Abstract
Conformational activation of integrins is generally required for ligand binding and cellular signalling. However, we have previously reported that the nonactivated conformation of α2β1 integrin can also bind to large ligands, such as human echovirus 1. In this study, we show that the interaction between the nonactivated integrin and a ligand resulted in the activation of focal adhesion kinase (FAK) in a protein kinase C dependent manner. A loss-of-function mutation, α2E336A, in the α2-integrin did not prevent the activation of FAK, nor did EDTA-mediated inactivation of the integrin. Full FAK activation was observed, since phosphorylation was not only confirmed in residue Y397, but also in residues Y576/7. Furthermore, initiation of downstream signaling by paxillin phosphorylation in residue Y118 was evident, even though this activation was transient by nature, probably due to the lack of talin involvement in FAK activation and the absence of vinculin in the adhesion complexes formed by the nonactivated integrins. Altogether these results indicate that the nonactivated integrins can induce cellular signaling, but the outcome of the signaling differs from conventional integrin signaling.
Highlights
The well-established model of integrin function stresses the importance of the conformational change from a nonactivated, bent conformation to an active, extended state prior to ligand binding and outside-in signaling[1]
We have previously showed that human echovirus 1 (EV1) binds with a higher avidity to the closed than to the open αI domain in the α2β1 integrin
Based on the confocal microscopy and immunofluorescence utilizing specific antibodies, the α2β1 integrin is located at the tips of the cellular protrusions, suggesting that these are the main plasma membrane areas involved in the EV1 interaction with the cell (Fig. 1b)
Summary
The well-established model of integrin function stresses the importance of the conformational change from a nonactivated, bent conformation to an active, extended state prior to ligand binding and outside-in signaling[1]. According to the conventional model of integrin outside-in signaling, the conformational changes in the ligand binding inserted domain (αI domain, often called as αA domain), and in the βI domain, lead to the separation of integrin legs, including transmembrane and intracellular domains. This allows intracellular signaling molecules to interact with integrins, which initiates the formation of focal adhesion sites[6]. EV1 binding does not seem to cause a conformational change in α2β1, whereas one virus particle can cluster several integrins together[15] This gave us an opportunity to utilize the interaction between EV1 and α2β1 integrin to analyse the signaling mechanisms related to nonactivated integrins. The phosphorylation of paxillin seems to be transient in nature, unlike after cell adhesion to collagen I
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