A direct interaction of JAM-C with the tight junction scaffold protein ZO-2

Tight junctions are sites of cell-cell contacts at the apical region of epithelial intercellular junctions that are involved in barrier formation, cellular signaling and cell-cell adhesion. Tight Junctions are formed by integral membrane proteins associated with cytoplasmic scaffolding and adapter proteins through which they are linked to the underlying actomyosin and microtubule cytoskeletons. Here, we have addressed the interaction of the Junctional Adhesion Molecule (JAM)-C with the zonula adherens (ZO) protein ZO-2. Using a combination of cell-based recruitment assays and biochemical in vitro experiments, we find that JAM-C and ZO-2 directly interact in a PDZ domain-dependent manner. Notably, the interaction requires PDZ domain 3 as well as the SH3 domain of ZO-2 indicating that ZO-2 forms a functional supramodule to interact with JAM-C. We also find that JAM-C recruits cingulin to cell-cell contacts, and that JAM-C localization at tight junctions is significantly upregulated in the absence of JAM-A. Our findings have implications on importants aspects of tight junction biology including mechanosensing and liquid-liquid phase separation.