A Structural Model for Vinculin Insertion into PIP-Containing Membranes and the Effect of Insertion on Vinculin Activation and Localization.

Vinculin, a scaffolding protein that localizes to focal adhesions (FAs) and adherens junctions, links the actin cytoskeleton to the adhesive super-structure. While vinculin binds to a number of cytoskeletal proteins, it can also associate with phosphatidylinositol 4,5-bisphosphate (PIP) to drive membrane association. To generate a structural model for PIP-dependent interaction of vinculin with the lipid bilayer, we conducted lipid-association, nuclear magnetic resonance, and computational modeling experiments. We find that two basic patches on the vinculin tail drive membrane association: the basic collar specifically recognizes PIP, while the basic ladder drives association with the lipid bilayer. Vinculin mutants with defects in PIP-dependent liposome association were then expressed in vinculin knockout murine embryonic fibroblasts. Results from these analyses indicate that PIP binding is not required for localization of vinculin to FAs or FA strengthening, but is required for vinculin activation and turnover at FAs to promote its association with the force transduction FA nanodomain.