The function and regulation of vinculin in cell-cell adhesions

Adherens junctions are essential for embryogenesis and tissue homeostasis. The major transmembrane adhesion receptors in adherens junctions are the cadherins, which mediate cell-cell adhesion by binding to cadherins on adjacent cells. Cadherin function is regulated by the protein complexes that assemble at its cytoplasmic tail. Vinculin is one cytoplasmic component of the cadherin adhesion complex, but unlike other junction components, it also is enriched in cell-matrix adhesions. The presence of vinculin in cellmatrix adhesions has commanded the most attention, while little is known about its role in cell-cell adhesions.

To define the role of vinculin in adherens junctions, I established a short hairpin RNA-based knockdown/substitution system that perturbs vinculin preferentially at sites of cell-cell adhesion. When this system was applied to epithelial cells, cell morphology was altered, and cell-cell adhesion was reduced owing to a lack of cadherin on the cell surface. I investigated the mechanism for this effect and found that vinculin must bind to beta-catenin to regulate E-cadherin surface expression.

Having established a role for vinculin in cell-cell adhesions, the critical question became how vinculin recruitment to and activation at cell-cell junctions are regulated. I found that á-catenin triggers activating vinculin conformational changes. Unlike all of the known vinculin activators in cell-matrix adhesions, alpha-catenin binds and activates vinculin independently of an A50I substitution. Thus, adherens junction activators and cell-matrix activators bind to distinct regions of vinculin to activate this molecule. Using mutant vinculins that cannot be tyrosine phosphorylated, I found that vinculin recruitment to cell-cell adhesions, but not cell-matrix adhesions, requires phosphorylation at Y822. Furthermore, this residue is phosphorylated by Abl tyrosine kinases during the assembly

of cell-cell adhesions. Taken together, these studies explain how vinculin is differentially recruited to adherens junctions and cell-matrix adhesions and describes the first known role for vinculin at cell-cell adhesions.