scholarly journals Evidence of a Role for SHP-1 in Platelet Activation by the Collagen Receptor Glycoprotein VI

2000 ◽  
Vol 275 (37) ◽  
pp. 28526-28531 ◽  
Author(s):  
Jean-Max Pasquet ◽  
Lynn Quek ◽  
Sophie Pasquet ◽  
Alastair Poole ◽  
James R. Matthews ◽  
...  
Keyword(s):  
Platelet Activation ◽  
Glycoprotein Vi ◽  
Collagen Receptor

A new monoclonal antibody, mAb 204-11, that influences the binding of platelet GPVI to fibrous collagen

2003 ◽  
Vol 89 (06) ◽  
pp. 996-1003 ◽  
Author(s):  
Jun Mizuguchi ◽  
Sachiko Kawashima ◽  
Michiko Nagamatsu ◽  
Yoshiki Miura ◽  
Tomohiro Nakagaki ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Platelet Aggregation ◽  
Binding Site ◽  
Tyrosine Phosphorylation ◽  
Platelet Activation ◽  
Collagen Binding ◽  
Glycoprotein Vi ◽  
Phosphorylation Of Proteins ◽  
Dimeric Form ◽  
Collagen Receptor

SummaryThe newly identified platelet collagen receptor glycoprotein VI binds to fibrous collagen, inducing platelet activation. Several antibodies against GPVI have been reported, including a patient’s auto-antibodies, that activates platelets through their ability to crosslink this glycoprotein. We have developed a monoclonal antibody (mAb) against GPVI using the recombinant extracellular domain of GPVI as an antigen. This antibody, mAb 204-11, induced platelet aggregation and tyrosine phosphorylation of proteins similar to those induced by GPVI-reactive proteins, collagen and convulxin. Its interaction with GPVI was analyzed by measuring the effect of the antibody on GPVI binding to collagen using a dimeric form of recombinant GPVI, GPVI-Fc2. MAb 204-11 inhibited the binding of GPVI-Fc2 to fibrous collagen particles, but enhanced the GPVI binding to immobilized collagen, suggesting that the antibody binds to a region near the collagen binding site of GPVI. MAb 204-11 also inhibited the GPVI binding to convulxin at a low concentration, but not completely. Since mAb 204-11 reacts specifically with GPVI and is applicable for immunoblotting and immunoprecipitation, this antibody would be useful for studies on GPVI.

Identification of P2Y12-dependent and -independent mechanisms of glycoprotein VI–mediated Rap1 activation in platelets

Blood ◽  
2003 ◽  
Vol 101 (4) ◽  
pp. 1409-1415 ◽  
Author(s):  
Mark K. Larson ◽  
Hong Chen ◽  
Mark L. Kahn ◽  
Anne M. Taylor ◽  
Jean-Etienne Fabre ◽  
...  
Keyword(s):  
Platelet Activation ◽  
Adenosine Diphosphate ◽  
Human Platelets ◽  
Independent Component ◽  
Glycoprotein Vi ◽  
Platelet Signaling ◽  
Guanosine Triphosphatase ◽  
Adp Receptor Antagonists ◽  
Rap1 Activation ◽  
Collagen Receptor

Glycoprotein (GP) VI is a critical platelet collagen receptor, yet the steps involved in GPVI-mediated platelet activation remain incompletely understood. Because activation of Rap1, an abundant small guanosine triphosphatase (GTPase) in platelets, contributes to integrin αIIbβ3 activation, we asked whether and how GPVI signaling activates Rap1 in platelets. Here we show that platelet Rap1 is robustly activated upon addition of convulxin, a GPVI-specific agonist. Using a reconstituted system in RBL-2H3 cells, we found that GPVI-mediated Rap1 activation is dependent on FcRγ but independent of another platelet collagen receptor, α2β1. Interestingly, GPVI-mediated Rap1 activation in human platelets is largely dependent on adenosine diphosphate (ADP) signaling through the P2Y12 and not the P2Y1 receptor. However, experiments with specific ADP receptor antagonists and platelets from knockout mice deficient in P2Y1 or the P2Y12-associated G-protein, Gαi2, indicate that human and murine platelets also have a significant P2Y12-independent component of GPVI-mediated Rap1 activation. The P2Y12-independent component is dependent on phosphatidylinositol 3-kinase and is augmented by epinephrine-mediated signaling. P2Y12-dependent and -independent components are also observed in GPVI-mediated platelet aggregation, further supporting a role for Rap1 in aggregation. These results define mechanisms of GPVI-mediated platelet activation and implicate Rap1 as a key signaling protein in GPVI-induced platelet signaling.

scholarly journals Physical and Functional Association of the Src Family Kinases Fyn and Lyn with the Collagen Receptor Glycoprotein VI-Fc Receptor γ Chain Complex on Human Platelets

1998 ◽  
Vol 188 (2) ◽  
pp. 267-276 ◽  
Author(s):  
Yasuharu Ezumi ◽  
Keisuke Shindoh ◽  
Masaaki Tsuji ◽  
Hiroshi Takayama
Keyword(s):  
Tyrosine Phosphorylation ◽  
Platelet Activation ◽  
Critical Role ◽  
Specific Inhibitor ◽  
Chain Complex ◽  
Human Platelets ◽  
Fc Receptor ◽  
Cross Linking ◽  
Glycoprotein Vi ◽  
Collagen Receptor

We have previously shown that uncharacterized glycoprotein VI (GPVI), which is constitutively associated and coexpressed with Fc receptor γ chain (FcRγ) in human platelets, is essential for collagen-stimulated tyrosine phosphorylation of FcRγ, Syk, and phospholipase Cγ2 (PLCγ2), leading to platelet activation. Here we investigated involvement of the Src family in the proximal signals through the GPVI–FcRγ complex, using the snake venom convulxin from Crotalus durissus terrificus, which specifically recognizes GPVI and activates platelets through cross-linking GPVI. Convulxin-coupled beads precipitated the GPVI–FcRγ complex from platelet lysates. Collagen and convulxin induced tyrosine phosphorylation of FcRγ, Syk, and PLCγ2 and recruited tyrosine-phosphorylated Syk to the GPVI–FcRγ complex. Using coprecipitation methods with convulxin-coupled beads and antibodies against FcRγ and the Src family, we showed that Fyn and Lyn, but not Yes, Src, Fgr, Hck, and Lck, were physically associated with the GPVI–FcRγ complex irrespective of stimulation. Furthermore, Fyn was rapidly activated by collagen or cross-linking GPVI. The Src family–specific inhibitor PP1 dose-dependently inhibited collagen- or convulxin-induced tyrosine phosphorylation of proteins including FcRγ, Syk, and PLCγ2, accompanied by a loss of aggregation and ATP release reaction. These results indicate that the Src family plays a critical role in platelet activation via the collagen receptor GPVI–FcRγ complex.

The Role of ITAM- and ITIM-coupled Receptors in Platelet Activation by Collagen

2001 ◽  
Vol 86 (07) ◽  
pp. 276-288 ◽  
Author(s):  
Naoki Asazuma ◽  
Ben Atkinson ◽  
Oscar Berlanga ◽  
Denise Best ◽  
Regis Bobe ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Platelet Activation ◽  
T Lymphocyte ◽  
Pivotal Role ◽  
Membrane Domains ◽  
Glycoprotein Vi ◽  
Ig Superfamily ◽  
Collagen Receptor ◽  
Lymphocyte Receptors

SummaryThe major activation-inducing collagen receptor glycoprotein VI (GPVI) has been cloned within the last two years. It is a member of the Ig superfamily of proteins and is constitutively associated with the ITAM-bearing Fc receptor γ-chain (FcR γ-chain). GPVI signals through a pathway that involves several of the proteins used by Fc, B- and T-lymphocyte receptors and which takes place in glycolipid-enriched membrane domains in the plasma membrane known as GEMs. Responses to GPVI are regulated by PECAM-1 (CD31) and possibly other ITIM-bearing receptors. Despite a pivotal role for GPVI, there are important differences between signalling events to collagen and GPVI-specific ligands. This may reflect a role for co-receptors in the response to collagen.

Interactions of 12-lipoxygenase with phospholipase A2isoforms following platelet activation through the glycoprotein VI collagen receptor

FEBS Letters ◽  
2004 ◽  
Vol 576 (1-2) ◽  
pp. 165-168 ◽  
Author(s):  
Marcus J. Coffey ◽  
Barbara Coles ◽  
Matthew Locke ◽  
Alexandra Bermudez-Fajardo ◽  
P.Claire Williams ◽  
...  
Keyword(s):  
Platelet Activation ◽  
Glycoprotein Vi ◽  
12 Lipoxygenase ◽  
Collagen Receptor

Tec regulates platelet activation by GPVI in the absence of Btk

Blood ◽  
2003 ◽  
Vol 102 (10) ◽  
pp. 3592-3599 ◽  
Author(s):  
Ben T. Atkinson ◽  
Wilfried Ellmeier ◽  
Steve P. Watson
Keyword(s):  
Platelet Activation ◽  
Functional Role ◽  
Adenosine Diphosphate ◽  
Human Platelets ◽  
Essential Requirement ◽  
Glycoprotein Vi ◽  
Tec Family Kinase ◽  
A Minor ◽  
Collagen Receptor

AbstractThe Tec family kinase Btk plays an important role in the regulation of phospholipase Cγ2 (PLCγ2) downstream of the collagen receptor glycoprotein VI (GPVI) in human platelets. Platelets also express a second member of this family, Tec; however, its function has not been analyzed. To address the role of Tec, we analyzed Btk-/-, Tec-/-, and Btk/Tec double-deficient (Btk-/-/Tec-/-) platelets. Tec-/- platelets exhibit a minor reduction in aggregation to threshold concentrations of collagen or the GPVI-specific agonist collagen-related peptide (CRP), whereas responses to higher concentrations are normal. Tyrosine phosphorylation of PLCγ2 by collagen and CRP is not altered in Tec-/- platelets. However, Btk-/-/Tec-/- platelets exhibit a greater reduction in PLCγ2 phosphorylation than is seen in the absence of Btk, thus revealing an important role for Tec in this situation. Furthermore, Btk-/-/Tec-/- platelets fail to undergo an increase in Ca2+, aggregation, secretion, and spreading in response to collagen or CRP, whereas they aggregate normally to adenosine diphosphate (ADP) and spread on fibrinogen. A residual GPVI signal exists in the Btk-/-/Tec-/- platelets as CRP synergizes with ADP to mediate aggregation. These results demonstrate an essential requirement for Tec and Btk in platelet activation by GPVI and reveal a functional role for Tec in the regulation of PLCγ2 in the absence of Btk. (Blood. 2003;102: 3592-3599)

scholarly journals Platelet Ca2+ responses coupled to glycoprotein VI and Toll-like receptors persist in the presence of endothelial-derived inhibitors: roles for secondary activation of P2X1 receptors and release from intracellular Ca2+ stores

Blood ◽  
2012 ◽  
Vol 119 (15) ◽  
pp. 3613-3621 ◽  
Author(s):  
C. Y. Eleanor Fung ◽  
Sarah Jones ◽  
Adwoa Ntrakwah ◽  
Khalid M. Naseem ◽  
Richard W. Farndale ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Phospholipase C ◽  
Platelet Activation ◽  
Secretory Pathway ◽  
Atp Release ◽  
Activation Mechanism ◽  
Glycoprotein Vi ◽  
Kinase C ◽  
Spermine Nonoate ◽  
Secondary Activation

Abstract Inhibition of Ca2+ mobilization by cyclic nucleotides is central to the mechanism whereby endothelial-derived prostacyclin and nitric oxide limit platelet activation in the intact circulation. However, we show that ∼ 50% of the Ca2+ response after stimulation of glycoprotein VI (GPVI) by collagen, or of Toll-like 2/1 receptors by Pam3Cys-Ser-(Lys)4 (Pam3CSK4), is resistant to prostacyclin. At low agonist concentrations, the prostacyclin-resistant Ca2+ response was predominantly because of P2X1 receptors activated by ATP release via a phospholipase-C–coupled secretory pathway requiring both protein kinase C and cytosolic Ca2+ elevation. At higher agonist concentrations, an additional pathway was observed because of intracellular Ca2+ release that also depended on activation of phospholipase C and, for TLR 2/1, PI3-kinase. Secondary activation of P2X1-dependent Ca2+ influx also persisted in the presence of nitric oxide, delivered from spermine NONOate, or increased ectonucleotidase levels (apyrase). Surprisingly, apyrase was more effective than prostacyclin and NO at limiting secondary P2X1 activation. Dilution of platelets reduced the average extracellular ATP level without affecting the percentage contribution of P2X1 receptors to collagen-evoked Ca2+ responses, indicating a highly efficient activation mechanism by local ATP. In conclusion, platelets possess inhibitor-resistant Ca2+ mobilization pathways, including P2X1 receptors, that may be particularly important during early thrombotic or immune-dependent platelet activation.

scholarly journals Mice Lacking the Inhibitory Collagen Receptor LAIR-1 Exhibit a Mild Thrombocytosis and Hyperactive Platelets

2017 ◽  
Vol 37 (5) ◽  
pp. 823-835 ◽  
Author(s):  
Christopher W. Smith ◽  
Steven G. Thomas ◽  
Zaher Raslan ◽  
Pushpa Patel ◽  
Maxwell Byrne ◽  
...  
Keyword(s):  
Tyrosine Kinases ◽  
Kinase Activity ◽  
Thrombus Formation ◽  
Physiological Role ◽  
Src Family Kinase ◽  
Glycoprotein Vi ◽  
Collagen Receptor ◽  
Deficient Mice

Objective— Leukocyte-associated immunoglobulin-like receptor-1 (LAIR-1) is a collagen receptor that belongs to the inhibitory immunoreceptor tyrosine-based inhibition motif–containing receptor family. It is an inhibitor of signaling via the immunoreceptor tyrosine-based activation motif–containing collagen receptor complex, glycoprotein VI-FcRγ-chain. It is expressed on hematopoietic cells, including immature megakaryocytes, but is not detectable on platelets. Although the inhibitory function of LAIR-1 has been described in leukocytes, its physiological role in megakaryocytes and in particular in platelet formation has not been explored. In this study, we investigate the role of LAIR-1 in megakaryocyte development and platelet production by generating LAIR-1–deficient mice. Approach and Results— Mice lacking LAIR-1 exhibit a significant increase in platelet counts, a prolonged platelet half-life in vivo, and increased proplatelet formation in vitro. Interestingly, platelets from LAIR-1–deficient mice exhibit an enhanced reactivity to collagen and the glycoprotein VI–specific agonist collagen-related peptide despite not expressing LAIR-1, and mice showed enhanced thrombus formation in the carotid artery after ferric chloride injury. Targeted deletion of LAIR-1 in mice results in an increase in signaling downstream of the glycoprotein VI–FcRγ-chain and integrin αIIbβ3 in megakaryocytes because of enhanced Src family kinase activity. Conclusions— Findings from this study demonstrate that ablation of LAIR-1 in megakaryocytes leads to increased Src family kinase activity and downstream signaling in response to collagen that is transmitted to platelets, rendering them hyper-reactive specifically to agonists that signal through Syk tyrosine kinases, but not to G-protein–coupled receptors.

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