Article

Structural Basis of Integrin Activation by Talin

Department of Biochemistry, University of Oxford, South Parks Road, Oxford OX1 3QU, England, UK.
Cell (Impact Factor: 32.24). 02/2007; 128(1):171-82. DOI: 10.1016/j.cell.2006.10.048
Source: PubMed

ABSTRACT

Regulation of integrin affinity (activation) is essential for metazoan development and for many pathological processes. Binding of the talin phosphotyrosine-binding (PTB) domain to integrin beta subunit cytoplasmic domains (tails) causes activation, whereas numerous other PTB-domain-containing proteins bind integrins without activating them. Here we define the structure of a complex between talin and the membrane-proximal integrin beta3 cytoplasmic domain and identify specific contacts between talin and the integrin tail required for activation. We used structure-based mutagenesis to engineer talin and beta3 variants that interact with comparable affinity to the wild-type proteins but inhibit integrin activation by competing with endogenous talin. These results reveal the structural basis of talin's unique ability to activate integrins, identify an interaction that could aid in the design of therapeutics to block integrin activation, and enable engineering of cells with defects in the activation of multiple classes of integrins.

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Available from: Kate Louise Wegener
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    • "TLN consists of a N-terminal head and a C-terminal flexible rod domain. The head FERM domain binds membrane-proximal motifs of the -integrin tail which activates the integrin (Vinogradova et al., 2002; Jiang et al., 2003; Tadokoro et al., 2003; Wegener et al., 2007; Critchley, 2009; Moser et al., 2009; Kim et al., 2012; Pinon et al., 2014). The rod provides the major part of actin binding via actin binding sites (ABS), in particular the C-terminal ABS3 (Hemmings et al., 1996; Giannone et al., 2003; Jiang et al., 2003; Gingras et al., 2008). "
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    ABSTRACT: The molecular clutch (MC) model proposes that actomyosin-driven force transmission permits integrin-dependent cell migration. To investigate the MC, we introduced diverse talin (TLN) and integrin variants into Flp-In™ T-Rex™ HEK293 cells stably expressing uPAR. Vitronectin variants served as substrate providing uPAR-mediated cell adhesion and optionally integrin binding. This particular system allowed us to selectively analyse key MC proteins and interactions, effectively from the extracellular matrix substrate to intracellular f-actin, and to therewith study mechanobiological aspects of MC engagement also uncoupled from integrin/ligand binding. With this experimental approach, we found that for the initial PIP2-dependent membrane/TLN/f-actin linkage and persistent lamellipodia formation the C-terminal TLN actin binding site (ABS) is dispensable. The establishment of an adequate MC-mediated lamellipodial tension instead depends predominantly on the coupling of this C-terminal TLN ABS to the actomyosin-driven retrograde actin flow force. This lamellipodial tension is crucial for full integrin activation eventually determining integrin-dependent cell migration. In the integrin/ligand-independent condition the frictional membrane resistance participates to these processes. Integrin/ligand binding can also contribute but is not necessarily required.
    Full-text · Article · Oct 2015 · European Journal of Cell Biology
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    • "The retention of slow rolling also suggests that shear forces applied to L2- ICAM-1 bonds are sufficient for integrin extension (Chen et al., 2012). In static systems, the L325R mutation prevents the talin-induced tilting of the 3 transmembrane domain required for allosteric activation of IIb3 (Wegener et al., 2007; Kim et al., 2012). Our finding that talin binding to the MP domain of the  subunit, which is blocked by the L325R mutation, is required for arrest indicates that the forces experienced during slow rolling are not sufficient to cause the headpiece opening associated with neutrophil arrest, even in the presence of chemokines. "
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    ABSTRACT: Neutrophil recruitment, mediated by β2 integrins, combats pyogenic infections but also plays a key role in ischemia-reperfusion injury and other inflammatory disorders. Talin induces allosteric rearrangements in integrins that increase affinity for ligands (activation). Talin also links integrins to actin and other proteins that enable formation of adhesions. Structural studies have identified a talin1 mutant (L325R) that perturbs activation without impairing talin's capacity to link integrins to actin and other proteins. Here, we found that mice engineered to express only talin1(L325R) in myeloid cells were protected from renal ischemia-reperfusion injury. Dissection of neutrophil function in vitro and in vivo revealed that talin1(L325R) neutrophils had markedly impaired chemokine-induced, β2 integrin-mediated arrest, spreading, and migration. Surprisingly, talin1(L325R) neutrophils exhibited normal selectin-induced, β2 integrin-mediated slow rolling, in sharp contrast to the defective slow rolling of neutrophils lacking talin1 or expressing a talin1 mutant (W359A) that blocks talin interaction with integrins. These studies reveal the importance of talin-mediated activation of integrins for renal ischemia-reperfusion injury. They further show that neutrophil arrest requires talin recruitment to and activation of integrins. However, although neutrophil slow rolling requires talin recruitment to integrins, talin-mediated integrin activation is dispensable. © 2015 Yago et al.
    Full-text · Article · Jul 2015 · Journal of Experimental Medicine
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    • "A cluster of negatively charged residues at the a IIb CT MD region may disfavor the negatively charged cell membrane and inhibit the membrane embedding of a IIb TM-CT, while an interaction between the talin-head and the a IIb CT MD acidic residues as indicated by a molecular dynamic simulation study (Provasi et al., 2014) might facilitate the membrane embedding of a IIb TM-CT to activate integrin. In addition, the capability of talin to activate integrin depends on its binding to both b-integrin CT and cell membrane (Wegener et al., 2007;Anthis et al., 2009;Moore et al., 2012;Song et al., 2012;Kalli et al., 2013). The interaction between a CT and talin might help talin maintain a proper orientation for the favorable interaction with b-integrin CT and cell membrane in order to stabilize the active conformation of TM-CT (supplementary material Fig. S4C). "
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    ABSTRACT: Studies on the mechanism of integrin inside-out activation have been focused on the role of β cytoplasmic tails that are relatively conserved and bear binding sites for the intracellular activators including talin and kindlin. Integrin α cytoplasmic tails share a conserved GFFKR motif at the membrane-proximal region forming specific interface with β membrane-proximal region that keeps integrin inactive. The α membrane-distal regions after the GFFKR motif are diverse both in length and sequence and their roles in integrin activation have not been well-defined. In this study, we report that the α cytoplasmic membrane-distal region contributes to maintaining integrin in the resting state and to integrin inside-out activation. Complete deletion of the α membrane-distal region diminished talin and kindlin mediated integrin ligand binding and conformational change. A proper length and amino acids of α membrane-distal region is important for integrin inside-out activation. Our data establish an essential role of the α integrin cytoplasmic membrane-distal region in integrin activation and provide new insights into how talin and kindlin induce the high affinity integrin conformation that is required for fully functional integrins.
    Full-text · Article · Mar 2015 · Journal of Cell Science
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