Regulation of vascular permeability by thrombin mediated signaling pathways

这项提议的目标是了解G蛋白下游多个G蛋白通路的信号整合 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PAR1下游的蛋白质信号通路，凝血酶受体。PAR1 已知通过与多个G蛋白偶联来调节其在内皮中的复杂作用 包括GI、GQ和G12/13家族的成员，从而导致这些信号通路的激活 调节内皮屏障功能。G蛋白信号转导模型的数学基础 路径问题还没有被应用于这样的多路径集成问题。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 屏障通透性调节器。屏障功能障碍会导致浮肿，这是创伤和 炎症，但当不加以控制时，是许多疾病的毁灭性组成部分，如急性肺损伤 还有败血症。我们的目标是1)构建一个数学和计算模型来描述 PARs下游的G蛋白信号通路，将作为系统研究 调节内皮通透性的详细分子机制。2)通过以下方式确定机制 哪些单独的G蛋白对生理反应有贡献。实验室已经研制出 创新的工具，一次解偶联一个G蛋白途径，并确定对细胞反应的影响。 3)使用该模型帮助确定将成为这些治疗干预的新靶点的关键部位 精神错乱。利用模型的早期迭代，我们发现凝血酶和凝血酶受体 激活肽对不同类别的G蛋白信号通路有不同的调节作用。我们会 研究不同构象的PAR1向不同的G蛋白传递的假设，以及 差别化调节屏障功能。这将为寻找变构药物提供一个治疗窗口 作用于不同位点的PAR的调节物，而不是被拴住的配体。在整个授奖过程中，模特， 模拟和实验将迭代进行。长期目标是为实现 用于凝血酶的复杂信号过程的有效计算模型的产生 内皮屏障的调节。该模型将有助于确定将成为新目标的关键地点 对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.