Vinculin controls talin engagement with the actomyosin machinery.

Paul Atherton,Pengbo Wang,Devina Jethwa,Eugenia Meiler-Rodriguez,Neil Bate,De-Yao Wang,David R Critchley,Ricky Tsang,Christoph Ballestrem,Igor L Barsukov,Benjamin T Goult,Roy Zent,Ben Stutchbury

doi:10.1038/ncomms10038

Abstract

The link between extracellular-matrix-bound integrins and intracellular F-actin is essential for cell spreading and migration. Here, we demonstrate how the actin-binding proteins talin and vinculin cooperate to provide this link. By expressing structure-based talin mutants in talin null cells, we show that while the C-terminal actin-binding site (ABS3) in talin is required for adhesion complex assembly, the central ABS2 is essential for focal adhesion (FA) maturation. Thus, although ABS2 mutants support cell spreading, the cells lack FAs, fail to polarize and exert reduced force on the surrounding matrix. ABS2 is inhibited by the preceding mechanosensitive vinculin-binding R3 domain, and deletion of R2R3 or expression of constitutively active vinculin generates stable force-independent FAs, although cell polarity is compromised. Our data suggest a model whereby force acting on integrin-talin complexes via ABS3 promotes R3 unfolding and vinculin binding, activating ABS2 and locking talin into an actin-binding configuration that stabilizes FAs.

Highlights

The link between extracellular-matrix-bound integrins and intracellular F-actin is essential for cell spreading and migration
Cell migration involves the cyclical attachment and detachment of the integrin family of adhesion molecules to extracellular matrix (ECM), as well as the generation of force required to translocate the cell body. Such events occur in focal adhesions (FA), dynamic macromolecular complexes in which integrins are linked via cytoplasmic adhesion plaque proteins to the actomyosin contractile machinery[1,2,3]
We demonstrate that vinculin binding within domains R2R3 acts to ‘unlock’ ABS2 of talin, and that this is regulated either by prior activation of vinculin, or by the application of force across talin provided by actin-binding to ABS3

Summary

Introduction

The link between extracellular-matrix-bound integrins and intracellular F-actin is essential for cell spreading and migration. Cell migration involves the cyclical attachment and detachment of the integrin family of adhesion molecules to extracellular matrix (ECM), as well as the generation of force required to translocate the cell body Such events occur in focal adhesions (FA), dynamic macromolecular complexes in which integrins are linked via cytoplasmic adhesion plaque proteins to the actomyosin contractile machinery[1,2,3]. Structural and biochemical studies on talin show that the vinculin binding sites (VBSs) within the helical bundles that make up the talin rod are cryptic[12,14,20,21,22], and single molecule experiments[9,23,24,25] indicate that force-induced unfolding of the bundles is required to unmask the VBSs. it is hypothesised that force-induced conformational changes in talin lead to the recruitment of vinculin and the stabilization of FAs. to what extent such mechanisms operate in cells is unclear. Structure–function studies on talin have been hampered by the fact that (i) most cell types express two structurally and functionally related talin isoforms[26,27], (ii) knockout or knockdown of talin[1] leads to upregulation of talin[24] and (iii) knockdown leaves residual talin[4,28], complicating the interpretation of results

Methods

Results

Conclusion