Outside-in signaling mechanisms of platelet integrin alpha-llb-beta3

血小板整合素α-IIb-β3由外而内的信号传导机制

• 批准号: 8186790
• 负责人: Xiaoping Du
• 金额: $ 39.75万
• 依托单位: UNIVERSITY OF ILLINOIS AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-12-15 至 2015-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8186790
• 关键词: Adhesions; Adhesives; Agonist; Binding; Blood; Blood Platelets; Blood Vessels; Bone Marrow Transplantation; Calpain; Cause of Death; Chinese Hamster Ovary Cell; Clot retraction; Coagulation Process; Cytoplasmic Granules; Cytoplasmic Tail; Cytoskeleton; Data; Development; Disease; Extracellular Matrix; Family; Fibrin; Fibrinogen; Funding; GTP-Binding Proteins; Goals; In Vitro; Injury; Integrin Signaling Pathway; Integrin alpha Chains; Integrins; Ligand Binding; Ligands; Ligation; Mediating; Membrane; Modeling; Monomeric GTP-Binding Proteins; Movement; Myocardial Infarction; Names; Peptides; Pharmaceutical Preparations; Platelet Activation; Play; Protein Subunits; Publishing; Regulation; Role; SRC gene; Science; Side; Signal Pathway; Signal Transduction; Site; Stroke; Surface; Techniques; Testing; Thrombosis; Thrombus; Transgenic Mice; Tyrosine Phosphorylation; United States; Wound Healing; adhesion receptor; base; extracellular; fighting; in vivo; inhibitor/antagonist; mutant; response; rho guanine nucleotide exchange factor p115; seal; src-Family Kinases; wound

DESCRIPTION (provided by applicant): The platelet integrin, aIIb¿3, interacts with adhesive ligands such as fibrinogen and fibrin, and mediates platelet adhesion and aggregation. Integrin aIIb¿3 thus plays a critical role in the development of thrombotic diseases such as heart attack and stroke. Integrin aIIb¿3 also transmits signals bidirectionally: The ligand binding function of integrin aIIb¿3 is activated by signals from within platelets ("inside-out" signaling). Ligand binding to aIIb¿3 induces "outside-in" signals, which transmit into platelets and elicit cellular response critical in amplifying and stabilizing thrombi, in wound healing and the re-canalization of blood vessels. The goal of this application is to understand the mechanisms of integrin outside-in signaling. It is known that integrin outside-in signaling requires the activation of the Src family of kinases and regulation of small GTPases such as RhoA. However, the upstream mechanisms that initiate integrin-mediated Src activation have been unknown. Furthermore, integrin aIIb¿3 initially mediates platelet spreading, but later induces platelet-mediated clot retraction, which requires the movement of platelet membranes in the opposite direction. It has been unclear how integrin aIIb¿3 switches the direction of membrane movements. During the current funding period, we have made the following discoveries: (1) The G protein subunit, Ga13, directly interacts with the cytoplasmic domain of the integrin ¿3 subunit, and this interaction is required for the integrin-dependent c-Src activation, RhoA inhibition and platelet spreading (Gong et al, Science, 2010). (2) a calpain cleavage of ¿3 disrupts the c- Src-dependent RhoA inhibition, inducing RhoA-dependent clot retraction. (3) the small G proteins, RhoA and Rac1, are both important in integrin-dependent retractile signaling and clot retraction. Based on data from these studies, we propose following hypotheses: (1) integrin aIIb¿3 outside-in signaling is mediated by the direct binding of Ga13 to the cytoplasmic domain of ¿3. (2) Ga13 transmits integrin signal to activate c-Src and induce c-Src-dependent inhibition of RhoA, resulting in inhibition of platelet retraction and promotion of platelet spreading. (3) Calpain cleavage of ¿3 serves as a mechanism that down regulates Ga13 signaling by inactivating the Ga13/c-Src-dependent signaling pathway that inhibits RhoA. This facilitates integrin-dependent RhoA activation and stimulates clot retraction. To test these hypotheses, we propose the following specific aims: Aim 1, to determine the role of Ga13 binding to ¿3 in mediating integrin outside-in signaling. In this aim, we will determine the importance of Ga13, particularly the role of Ga13-¿3 interaction, in various integrin signaling pathways, and determine how Ga13-¿3 interaction is regulated and how this interaction activates c- Src. Aim 2, to investigate the mechanism of Ga13 and integrin-dependent dynamic regulation of RhoA in controlling platelet spreading and retraction. In this aim, we will investigate the role of Ga13-¿3 interaction in inhibiting RhoA and stimulating p190RhoGAP, competitive inhibition of Ga13-p115RhoGEF by ¿3, and the mechanisms of calpain-induced late RhoA activation and retractile signaling. PUBLIC HEALTH RELEVANCE: Thrombotic diseases such as heart attack and stroke are a leading cause of death in United States. Integrins are a family of adhesion receptors expressed on the surface of blood platelets that are critical to thrombosis ("clotting" in blood vessels). Following integrin mediated platelet adhesion to the site of vascular injury, there is a signal sent by integrins from the outside to the inside of platelets (outside-in signaling) that induces platelet responses including platelet spreading, clot retraction and platelet release of pro-thrombotic factors that greatly amplify thrombus formation. In the previous funding period, we discovered that integrin outside-in signaling requires the binding of G protein, Ga13 to the inner side of integrin ¿3 subunit (published in Science, 2010). The binding of Ga13 to ¿3 sends a signal inhibiting another G protein named RhoA. The function of RhoA is to transmit a signal causing platelet contraction. By inhibiting contraction, integrins signaling allows platelets to spread out helping sealing the wound. This competitive renewal application is aimed at further investigating how the binding of Ga13 to integrins transmits signals that induce platelet responses and thrombosis. This study will help in the development of new concepts and new drugs in fighting thrombotic diseases.

描述（由申请人提供）：血小板整合素，aiib¿ 3，与纤维蛋白原和纤维蛋白等粘合剂配体相互作用，以及介导血小板粘附和聚集。 3因此，在血栓形成疾病（例如心脏病发作和中风）中起着至关重要的作用。 3还双向传输信号：整联蛋白AIIB课的配体结合函数3通过血小板内的信号激活（“内部”信号传导）。 3诱导“外部”信号，该信号在放大和稳定血栓愈合中的血小板响应至关重要，而血管的量很大。外部信号表明激酶的SRC家族和Rhoa的调节3最初介导血小板扩散，但后来诱导血小板介导的凝块回缩，这需要血小板膜在相反的方向上移动。 3切换膜运动的方向，在当前的融资期间，我们已经发现了以下发现：（1）与整联蛋白的细胞质结构界键相互作用。 3亚基，这是整联蛋白依赖性C-SRC激活所必需的（2）cal 3破坏了THOA抑制作用，诱导Rhoa依赖性凝块回缩。 3外部信号传导是由GA13与»细胞质结构域的直接结合介导的。 3。（2）GA13将整联蛋白信号激活c-Src依赖性抑制作用。 3用作抑制ga13/c-src的信号传导的机制，抑制了依赖于整联蛋白的RhoA激活和刺激凝块回收的机制。确定GA13与»结合的作用3在此目标中介导整联蛋白的信号传导中，我们将确定GA13的重要性。 3相互作用，在各种整联蛋白信号通路中，并确定GA13-€的方式3相互作用是常规的，这种相互作用如何激活RHOA的离子在控制血小板扩散和缩回中。 3抑制RhoA和刺激P190RHOGAP的相互作用3，以及Calpain诱导的RhoA晚期激活和伸缩信号的机制。 公共卫生相关性：诸如中风之类的血栓性疾病是在血小板上表达的主要原因。外部的信号是诱导血小板散布，凝块缩回和血小板释放的血小板释放的信号（外部信号传导），这些因子的血小板释放在先前的资助周期G蛋白质中会散发出良好的促进性。 ，GA13到整联蛋白的内侧» 3个亚基（科学上的发表，2010年）。 3发出抑制一个名为RhoA的G蛋白的信号。反应和血栓形成。