Tyrosine phosphorylation of SLP-76 is downstream of Syk following stimulation of the collagen receptor in platelets
Tyrosine phosphorylation of SLP-76 is downstream of Syk following stimulation of the collagen receptor in platelets
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DOI：
10.1074/jbc.274.9.5963
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发表时间：
1999-02-26
影响因子：
4.8
通讯作者：
Watson, SP
中科院分区：
文献类型：
--
作者：
Gross, BS;Lee, JR;Watson, SP
胶原相关肽（CRP）是一种胶原同源物，通过酪氨酸激酶依赖途径诱导血小板活化，导致Pc受体(FcR) γ链、Syk和磷脂酶c - γ的顺序酪氨酸磷酸化(2)。在这里，我们报道了CRP和血小板低亲和力免疫受体Fc γ RIIA刺激T细胞适配器SLP-76的酪氨酸磷酸化，而G蛋白偶联受体激动剂凝血酶仅诱导轻微的酪氨酸磷酸化。这表明SLP-76在受体下游具有特定作用，通过免疫受体酪氨酸激活基元发出信号。免疫沉淀研究表明，SLP-76与crp刺激血小板中的slp -130、Vav、Fyn、Lyn和FcR γ链有关。在体外对SLP-76免疫沉淀物进行的激酶测定中，这些蛋白中的一些，包括SLP-76，经历酪氨酸磷酸化。在体外激酶实验中，所有这些蛋白的酪氨酸磷酸化都被Src家族激酶抑制剂PP1所消除，这表明它是由Fyn或Lyn介导的。然而，这一生理意义尚不确定，因为在体内，在Fyn或Lyn缺陷血小板中，SLP-76的酪氨酸磷酸化都没有改变，因此，对Syk缺陷血小板的CRP刺激表明，体内SLP-76的酪氨酸磷酸化位于Syk的下游。SLP-76免疫沉淀中Syk的缺失增加了另一种蛋白质将SLP-76带入Syk的可能性，候选蛋白包括那些与SLP-76共同免疫沉淀的蛋白质，包括FcR γ链，plc - γ的酪氨酸磷酸化(2)和Ca2+动员在CRP刺激后SLP-76缺陷血小板中显着减弱，这表明该受体在磷脂酶的调节中起着关键作用。slp -130响应CRP的酪氨酸磷酸化的增加在SLP-76缺乏的血小板中也被抑制，使其处于SLP-76的下游。这项研究确定SLP-76是一个重要的适配器分子，由Syk调节，位于slp -130和plc - γ(2)的上游。
Collagen-related peptide (CRP), a collagen homologue, induces platelet activation through a tyrosine kinase-dependent pathway, leading to sequential tyrosine phosphorylation of Pc receptor (FcR) gamma-chain, Syk, and phospholipase C-gamma(2). Here we report that CRP and the platelet low affinity immune receptor Fc gamma RIIA stimulate tyrosine phosphorylation of the T cell adapter SLP-76, whereas the G protein-coupled receptor agonist thrombin induces only minor tyrosine phosphorylation. This suggests that SLP-76 has a specific role downstream of receptors that signal via an immunoreceptor tyrosine-based activation motif, Immunoprecipitation studies demonstrate association of SLP-76 with SLAP-130, Vav, Fyn, Lyn, and the FcR gamma-chain in CRP-stimulated platelets. Several of these proteins, including SLP-76, undergo tyrosine phosphorylation in in vitro kinase assays performed on SLP-76 immunoprecipitates. Tyrosine phosphorylation of all of these proteins in the in vitro kinase assay was abrogated by the Src family kinase inhibitor PP1, suggesting that it is mediated by either Fyn or Lyn, The physiological significance of this is uncertain, however, since tyrosine phosphorylation of SLP-76 in vivo is not altered in either Fyn- or Lyn-deficient platelets, CRP stimulation of Syk-deficient platelets demonstrated that in vivo tyrosine phosphorylation of SLP-76 is downstream of Syk, The absence of Syk in the SLP-76 immunoprecipitates raises the possibility that another protein is responsible for bringing SLP-76 to Syk, Candidates for this include those proteins that co-immunoprecipitate with SLP-76, including the FcR gamma-chain, Tyrosine phosphorylation of PLC-gamma(2) and Ca2+ mobilization is markedly attenuated in SLP-76-deficient platelets following CRP stimulation, suggesting that the adapter plays a critical role in the regulation of the phospholipase. The increase in tyrosine phosphorylation of SLAP-130 in response to CRP is also inhibited in SLP-76-deficient platelets, placing it downstream of SLP-76, This work identifies SLP-76 as an important adapter molecule that is regulated by Syk and lies upstream of SLAP-130 and PLC-gamma(2) in CRP-stimulated platelets.