Integrin activation

Abstract

Agonist stimulation of integrin receptors, composed of transmembrane alpha and beta subunits, leads cells to regulate integrin affinity ('activation'), a process that controls cell adhesion and migration, and extracellular matrix assembly. A final step in integrin activation is the binding of talin to integrin beta cytoplasmic domains. We used forward, reverse and synthetic genetics to engineer and order integrin activation pathways of a prototypic integrin, platelet alphaIIbbeta3. PMA activated alphaIIbbeta3 only after expression of both PKCalpha (protein kinase Calpha) and talin at levels approximating those in platelets. Inhibition of Rap1 GTPase reduced alphaIIbbeta3 activation, whereas expression of constitutively active Rap1A(G12V) bypassed the requirement for PKCalpha. Overexpression of a Rap effector, RIAM (Rap1-GTP-interacting adaptor molecule), activated alphaIIbbeta3 and bypassed the requirement for PKCalpha and Rap1. In addition, shRNA (short hairpin RNA)-mediated knockdown of RIAM blocked talin interaction with and activation of integrin alphaIIbbeta3. Rap1 activation caused the formation of an 'activation complex' containing talin and RIAM that redistributed to the plasma membrane and activated alphaIIbbeta3. The central finding was that this Rap1-induced formation of an 'integrin activation complex' leads to the unmasking of the integrin-binding site on talin, resulting in integrin activation.

Figure 1. Model of talin-induced integrin activation

Left: the talin F3 domain (surface representation; coloured by charge), freed from its autoinhibitory interactions in the full-length protein, becomes available for binding to the integrin. Middle: F3 engages the membrane-distal (MD) part of the β3-integrin tail (in red), which becomes ordered, but the α–β integrin interactions that hold the integrin in the low-affinity conformation remain intact. Right: in a subsequent step, F3 engages the membrane-proximal (MP) portion of the β3 tail while maintaining its MD interactions. Consequences of this additional interaction are: (i) destabilization of the putative integrin salt bridge; (ii) stabilization of the helical structure of the MP region; and (iii) electrostatic interactions between F3 and the acidic lipid headgroups. The net result is a change in the position of the transmembrane helix, which is continuous with the MP β tail helix. This position change causes a packing mismatch with the αIIb transmembrane helix, separation or reorientation of the integrin tails and activation (inset). Mutants of F3 that have compromised interactions with the MP region and other PTB domains that lack an MP-binding site stall at point B, consistent with their dominant-negative behaviour. Reprinted from Cell, 128, Wegener, K.L., Partridge, A.W., Han, J., Pickford, A.R., Liddington, R.C., Ginsberg, M.H. and Campbell, I.D., ’Structural basis of integrin activation by talin’, pp. 171–182. © 2007, with permission from Elsevier.

Figure 2. Connecting agonist stimulation to integrin activation

Depicted are core connections between agonists and integrins. Agonist receptors (e.g. G-protein-coupled receptors, tyrosine-kinase-coupled receptors) induce the formation of DAG (diacylglycerol) and increased Ca²⁺, leading to the activation and/or translocation of active GTP-bound Rap1 to the plasma membrane via activation of PKC or a Rap-GEF. At the plasma membrane, activated Rap interacts with RIAM, leading to the recruitment of talin to form the integrin-activation complex, thus unmasking the integrin-binding site on talin, leading to integrin activation. Reprinted from Curr. Biol., 16, Han, J., Lim, C.J., Watanabe, N., Soriani, A., Ratnikov, B., Calderwood, D.A., Puzon-McLaughlin, W., Lafuente, E.M., Boussiotis, V.A., Shattil, S.J. and Ginsberg, M.H., ’Reconstructing and deconstructing agonist-induced activation of integrin αIIbβ3’, pp. 1796–1806. © 2006, with permission from Elsevier.