Vinculin acts as a sensor in lipid regulation of adhesion-site turnover

Abstract

The dynamics of cell adhesion sites control cell morphology and motility. Adhesion-site turnover is thought to depend on the local availability of the acidic phospholipid phosphatidylinositol-4,5-bisphosphate (PIP2). PIP2 can bind to many cell adhesion proteins such as vinculin and talin, but the consequences of this interaction are poorly understood. To study the significance of phospholipid binding to vinculin for adhesion-site turnover and cell motility, we constructed a mutant, vinculin-LD, deficient in acidic phospholipid binding yet with functional actin-binding sites. When expressed in cells, vinculin-LD was readily recruited to adhesion sites, as judged by fluorescence recovery after photobleaching (FRAP) analysis, but cell spreading and migration were strongly impaired, and PIP2-dependent disassembly of adhesions was suppressed. Thus, PIP2 binding is not essential for vinculin activation and recruitment, as previously suggested. Instead, we propose that PIP2 levels can regulate the uncoupling of adhesion sites from the actin cytoskeleton, with vinculin functioning as a sensor.

Modulation of cell spreading and cell-substrate adhesion dynamics by dystroglycan

“…Dystroglycan with a vinexin binding site mutation could not rescue cell spreading in dystroglycan-depleted cells on E3 laminin, suggesting a possible role for the dystroglycan-vinexinvinculin complex in cell spreading. A number of studies on vinculin-depleted and/or mutated cells have also documented a defect in cell spreading on laminin (Varnum-Finney and Reichardt, 1994;Volberg et al, 1995;Chandrasekar et al, 2005), and similarly to dystroglycan-depleted cells, these cells can still form adhesions because of compensatory changes in other adhesion proteins (Volberg et al, 1995). Furthermore, mutations in the regulatory PIP2-binding site in vinculin inhibit radial cell spreading and subsequent cell polarisation, and thus cell motility (Chandrasekar et al, 2005), a phenotype that is similar to that observed in dystroglycan-depleted myoblast cells.…”

Vinculin directly binds zonula occludens-1 and is essential for stabilizing connexin 43 containing gap junctions in cardiac Myocytes

“…Vcl can regulate the coupling of adhesion sites to the actin cytoskeleton (Chandrasekar et al, 2005), and is also known for binding to cadherin-based cell-cell contact sites through a-and b-catenin (Peng et al, 2010;Weiss et al, 1998). Recently it has been described that ZO-1 recruitment to a-catenin links the assembly of the endothelial barrier to adherens junctions (Maiers et al, 2013).…”

“…The acidic phospholipid phosphatidylinositol (4,5)-bisphosphate [PtdIns(4,5)P 2, PIP 2 ] directly regulates the actin cytoskeleton by modulating the activity and targeting of actin regulatory proteins (Yin and Janmey, 2003). Vcl binds PIP 2 , thereby acting as a sensor that can modify how phospholipids regulate the coupling of adhesion sites to the actin cytoskeleton (Chandrasekar et al, 2005;Palmer et al, 2009). Class I phosphoinositide 3-kinases (PI3Ks) phosphorylate phospholipids including PIP 2 , and act as scaffolding adapters (Costa and Hirsch, 2010).…”

Vinculin Is a Dually Regulated Actin Filament Barbed End-capping and Side-binding Protein

“…Although phosphatidylinositol 4,5-bisphosphate disrupts the intramolecular interaction Vh-Vt (50), it is unlikely that it unmasks the capping domain of Vt, because it also inhibits the binding of Vt to actin filaments (51). The interaction of phosphatidylinositol 4,5-bisphosphate with the C-terminal arm is also known to control the turnover of focal adhesions (52). Whether the regulation of the capping function by phosphatidylinositol 4,5-bisphosphate contributes to the regulation of focal adhesion turnover is an interesting open issue.…”