to organize tricellular junctions for robust multicellular movement.