G Protein Regulation of Endothelial Barrier Function

G 蛋白对内皮屏障功能的调节

基本信息

批准号：
7843673
负责人：
TATYANA A VOYNO-YASENETSKAYA
金额：
$ 40.87万
依托单位：
UNIVERSITY OF ILLINOIS AT CHICAGO
依托单位国家：
美国
项目类别：
财政年份：
资助国家：
美国
起止时间：
至
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/7843673
关键词：
A kinase anchoring protein Actin-Binding Protein Actins Address Adult Respiratory Distress Syndrome Aleurites Alveolar Binding Blood Vessels Catalytic Domain Cells Complex Cyclic AMP Cyclic AMP-Dependent Protein Kinases Cytoskeleton Cytoskeleton Alteration Data Dominant-Negative Mutation Down-Regulation Endothelial Cells F2R gene Feedback Floods Focal Adhesions G-Protein-Coupled Receptors G13 Protein GTP-Binding Proteins Goals Guanine Nucleotide Dissociation Inhibitors Heterotrimeric GTP-Binding Proteins Hour Inflammation Mediators Lead Life Link Liquid substance Lung MAP Kinase Kinase Kinase MAP3K1 gene MAP3K5 gene MEKKs Maintenance Mediating Microfilaments Mitogen-Activated Protein Kinases Modeling Molecular Morbidity - disease rate Mus NF-kappa B PAR-1 Receptor Pathway interactions Permeability Phosphorylation Phosphotransferases Physiological Play Principal Investigator Production Proteins Pulmonary Edema Recovery Regulation Role Signal Pathway Signal Transduction Signaling Molecule Stress Fibers Testing Thrombin Vascular Permeabilities base depolymerization fiber cell inhibitor/antagonist lung injury mortality novel polymerization prevent programs radixin protein response restoration rho rho guanine nucleotide exchange factor p115 rhoB p20 GDI vasodilator-stimulated phosphoprotein

项目摘要

Loss of lung vascular endothelial barrier integrity leads to increased vascular permeability and alveolar flooding, and contributes to the morbidity and mortality associated with ARDS. Thrombin activates G protein-coupled receptor PAR-1 in endothelial cells and induces actin stress fiber formation and the resultant increase in transendothelial permeability. The activation of the alpha subunit of the heterotrimeric G13 protein by thrombin in endothelial cells is critical for the activation of Rho proteins, and the subsequent increased endothelial permeability response. We have shown that Galpha-13 protein interacts with the actin-binding protein, radixin, which is critical for the assembly of focal adhesions and actin filaments. Thus, radixin may be an essential effector in signaling the Galpha-13-induced Rho activation via the Rho guanine nucleotide dissociation inhibitor, RhoGDI. In Project 3, we will address a new and potentially important signaling pathway by which Galpha-13-dependent Rho activation results in increased lung endothelial permeabihty. We will define the upstream regulation of Rho involving radixin and the signaling pathways mediating the increase in endothelial permeability. In response to inflammatory mediators such thrombin, the actin cytoskeleton alterations and the increased endothelial permeability are typically reversible within hours. The reversibility ot the response restores endothelial barrier integrity; however, its molecular and cellular bases are poorly understood. Protein kinase A, PKA, a kinase linked to enhancing endothelial barrier function is usually activated by cyclic AMP. We have discovered that both thrombin and Galpha-13 can stimulate PKA via two novel mechanisms that do not require cAMP. One mechanism is dependent on the interaction of Galpha-13 with radixin, which activates the reversal of the permeability response. Another mechanism is dependent on the stimulation of the NF-kappaB signaling pathway via mitogen-activated protein kinases. The end result is that PKA may phosphorylate Galpha-13, and thereby inhibit Rho activation. We will address the concept that this is a key mechanism for the down-regulation of Galpha-13 activity and the reversal of the permeability response. Another possibility to be addressed is that Galpha-13 induces the phosphorylation of vasodilator-stimulated phosphoprotein, VASP, which can prevent actin polymerization and thus re-establish the endothelial barrier. Project 3 will test the general hypothesis that signaling complexes formed by PAR-1 activation of Galpha-13 signal the loss of endothelial barrier function and sequentially activate signals that lead to endothelial barrier recovery, and the reversal of the permeability-increase. We believe that the proposed studies will generate novel information elucidating the regulation of lung vascular permeability. Moreover, with the unraveling of the signals regulating the reversal of the increase in lung vascular permeability, it will be possible to identify novel targets tor therapies whereby the inappropriate increase in endothelial permeability can be controlled reducing the vascular "leak" associated with ARDS.

肺血管内皮屏障完整性的丧失会导致血管渗透性增加和肺泡洪水，并有助于与ARDS相关的发病率和死亡率。凝血酶在内皮细胞中激活G蛋白偶联受体PAR-1，并诱导肌动蛋白应激纤维的形成以及产生的跨内皮渗透性增加。凝血酶在内皮细胞中通过凝血酶的α亚基的激活对 Rho蛋白的激活以及随后增加的内皮渗透性反应。我们已经表明，galpha-13蛋白与肌动蛋白结合蛋白Radixin相互作用，这对于聚焦粘连和肌动蛋白丝的组装至关重要。因此，radixin可能是通过Rho Guanine向Galpha-13诱导的RHO激活发出信号的重要效应。核苷酸解离抑制剂Rhogdi。在项目3中，我们将解决一种新的且潜在的重要信号传导途径，通过该途径Galpha-13依赖性RHO激活导致肺部内皮通透性增加。我们将定义涉及radixin的Rho的上游调节以及介导内皮通透性增加的信号通路。响应炎性介质，例如凝血酶，肌动蛋白细胞骨架的改变和增加内皮渗透性通常在几个小时内可逆。响应的可逆性恢复了内皮屏障完整性；然而，其分子和细胞碱基的理解很少。蛋白激酶A，PKA，一种与增强内皮屏障功能相关的激酶通常通过环状AMP激活。我们已经发现，凝血酶和Galpha-13都可以通过两种不需要扎营的新型机制刺激PKA。一种机制是取决于Galpha-13与radixin的相互作用，这激活了渗透率响应的逆转。另一种机制取决于通过有丝分裂原激活的蛋白激酶刺激NF-kappab信号通路。最终结果是PKA可能会磷酸化Galpha-13，从而抑制RHO激活。我们将解决一个概念这是下调Galpha-13活性和渗透性反应的逆转的关键机制。要解决的另一种可能性是Galpha-13诱导血管扩张剂刺激的磷酸蛋白VASP的磷酸化，这可以防止肌动蛋白聚合并因此重新建立内皮障碍。项目3将检验一个总体假设，即通过Galpha-13的PAR-1激活形成的信号传导复合物信号信号信号是内皮屏障功能的丧失，并依次激活信号，从而导致内皮屏障恢复，并逆转渗透性。我们认为，拟议的研究将产生新的信息，以阐明肺血管通透性的调节。此外，随着信号调节肺血管通透性增加的逆转，可以鉴定出新的靶标疗法的新靶标，从而可以控制内皮通透性的不适当增加，从而减少与ARDS相关的血管“泄漏”。