Protease-activated receptors in embryonic development

描述(申请人提供)：在成人中，蛋白酶激活的受体(PARs)对凝血酶作出反应，以帮助协调参与止血、炎症和修复的细胞反应。在之前的项目期间，我们展示了PARs在胚胎发育中也扮演着明显而重要的角色。事实上，内皮细胞中的PAR1信号对于小鼠胚胎中发育中的血管的正确重塑和/或完整性是必需的。PAR1在血管形成过程中监控哪些生化和生理过程？哪些内皮细胞对PAR1激活的反应对血管的正常发育很重要？我们假设凝血级联和PARS一起提供了一个系统，用于监测和调节发育中的血管的形成、重塑和完整性。为了验证这一假设，我们首先要确定PARs在小鼠胚胎发育过程中是否感知凝血酶，以及PARs的激活是否解释了凝血因子在这一背景下的作用。具体问题包括a)PARS表型缺乏症是否合并组织因子缺乏？B)组织因子在血管内或血管周围的表达对于支持胚胎发育是否必要且足够？和c)携带凝血酶原突变的胚胎能否通过PAR1的互补突变来挽救？接下来，我们将通过去除小鼠胚胎内皮细胞中特定的G蛋白通路的功能，来确定哪些信号通路对血管发育过程中内皮细胞中PAR1和其他GPCRs的功能起重要作用。我们期望这些研究能够阐明凝血级联反应和PARS的新作用，并为控制血管发育的机制以及其他环境下的新血管形成提供新的见解。

DESCRIPTION (provided by applicant): In the adult, protease-activated receptors (PARs) respond to coagulation proteases to help orchestrate cellular responses involved in hemostasis, inflammation and repair. In the previous project period, we showed that PARs also play distinct and important roles in embryonic development. Indeed, PAR1 signaling in endothelial cells is required for proper remodeling and/or integrity of developing blood vessels in mouse embryos. What biochemical and physiological processes does PAR1 monitor during blood vessel formation? Which endothelial cell responses to PAR1 activation are important for proper vessel development? We hypothesize that the coagulation cascade and PARs together provide a system for monitoring and regulating the formation, remodeling, and integrity of developing blood vessels. Toward testing this hypothesis we shall first determine whether PARs sense coagulation proteases during mouse embryonic development, and whether activation of PARs account for the roles of coagulation factors in this context. Specific questions include a) Does combined deficiency of PARs phenocopy tissue factor deficiency? b) Is tissue factor expression in or around blood vessels necessary and sufficient to support embryonic development? and c) Can embryos bearing prothrombin mutations be rescued by complementary mutations in PAR1? We shall next determine which signaling pathways are important for the function of PAR1 and other GPCRs in endothelial cells during vascular development by ablating the function of specific G protein pathways in endothelial cells in the mouse embryo. We expect these studies to illuminate a novel role for the coagulation cascade and PARs and to provide new insights regarding the mechanisms governing vascular development and perhaps new blood vessel formation in other settings.