Microtubule-associated Rac

微管相关Rac

• 批准号: 8474833
• 负责人: Anna Birukova
• 金额: $ 36.76万
• 依托单位: UNIVERSITY OF CHICAGO
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-08-01 至 2015-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8474833
• 关键词: Actins; Actomyosin; Acute; Acute Lung Injury; Adaptor Signaling Protein; Adenomatous Polyposis Coli; Adherens Junction; Adult Respiratory Distress Syndrome; Agonist; Animal Model; Attenuated; Automobile Driving; Binding; Biochemical; Biological Preservation; Blood Vessels; Cell Adhesion; Cells; Complex; Complication; Cytoskeleton; Data; Development; Disease; Down-Regulation; Drug Design; Elements; Endothelial Cells; Endothelium; Equilibrium; Functional disorder; Future; Guanine Nucleotide Exchange Factors; Guanosine Triphosphate; Guanosine Triphosphate Phosphohydrolases; Hepatocyte Growth Factor; Human; In Vitro; Inflammatory; Injury; Intercellular Junctions; Kinesin; Knockout Mice; Lead; Link; Lung; Lung Inflammation; Lung diseases; Mechanics; Mediating; Mediation; Microtubule Alteration; Microtubule Stabilization; Microtubule-Associated Proteins; Microtubules; Modeling; Molecular; Molecular Analysis; Molecular Target; Monomeric GTP-Binding Proteins; Motor; Pathogenesis; Peripheral; Permeability; Pilot Projects; Play; Prevention; Prevention therapy; Property; Protective Agents; Proteins; Publishing; Pulmonary Edema; Regulation; Research Proposals; Role; Sepsis; Signal Pathway; Signal Transduction; Small Interfering RNA; Thrombin; Time; Transducers; Translational Research; Vascular Permeabilities; Ventilator-induced lung injury; base; effective therapy; in vivo; in vivo Model; injured; insight; lung injury; lung preservation; lung repair; monolayer; mortality; multidisciplinary; novel; protective effect; protein complex; restoration; rho; rho GTP-Binding Proteins; targeted delivery

DESCRIPTION (provided by applicant): Microtubule-associated Rac regulation and acute lung injury. Compromised lung endothelial cell (EC) permeability leads to pulmonary edema, a serious complication observed in various lung diseases and associated with high mortality. Small GTPases Rac and Rho play opposing roles in EC cytoskeletal remodeling and cell junction dynamics, the critical mechanisms of endothelial barrier regulation. Precise regulation of small GTPase activities in different subcellular compartments may be a key mechanism of Rac-dependent enhancement of peripheral actin, cell junction complexes and EC barrier protective potential. It is known that increased microtubule (MT) disassembly leads to EC barrier dysfunction. Our published studies show that edemagenic and pro-inflammatory agonists thrombin, TGF2 and TNF1 increase EC permeability via Rho-dependent alteration of MT dynamics leading to MT- dependent actomyosin contraction. In contrast, effects of barrier-protective agents on crosstalk between MT, cell adhesions, and actin cytoskeleton are not clear, nor the role of MT changes in Rac regulation is described. Our novel data show that MT-mediated signaling is directly involved in the EC barrier preservation by hepatocyte growth factor (HGF), a circulating molecule with proven protective effects in the models of acute ling injury (ALI). These results have led us to a hypothesis that microtubules may mediate barrier-protective effects of HGF and attenuate acute pulmonary EC dysfunction via MT-associated signaling complexes. We hypothesize that MT-associated adaptor protein APC (adenomatous polyposis coli) can bind and deliver a novel Rac-specific guanine nucleotide exchange factor Asef along MT to the cell periphery, where Asef switches its cytoskeletal localization from MT to actin and targets actin-associated Rac effector IGQAP1. This mechanism may lead to a novel paradigm of microtubule- dependent enhancement of peripheral actin cytoskeleton and cell-cell junctions via targeted delivery of Rac activator Asef, which promote EC barrier properties and lung repair mechanisms in ALI. Specific Aim #1 will examine the role of APC and Asef in the Rac GTPase activation and HGF-induced barrier enhancement in pulmonary endothelial cells. Specific Aim #2 will study the role of MT in APC/Asef activation and intracellular localization. Specific Aim #3 will examine HGF-induced APC/Asef interactions with IQGAP1 and study the role of APC/Asef/IQGAP1 complex in HGF-induced EC barrier enhancement. Specific Aim #4 will use siRNA-based Asef knockdown in vivo, Asef knockout mice, and rescue strategies to evaluate Asef role in the lung protective mechanisms against ALI. These studies will characterize novel barrier protective mechanisms and identify new protein targets for future therapies aimed at prevention of the pulmonary vascular barrier dysfunction associated with acute lung injury.

描述（由申请人提供）：微管相关的RAC调节和急性肺损伤。肺内皮细胞（EC）渗透率受损会导致肺水肿，这是一种严重的并发症，在各种肺部疾病中观察到，并且与高死亡率有关。小型GTPases RAC和RHO在EC细胞骨架重塑和细胞连接动力学中扮演着相反的角色，这是内皮屏障调节的关键机制。对不同亚细胞室中小的GTPase活性的精确调节可能是RAC依赖性增强外周肌动蛋白，细胞连接络合物和EC屏障保护潜力的关键机制。众所周知，增加的微管（MT）拆卸会导致EC屏障功能障碍。我们发表的研究表明，通过RHO依赖性改变MT动力学的变化，杂志和促炎激动剂凝血酶，TGF2和TNF1增加了EC的渗透性，从而导致MT依赖性肌动蛋白收缩。相反，屏障保护剂对MT，细胞粘连和肌动蛋白细胞骨架之间串扰的影响尚不清楚，也描述了MT变化在RAC调节中的作用。我们的新数据表明，MT介导的信号传导直接参与了肝细胞生长因子（HGF）的EC屏障，这是一种循环分子，在急性Ling损伤（ALI）模型中具有证实的保护作用。这些结果使我们提出了一个假设，即微管可能介导HGF的屏障保护作用，并通过MT相关的信号传导复合物减轻急性肺EC功能障碍。我们假设与MT相关的衔接蛋白APC（腺瘤息肉大肠杆菌）可以结合并提供新型的RAC特异性鸟嘌呤核苷酸核苷酸交换因子ASEF沿MT沿MT与细胞外围关系，在那里，ASEF将其ASEF切换为其细胞骨架的位置，从MT到Actins Actins acts acts acts acts acts acts acts-Actapiend-Racied Raceciped Raceciped Raceciped Racefor iGQAP1。这种机制可能导致通过针对RAC Activator的靶向递送ASEF的靶向递送，促进ALI中的EC屏障特性和肺修复机制，从而导致了周围依赖于微管依赖性的增强依赖于微管的范式。具体的目标＃1将检查APC和ASEF在RAC GTPase激活中的作用，以及HGF诱导的肺内皮细胞的屏障增强。具体目标＃2将研究MT在APC/ASEF激活和细胞内定位中的作用。特定的目标＃3将检查HGF诱导的APC/ASEF与IQGAP1的相互作用，并研究APC/ASEF/IQGAP1复合物在HGF诱导的EC屏障增强中的作用。特定的目标＃4将使用基于siRNA的ASEF敲除在体内，ASEF敲除小鼠和救援策略来评估ASEF在针对ALI的肺部保护机制中的作用。这些研究将表征新颖的屏障保护机制，并确定旨在预防与急性肺损伤相关的肺血管屏障功能障碍的未来疗法的新蛋白质靶标。