Regulation of Platelet Shape Changes

描述（由申请人提供）：形状变化对于血小板防止出血的损伤控制功能至关重要，并且依赖于肌动蛋白丝响应于连接受体产生的信号的重塑和组装。血小板表面形状变化差异的细节可能对体内伴随止血和血栓形成反应的血小板分层具有重要影响，并可能为未来的抗血栓治疗提供靶点。我们以前定义了某些血小板信号级联，最终在净肌动蛋白组装/拆卸，并集中在钙，GTP酶，磷酸肌醇生产的形状变化的调节和这些代理商的下游目标的核心作用。我们还努力将肌动蛋白组装反应置于一个结构框架内，首先定义了肌动蛋白的组织和静息血小板的膜骨架，然后解剖了启动和维持形状变化反应所需的结构变化。该项目的总体目标是了解血小板如何使用大的lameprods和filepods传播，并将这些信息整合到介导血小板形状变化的信号通路和蛋白质中。这项建议有三个具体目标。在第一，我们强调的核心作用的rac，ralA和钙在lamellapodial大会。我们采取了一种新的结构方法，将选择性透化的细胞分散在我们可以引入外源分子的限定表面上。在第二个目标中，我们关注cdc 42在血小板丝状伪足形成中的作用。第三个目标详细介绍了一个基于珠的肌动蛋白组装系统，依赖于GTdR的活动，我们将使用它来开始重建层状和丝状伪足肌动蛋白组装的生化简化，然后重建的基本组成部分的目标。

DESCRIPTION (provided by applicant): Shape change is essential for the damage control functions of blood platelets that prevent bleeding and is dependent on the remodeling and assembly of actin filaments in response to signals generated by ligated receptors. Details in differences of platelet surface shape changes may have important ramifications for the layering of platelets accompanying hemostatic and thrombotic reactions in vivo and may provide targets for future antithrombotic therapies. We previously defined certain platelet signaling cascades that end in net actin assembly/disassembly and have focused on the central roles of calcium, GTPases, and phosphoinositide production in the regulation of shape change and on the downstream targets of these agents. We have also labored to put the actin assembly reactions within a structural framework by first having defined the organization of actin and the membrane skeleton of the resting platelet and then dissecting the changes in this architecture that are required to initiate and sustain the shape change reaction. The overall goal of this project is to understand how platelets spread using large lamellae and filopods and integrate this information into the signaling pathways and proteins that mediate platelet shape change. This proposal has three specific aims. In the first, we highlight the central roles of rac, ralA and calcium in lamellapodial assembly. We take a novel structural approach that combines selectively permeabilized cells spread on defined surfaces to which we can introduce exogenous molecules. In the second aim, we focus on the role of cdc42 in platelet filopodial elaboration. The third aim details a bead-based actin assembly system, dependent on GTPase activity, which we will use to begin to reconstruct lamellar and filopodial actin assembly with the goal of biochemical simplification and then reconstitution of the essential components.