Grb2 Contributes to (hem)ITAM-Mediated Signaling in Platelets
Rationale: Platelets are anuclear cell fragments derived from bone marrow megakaryocytes (MKs) that safeguard vascular integrity but may also cause pathological vessel occlusion. One major pathway of platelet activation is triggered by two receptors that signal through an (hem) immunoreceptor tyrosine-based activation motif (ITAM), the activating collagen receptor glycoprotein (GP) VI and the C-type lectin-like receptor 2 (CLEC-2). Growth-factor receptor-bound protein 2 (Grb2) is a ubiquitously expressed adapter molecule involved in signaling processes of numerous receptors in different cell types, but its function in platelets and MKs is unknown.
Objective: We tested the hypothesis that Grb2 is a crucial adapter protein in (hem)ITAM signaling in platelets.
Methods and Results: Here we show that genetic ablation of Grb2 in MKs and platelets did not interfere with MK differentiation or platelet production. However, Grb2-deficiency severely impaired GPVI-mediated platelet activation due to defective stabilization of the LAT signalosome and activation of downstream signaling proteins which resulted in reduced adhesion, aggregation and coagulant activity on collagen in vitro. Similarly, CLEC-2-mediated signaling was impaired in Grb2-deficient platelets, whereas the cells responded normally to stimulation of G-protein coupled receptors (GPCR). In vivo this selective (hem)ITAM signaling defect resulted in prolonged bleeding times but affected arterial thrombus formation only after concomitant treatment with acetylsalicylic acid (ASA), indicating that defective GPVI signaling in the absence of Grb2 can be compensated through thromboxane A2 (TxA2)-induced GPCR signaling pathways.
Conclusions: These results reveal an important contribution of Grb2 in (hem)ITAM signaling in platelets in hemostasis and thrombosis by stabilizing the LAT signalosome.
- Received September 20, 2013.
- Revision received November 20, 2013.
- Accepted November 21, 2013.