Cadherin Regulation of Epithelial Barriers

钙粘蛋白对上皮屏障的调节

• 批准号: 8706916
• 负责人: BARRY M. GUMBINER
• 金额: $ 39.5万
• 依托单位: UNIVERSITY OF VIRGINIA
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-08-01 至 2017-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8706916
• 关键词: Actomyosin; Acute; Adhesions; Adhesives; Affinity; Allergens; Animal Model; Antibodies; Binding; Binding Sites; Biophysics; Cadherins; Calcium Binding; Cell Culture Techniques; Cell Line; Cell surface; Cells; Collaborations; Complex; Cytoskeleton; Dendritic Cells; Disease; Dissociation; E-Cadherin; Endothelium; Enzymes; Epithelial; Epithelium; Epitopes; Event; Goals; Illinois; Immune; Inflammation; Inflammation Mediators; Inflammatory; Intercellular Junctions; Intestines; Kinetics; Laboratories; Left; Lung; Lung Inflammation; Maintenance; Measurement; Microbe; Microtubules; Modeling; Molecular; Molecular Conformation; Monoclonal Antibodies; Mutation; N-terminal; Organ; Permeability; Phosphorylation; Physiological; Physiological Processes; Play; Process; Property; Proteins; Regulation; Role; Site; Structure; Surface; System; Therapeutic; Tissues; base; extracellular; in vivo; insight; migration; neutrophil; novel; novel strategies; public health relevance; response; seal

DESCRIPTION (provided by applicant): Epithelia form dynamic barriers between tissue compartments and modulate their permeability properties during physiological and pathophysiological processes. Inappropriate maintenance or control of epithelial paracellular barriers contributes to many disease processes, especially inflammation. Cadherins are particularly important for the regulation of paracellular permeability in epithelia as well as endothelia, and the cadherin-catenin system has been directly implicated in inflammatory disease. New findings from my laboratory provide important new insights into the mechanisms of E-cadherin regulation in epithelia as well as novel approaches to manipulate cadherin activity experimentally. We discovered a mechanism that controls cadherin conformation and adhesive activity at the cell surface independent of any changes in levels of expression or amounts of associated catenins. This included the discovery of E-cadherin activating monoclonal antibodies (mAbs) and a role for p120-catenin phosphorylation and microtubules in the control of E-cadherin activity state. I hypothesize that regulation of the adhesive homophilic bond itself is a key event in cell junction regulation, allowing the catenin-associated cytoskeleton to pull the junctions apart once the adhesive bond is broken, and that this mechanism plays a role in barrier regulation in both epithelia and endothelia during inflammatory processes. The overall goals of this proposal are to determine the roles of cadherin cell surface regulation in the contro of epithelial barrier function and to understand in greater depth the mechanisms by which cadherins are regulated at the cell surface. The specific aims are: A. Investigate the role of E-cadherin activity state in the regulation of the epithelial barrier during inflammatory processes i cell culture and animal models. Regulation by soluble inflammatory mediators, control of neutrophil transmigration, and regulation of the interactions of dendritic cells with epithelium wil be investigated. The role of E-cadherin regulation in vivo will be assessed in an animal model of lung inflammation. B. Determine the mechanisms regulating cadherin adhesive binding activity at the cell surface. Analysis of the structure of epitopes recognized by activity associated mAbs, the biophysical changes in the properties of the homophilic adhesive bond, and the role of cadherin oligomerization, clustering, and localization in adhesion sites, all will provide insights into how the extracellular adhesive domain is regulated. C. Investigate cytoplasmic mechanisms of E-cadherin activation and cell surface regulation. Determining the enzymes that control p120-catenin phosphorylation, the role of microtubules in controlling E-cadherin activity and p120-catenin phosphorylation, and the effectors through which p120-catenin phosphorylation controls adhesion activity, will provide insights into the molecular mechanisms underlying adhesion regulation. Findings from these studies may allow us to develop cadherin- based approaches, or even therapeutics, to manipulate barrier function during inflammation and related disease processes.

描述（由申请人提供）：上皮细胞在组织区室之间形成动态屏障，并在生理和病理生理过程中调节其渗透性。上皮细胞旁屏障的不适当维护或控制会导致许多疾病过程，尤其是炎症。钙粘蛋白对于上皮和内皮细胞的细胞旁通透性的调节特别重要，并且钙粘蛋白-连环蛋白系统与炎症疾病直​​接相关。我实验室的新发现为上皮细胞中 E-钙粘蛋白调节机制提供了重要的新见解，并提供了通过实验操纵钙粘蛋白活性的新方法。我们发现了一种控制细胞表面钙粘蛋白构象和粘附活性的机制，与相关连环蛋白表达水平或数量的任何变化无关。其中包括发现 E-钙粘蛋白激活单克隆抗体 (mAb) 以及 p120-连环蛋白磷酸化和微管在控制 E-钙粘蛋白活性状态中的作用。我假设粘附同亲键本身的调节是细胞连接调节中的关键事件，一旦粘附键被破坏，连环蛋白相关的细胞骨架就可以将连接拉开，并且这种机制在两个上皮细胞的屏障调节中都发挥着作用和炎症过程中的内皮细胞。该提案的总体目标是确定钙粘蛋白细胞表面调节在控制上皮屏障功能中的作用，并更深入地了解钙粘蛋白在细胞表面的调节机制。具体目标是： A. 研究细胞培养和动物模型炎症过程中 E-钙粘蛋白活性状态在上皮屏障调节中的作用。将研究可溶性炎症介质的调节、中性粒细胞迁移的控制以及树突细胞与上皮细胞相互作用的调节。 E-钙粘蛋白的体内调节作用将在肺部炎症动物模型中进行评估。 B. 确定调节细胞表面钙粘蛋白粘附结合活性的机制。分析与活性相关的 mAb 识别的表位结构、同亲粘附键特性的生物物理变化，以及粘附位点中钙粘蛋白寡聚、聚类和定位的作用，所有这些都将提供见解 研究细胞外粘附结构域是如何调节的。 C. 研究 E-钙粘蛋白激活和细胞表面调节的细胞质机制。确定控制 p120-连环蛋白磷酸化的酶、微管在控制 E-钙粘蛋白活性和 p120-连环蛋白磷酸化中的作用，以及 p120-连环蛋白磷酸化控制粘附活性的效应器，将有助于深入了解粘附调节的分子机制。这些研究的结果可能使我们能够开发基于钙粘蛋白的方法，甚至是治疗方法，以在炎症和相关疾病过程中操纵屏障功能。