Regulation of vascular permeability by thrombin mediated signaling pathways

该提案的目标是了解多个G蛋白通路下游的信号整合， 单一受体我们建议建立多个可计算的“信令模块”，它们代表G 多重偶联受体（凝血酶受体PAR 1）下游的蛋白质信号传导途径。PAR1 已知通过其与多种G蛋白偶联的能力介导其在内皮中的复杂作用 包括Gi、Gq和G12/13家族的成员，从而导致这些信号传导途径的激活， 调节内皮屏障功能。G蛋白信号建模的数学基础 pathways尚未应用于这样的多途径整合问题。内皮细胞（EC） 在血液和间质组织之间形成动态调节的屏障;凝血酶是最有效的 屏障渗透性的调节剂。屏障功能障碍导致水肿，这是创伤的正常后果， 炎症，但当不受控制时，是许多疾病（如急性肺损伤）的破坏性组成部分 和败血症我们的目标是：1）构建一个数学和计算模型，描述多个 PAR下游的G蛋白信号通路，将作为系统研究 详细的调节内皮通透性的分子机制。2)确定机制， 哪些G蛋白参与了生理反应。该实验室开发了 创新的工具，解偶联一个G蛋白通路的时间，并确定对细胞反应的影响。 3)使用该模型来帮助确定关键部位，这些部位将成为治疗干预的新靶点。 紊乱使用模型的早期迭代，我们发现凝血酶和凝血酶受体 活化肽差异调节不同类别的G蛋白信号传导途径。我们将 研究PAR 1的不同构象运输到不同G蛋白的假设，以及 差异调节屏障功能。这将提供一个治疗窗口，以寻找变构 PAR的调节剂，其在与系链配体不同的位点起作用。在整个拨款过程中， 将反复进行模拟和实验。长期目标是朝着 凝血酶中使用的复杂信号传导过程的验证计算模型的产生 调节内皮屏障。该模型将有助于确定关键网站，将是新的目标， 这些疾病的治疗干预。

The goal of this proposal is to understand signal integration of multiple G protein pathways downstream of a single receptor. We propose to build up multiple computable "signaling modules", which represent G protein signalingpathways downstream of a multiply coupled receptor, PAR1, the thrombin receptor. PAR1 is known to mediate its complex actions in endothelium via its ability to couple to multiple Gproteins including members of Gi, Gq and G12/13 families thus leading to activation of these signaling pathways to modulate endothelial barrier function. The mathematical ground work laid in modeling of G protein signaling pathways has not yet been applied to such a multiple pathway integration problem. Endothelial cells (EC) form a dynamicallyregulated barrier between blood and interstitial tissues; thrombin is the most potent regulator of barrierpermeability. Barrier dysfunctionleads to edema, a normal consequence of wounding and inflammation, but when uncontrolled is a devastating component of many diseases such as acute lung injury and sepsis. Our aims are to 1) Construct a mathematical and computational model that describes the multiple G protein signaling pathways downstream of PARs that will serve as a tool to systematically investigate the detailed molecular mechanisms that regulate endothelial permeability. 2) Determine the mechanisms by which individualG proteins contribute to the physiological response. The laboratory has developed innovative tools to uncouple one G protein pathway at a time and determine the effects on cellular responses. 3) Use the model to help to identify key sites that would be novel targets for therapeutic intervention in these disorders. Using an early iteration of the model, we showed that thrombin and the thrombin receptor activating peptide differentially regulate different classes of G protein signaling pathways. We will investigate the hypothesis that different conformations of PAR1 traffic to different G proteins, and differentially regulate barrier function. This would provide a therapeutic window to search for allosteric modulators of PARs that work at different sites than the tethered ligand. Throughout the grant, the modeling, simulations and experimentation will be conducted iteratively. The long term goals are to build toward production of a validated computational model of the complex signaling processes used in thrombin regulation of the endothelial barrier. The model will help to identify key sites that would be novel targets for therapeutic intervention in these disorders.