Mori Jun, Pearce Andrew C, Spalton Jennifer C, Grygielska Beata, Eble Johannes A, Tomlinson Michael G, Senis Yotis A, Watson Steve P
Centre for Cardiovascular Sciences, Institute of Biomedical Research, University of Birmingham, B15 2TT, UK.
J Biol Chem. 2008 Dec 19;283(51):35419-27. doi: 10.1074/jbc.M806895200. Epub 2008 Oct 27.
Platelets play an essential role in wound healing by forming thrombi that plug holes in the walls of damaged blood vessels. To achieve this, platelets express a diverse array of cell surface receptors and signaling proteins that induce rapid platelet activation. In this study we show that two platelet glycoprotein receptors that signal via an immunoreceptor tyrosine-based activation motif (ITAM) or an ITAM-like domain, namely the collagen receptor complex glycoprotein VI (GPVI)-FcR gamma-chain and the C-type lectin-like receptor 2 (CLEC-2), respectively, support constitutive (i.e. agonist-independent) signaling in a cell line model using a nuclear factor of activated T-cells (NFAT) transcriptional reporter assay that can detect low level activation of phospholipase Cgamma (PLCgamma). Constitutive and agonist signaling by both receptors is dependent on Src and Syk family kinases, and is inhibited by G6b-B, a platelet immunoglobulin receptor that has two immunoreceptor tyrosine-based inhibitory motifs in its cytosolic tail. Mutation of the conserved tyrosines in the two immunoreceptor tyrosine-based inhibitory motifs prevents the inhibitory action of G6b-B. Interestingly, the inhibitory activity of G6b-B is independent of the Src homology 2 (SH2)-domain containing tyrosine phosphatases, SHP1 and SHP2, and the inositol 5'-phosphatase, SHIP. Constitutive signaling via Src and Syk tyrosine kinases is observed in platelets and is associated with tyrosine phosphorylation of GPVI-FcR gamma-chain and CLEC-2. We speculate that inhibition of constitutive signaling through Src and Syk tyrosine kinases by G6b-B may help to prevent unwanted platelet activation.
血小板在伤口愈合过程中发挥着至关重要的作用，它们通过形成血栓来堵塞受损血管壁上的漏洞。为实现这一功能，血小板表达了多种细胞表面受体和信号蛋白，这些蛋白可诱导血小板快速激活。在本研究中，我们发现两种通过免疫受体酪氨酸激活基序（ITAM）或ITAM样结构域进行信号传导的血小板糖蛋白受体，即胶原受体复合物糖蛋白VI（GPVI）-FcRγ链和C型凝集素样受体2（CLEC-2），在细胞系模型中利用可检测磷脂酶Cγ（PLCγ）低水平激活的活化T细胞核因子（NFAT）转录报告基因检测法，支持组成性（即不依赖激动剂）信号传导。两种受体的组成性和激动剂信号传导均依赖于Src和Syk家族激酶，并受到G6b-B的抑制，G6b-B是一种血小板免疫球蛋白受体，其胞质尾部有两个基于免疫受体酪氨酸的抑制基序。两个基于免疫受体酪氨酸的抑制基序中保守酪氨酸的突变可阻止G6b-B的抑制作用。有趣的是，G6b-B的抑制活性独立于含Src同源2（SH2）结构域的酪氨酸磷酸酶SHP1和SHP2以及肌醇5'-磷酸酶SHIP。在血小板中观察到通过Src和Syk酪氨酸激酶的组成性信号传导，并且这与GPVI-FcRγ链和CLEC-2的酪氨酸磷酸化相关。我们推测，G6b-B通过Src和Syk酪氨酸激酶抑制组成性信号传导可能有助于防止不必要的血小板激活。
