Ras/Rap Signaling and Endothelial Morphogenesis

内皮细胞在发育过程中通过细胞-细胞相互作用相互交流， 船舶.一个重要的结构是粘附连接，它连接个体的肌动蛋白细胞骨架。 细胞通过钙粘蛋白的相互作用。虽然内皮粘附连接在血管内皮细胞中很重要， 然而，令人惊讶的是，人们对它们的形成和调控知之甚少。在成熟的血管中， 需要连接以形成控制流体和大分子在 血液和组织部位。第二信使甘油二酯（DAG）促进血管通透性， 破坏内皮屏障功能，佛波醇酯模拟DAG活性。本提案的目的是 确定我们最近发现的一种新的内皮DAG/佛波醇酯受体RasGRPS如何影响 粘附连接发育，并影响成人血管中的血管通透性。RasGRPS 激活小GTP酶Ras和/或Rap，我们的初步数据指出RasGRPS的关键作用 如果发育中的血管对DAG/佛波酯的反应，通过类似于渗透性的机制， 成年人的反应。因此，我们假设RasGRPS介导的Ras/Rap信号转导是一个关键的 在细胞信号传导的情况下， 通过DAG是主导的。我们通过外源性给予佛波醇酯进行实验， 我们认为这种情况在以升高的DAG水平为特征的疾病状态中重演， 例如糖尿病。我们认为，成人和发展中国家糖尿病的血管并发症 胎儿可以通过阻断RasGRPS活性来改善。这些假设将在三个测试 目的是充分表征发育中血管和内皮细胞的RasGRPS依赖性反应 DAG/佛波酯在细胞和分子水平，并检查遗传的后果， RasGRPS对内皮屏障功能和糖尿病血管并发症的影响 胚胎和成年人

Endothelial cells communicate with each other via cell-cell interactions during development and in stable vessels. One important structure is the adherens junction, which links the actin cytoskeleton of individual cells via cadherin-based interactions. Although endothelial adherens junctions are important in vascular development, surprisingly little is known about their formation and regulation. In mature vessels, adherens junctions are required to form a barrier that controls the movement of fluids and macromolecules between blood and tissue sites. The second messenger diacylglycerol (DAG) promotes vascular permeability and disrupts endothelial barrier function, and phorbol esters mimic DAG activity. The goal of this proposal is to determine how a novel endothelial DAG/phorbol ester receptor we recently identified, RasGRPS, affects adherens junctions developmentally and impacts on .vessel permeability in adult vessels. RasGRPS activates the small GTPases Ras and/or Rap, and our preliminary data point to a crucial role for RasGRPS irf the response of developing vessels to DAG/phorbol esters, via a mechanism similar to the permeability response of adult vessels. Thus we hypothesize that RasGRPS-mediated Ras/Rap signaling is a critical endothelial control point for the regulation of adherens junctions, in situations where cellular signaling through DAG is dominant. We set this up experimentally by exogenous administration of phorbol esters, but we posit that this scenario is recapitulated in disease states that are characterized by elevated DAG levels, such as diabetes. We propose that the vascular complications of diabetes in both adults and developing fetuses can be ameliorated by blockade of RasGRPS activity. These hypotheses will be tested in three aims that fully characterize the RasGRPS-dependent response of developing vessels and endothelial cells to DAG/phorbol esters at the cellular and molecular levels, and examine the consequences of genetic manipulation of RasGRPS on endothelial barrier function and the vascular complications of diabetes in embryos and adults.