Vinculin Regulates Assembly of Talin: β3 Integrin Complexes

Abstract

Vinculin is a talin-binding protein that promotes integrin-mediated cell adhesion, but the mechanisms are not understood. Because talin is a direct activator of integrins, we asked whether and how vinculin regulates the formation of integrin: talin complexes. We report that VD1 (aa 1-258) and its talin-binding mutant, VD1A50I, bind directly and equally to several β integrin cytoplasmic tails (βCT). Results from competition assays show that VD1, but not VD1A50I, inhibits the interaction of talin (Tn) and talin rod (TnR), but not talin head (TnH) with β3CT. The inhibition observed could be the result of VD1 binding to one or more of the 11 vinculin binding sites (VBSs) in the TnR domain. Our studies demonstrate that VD1 binding to amino acids 482-911, a VBS rich region, in TnR perturbs the interaction of rod with β3CT. The integrin activation assays done using CHOA5 cells show that activated vinculin enhances αIIbβ3 integrin activation and that the effect is dependent on talin. The TnR domain however shows no integrin activation unlike TnH that shows enhanced integrin activation. The overall results indicate that activated vinculin promotes talin-mediated integrin activation by binding to accessible VBSs in TnR and thus displacing the TnR from the β3 subunit. The study presented, defines a novel direct interaction of VD1 with β3CT and provides an attractive explanation for vinculin's ability to potentiate integrin-mediated cell adhesion through directly binding to both TnR and the integrin cytoplasmic tail.

Computational and experimental analysis of bioactive peptide linear motifs in the integrin adhesome

“…Vinculin present in focal adhesions may be tightly bound to ACTN1 and other proteins, and therefore not available for binding with peptide. Since vinculin has roles in both integrin activation and inhibition[14], the co-localization of peptide with vinculin seen here could be consistent with a damping of its potential inhibitory effects, but may act through a number of potential mechanisms.…”

“…Adhesome proteins that are embryonic lethal (see S1 Table) tend to have many interactors (i.e. are “hub” proteins, such as integrin, paxillin, Grb2 and FAK), while those with fewer interactors such as tensin, vimentin and IAP have less deleterious consequences [8][14]. Knockout of the highly connected Src is not lethal, perhaps because of compensation by the homologous kinases Lyn and Fyn[8].…”

Microenvironmental Stiffness Regulates Dental Papilla Cell Differentiation: Implications for the Importance of Fibronectin–Paxillin−β-Catenin Axis

“…Integrins as transmembrane proteins have the function of connecting the cytoskeleton and modulating intracellular signals, with respect to the cell adhesion . Integrins are αβ heterodimeric, and integrin β1 is the most ubiquitously expressed subunit, which links with a number of subunits, resulting in diverse functions . In recent years, integrin β3 has entered our line of sight.…”

Vinculin in cell–cell and cell–matrix adhesions

“…The recruitment of vinculin to talin stabilizes focal adhesions and promotes integrin clustering and enlargement [38, 39]. More recent work shows vinculin directly regulates integrin activation through talin [39–41]. Investigations into how this occurs suggest when a cell signals for increased integrin activation, talin is redistributed from the cytoplasm to the plasma membrane.…”