The mechanisms and dynamics of αvβ3 integrin clustering in living cells

Caroline Cluzel,FréDéRique Paulhe,Jost Lussi,Beat A Imhof,FréDéRic Saltel,Bernhard Wehrle-Haller

doi:10.1083/jcb.200503017

Abstract

During cell migration, the physical link between the extracellular substrate and the actin cytoskeleton mediated by receptors of the integrin family is constantly modified. We analyzed the mechanisms that regulate the clustering and incorporation of activated αvβ3 integrins into focal adhesions. Manganese (Mn2+) or mutational activation of integrins induced the formation of de novo F-actin–independent integrin clusters. These clusters recruited talin, but not other focal adhesion adapters, and overexpression of the integrin-binding head domain of talin increased clustering. Integrin clustering required immobilized ligand and was prevented by the sequestration of phosphoinositole-4,5-bisphosphate (PI(4,5)P2). Fluorescence recovery after photobleaching analysis of Mn2+-induced integrin clusters revealed increased integrin turnover compared with mature focal contacts, whereas stabilization of the open conformation of the integrin ectodomain by mutagenesis reduced integrin turnover in focal contacts. Thus, integrin clustering requires the formation of the ternary complex consisting of activated integrins, immobilized ligands, talin, and PI(4,5)P2. The dynamic remodeling of this ternary complex controls cell motility.

Highlights

Integrins are transmembrane receptors linking the actin cytoskeleton to the ECM
Nuclear magnetic resonance, and electronmicroscopic studies have revealed an allosteric switch in ␣v␤3 integrin that is induced upon activation (Xiong et al, 2001, 2002; Hynes, 2002; Takagi et al, 2002; Vinogradova et al, 2002; Xiao et al, 2004)
Despite this detailed model of integrin activation, the mechanisms that result in its lateral clustering remain controversial

Summary

Introduction

Integrins are transmembrane receptors linking the actin cytoskeleton to the ECM. This connection is dynamically reorganized in response to mechanical, chemokine, and growth factor signals. The distal parts of both ectodomains swing open in a switchblade motion, followed by the separation of the transmembrane and cytoplasmic domains (Xiao et al, 2004). This allosteric switch, or “integrin activation,” can be induced by effector binding to the cytoplasmic tail of the ␣ and ␤ subunit (Vinogradova et al, 2002; Katagiri et al, 2003; Tadokoro et al, 2003) or by ligand binding to the ectodomain (Takagi et al, 2002). Once the integrin heterodimer is in its open configuration, the cytoplasmic domain of the ␤ integrin chain is linked via cytoplasmic adaptor proteins, such as talin, to the actin cytoskeleton (Liddington and Ginsberg, 2002; Critchley, 2004)

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