These authors equally contributed to the work.
The proline-rich tyrosine kinase Pyk2 regulates platelet integrin αIIbβ3 outside-in signaling
Article first published online: 7 FEB 2013
© 2012 International Society on Thrombosis and Haemostasis
Journal of Thrombosis and Haemostasis
Volume 11, Issue 2, pages 345–356, February 2013
How to Cite
The proline-rich tyrosine kinase Pyk2 regulates platelet integrin αIIbβ3 outside-in signaling. J Thromb Haemost 2013; 11: 345–56., , , , , , , , , .
- Issue published online: 7 FEB 2013
- Article first published online: 7 FEB 2013
- Accepted manuscript online: 8 DEC 2012 04:10AM EST
- Manuscript Accepted: 15 NOV 2012
- Manuscript Received: 7 SEP 2012
- the Cariplo Foundation. Grant Number: 2011-0436
- the Ministero dell'Istruzione
- Università e Ricerca Scientifica (PRIN)
- Regione Lombardia and University of Pavia. Grant Number: REGLOM16
- British Heart Foundation. Grant Number: PG/06/022/20348
- phosphatidylinositol 3-kinase;
- platelet adhesion;
- signal transduction;
- tyrosine kinases
The proline-rich tyrosine kinase Pyk2 is a focal adhesion kinase expressed in blood platelets, and is activated downstream of G-protein coupled receptors as well as integrin α2β1.
In this study we have investigated the involvement of Pyk2 in integrin αIIbβ3 outside-in signaling in human and murine platelets.
We analyzed the stimulation of intracellular signaling pathways in platelets from Pyk2 knockout mice adherent to immobilized fibrinogen.
Pyk2 was rapidly phosphorylated and activated in human and murine platelets adherent to fibrinogen through integrin αIIbβ3. Activation of Pyk2 was Src-dependent, but did not require phospholipase Cγ2 activity. Platelets from Pyk2 knockout mice showed a defective ability to adhere and spread on fibrinogen, in association with a dramatic reduction of phosphatidylinositol 3-kinase (PI3K) activation and Akt phosphorylation. Pharmacological and genetic analysis demonstrated that integrin αIIbβ3 engagement selectively stimulated the β-isoform of PI3K (PI3Kβ), and that, as for Pyk2, PI3Kβ activation required Src family kinases activity, but not phospholipase Cγ2. In fibrinogen-adherent platelets, both Pyk2 and PI3Kβ were necessary for stimulation of the small GTPase Rap1b, a regulator of cell adhesion and spreading. Integrin αIIbβ3 engagement triggered the association of the PI3Kβ regulatory subunit p85 with the adaptor protein c-Cbl, which was mediated by the p85 SH3 domain, and was independent of c-Cbl tyrosine phosphorylation. However, p85-associated c-Cbl was tyrosine phosphorylated by activated Pyk2 in fibrinogen adherent platelets.
These results identify a novel pathway of integrin αIIbβ3 outside-in signaling and recognize the tyrosine kinase Pyk2 as a major regulator of platelet adhesion and spreading on fibrinogen.