Interaction of nectin with afadin is necessary for its clustering at cell-cell contact sites but not for its cis dimerization or trans interaction

We have recently found a novel functional unit of cell-cell adhesion at cadherin-based adherens junctions, consisting of at least nectin, a hemophilic cell adhesion molecule, and afadin, an actin filament-binding protein, which connects nectin to the actin cytoskeleton. Here we studied a mechanism of cell-cell adhesion of the nectin-afadin system by use of a cadherin-deficient L cell line stably expressing the intact form of mouse nectin-2 alpha, a truncated form of nectin-2 alpha incapable of interacting with afadin (nectin-2 alpha-Delta C), or a point-mutated form of nectin-2 alpha capable of interacting with afadin and a cadherin-expressing EL cell. line, which transiently expressed the point-mutated form of nectin-2 alpha, We found that the interaction of nectin-2 alpha with afadin was necessary for their clustering at cell-cell contact sites. However, nectin-2 alpha-Delta C showed cis dimerization and trans interaction, both of which did not require the interaction of nectin-2 alpha with afadin. We have previously shown in EL cells that the interaction of nectin-1 with afadin is necessary for its recruitment to adherens junctions. We found that the trans interaction of nectin-2 alpha was furthermore necessary for this recruitment. On the basis of these observations, we propose a model for the mechanism of cell-cell adhesion of nectin and roles of afadin in this mechanism.