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The mechanism of kindlin-mediated activation of integrin ?IIb?3.

ABSTRACT: Increased ligand binding to cellular integrins ("activation") plays important roles in processes such as development, cell migration, extracellular matrix assembly, tumor metastasis, hemostasis, and thrombosis. Integrin activation encompasses both increased integrin monomer affinity and increased receptor clustering and depends on integrin-talin interactions. Loss of kindlins results in reduced activation of integrins. Kindlins might promote talin binding to integrins through a cooperative mechanism; however, kindlins do not increase talin association with integrins. Here, we report that, unlike talin head domain (THD), kindlin-3 has little effect on the affinity of purified monomeric ?IIb?3, and it does not enhance activation by THD. Furthermore, studies with ligands of varying valency show that kindlins primarily increase cellular ?IIb?3 avidity rather than monomer affinity. In platelets or nucleated cells, loss of kindlins markedly reduces ?IIb?3 binding to multivalent but not monovalent ligands. Finally, silencing of kindlins reduces the clustering of ligand-occupied ?IIb?3 as revealed by total internal reflection fluorescence and electron microscopy. Thus, in contrast to talins, kindlins have little primary effect on integrin ?IIb?3 affinity for monovalent ligands and increase multivalent ligand binding by promoting the clustering of talin-activated integrins.

SUBMITTER: Ye F

PROVIDER: S-EPMC3912999 | biostudies-literature | 2013 Nov

REPOSITORIES: biostudies-literature

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The mechanism of kindlin-mediated activation of integrin αIIbβ3.

Ye Feng F Petrich Brian G BG Anekal Praju P Lefort Craig T CT Kasirer-Friede Ana A Shattil Sanford J SJ Ruppert Raphael R Moser Markus M Fässler Reinhard R Ginsberg Mark H MH

Current biology : CB 20131107 22

Increased ligand binding to cellular integrins ("activation") plays important roles in processes such as development, cell migration, extracellular matrix assembly, tumor metastasis, hemostasis, and thrombosis. Integrin activation encompasses both increased integrin monomer affinity and increased receptor clustering and depends on integrin-talin interactions. Loss of kindlins results in reduced activation of integrins. Kindlins might promote talin binding to integrins through a cooperative mecha ...[more]

PMID: 24210614

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Project description:Integrin-linked kinase (ILK) is an important signaling regulator that assembles into the heteroternary complex with adaptor proteins PINCH and parvin (termed the IPP complex). We recently reported that ILK is important for integrin activation in a Chinese hamster ovary (CHO) cell system. We previously established parental CHO cells expressing a constitutively active chimeric integrin (?IIb?6B?3) and mutant CHO cells expressing inactive ?IIb?6B?3 due to ILK deficiency. In this study, we further investigated the underlying mechanisms for ILK-dependent integrin activation. ILK-deficient mutant cells had trace levels of PINCH and ?-parvin, and transfection of ILK cDNA into the mutant cells increased not only ILK but also PINCH and ?-parvin, resulting in the restoration of ?IIb?6B?3 activation. In the parental cells expressing active ?IIb?6B?3, ILK, PINCH, and ?-parvin were co-immunoprecipitated, indicating the formation of the IPP complex. Moreover, short interfering RNA (siRNA) experiments targeting PINCH-1 or both ?- and ?-parvin mRNA in the parent cells impaired the ?IIb?6B?3 activation as well as the expression of the other components of the IPP complex. In addition, ILK mutants possessing defects in either PINCH or parvin binding failed to restore ?IIb?6B?3 activation in the mutant cells. Kindlin-2 siRNA in the parental cells impaired ?IIb?6B?3 activation without disturbing the expression of ILK. For CHO cells stably expressing wild-type ?IIb?3 that is an inactive form, overexpression of a talin head domain (THD) induced ?IIb?3 activation and the THD-induced ?IIb?3 activation was impaired by ILK siRNA through a significant reduction in the expression of the IPP complex. In contrast, overexpression of all IPP components in the ?IIb?3-expressing CHO cells further augmented THD-induced ?IIb?3 activation, whereas they did not induce ?IIb?3 activation without THD. These data suggest that the IPP complex rather than ILK plays an important role and supports integrin activation probably through stabilization of the active conformation.

Dataset Information

The mechanism of kindlin-mediated activation of integrin ?IIb?3.

Publications

The mechanism of kindlin-mediated activation of integrin αIIbβ3.

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