Calcium Dependent Signaling in Airway Epithelial Cells

描述(由申请人提供)：呼吸道上皮通过发出信号将吞噬细胞招募到感染部位来启动对吸入肺部病原体的防御。这一信号是由Toll样受体完成的，Toll样受体由保守的细菌成分激活，刺激MAPK和NF-β信号级联，诱导趋化因子如IL-8的表达，将PMN招募到呼吸道。为了使中性粒细胞跨越上皮屏障的紧密连接，必须对紧密连接和粘连连接的几种膜蛋白进行修饰。我们认为，由TLR2和TLR5启动的TLR信号通路激活了钙依赖的钙蛋白酶，该酶以上皮连接蛋白occludin和E-cadherin为靶标，促进中性粒细胞进入呼吸道。类似地，TLR4的内毒素激活可通过TACE等钙离子非依赖性的酶来裂解连接蛋白，从而促进PMN的迁移。这些相同的连接成分也是细菌毒力因子的靶标，细菌毒力因子是铜绿假单胞菌分泌的III型毒素，它进一步改变上皮屏障，促进细菌入侵。GTP酶抑制和ADP核糖化的联合作用，即铜绿假单胞菌毒素的活性，改变了对维持上皮细胞骨架及其屏障功能至关重要的连接蛋白的定位和功能。在提出的实验中，我们将描述促进PMN通过上皮屏障迁移的信号过程，以及细菌病原体如何利用相同的靶点。公共卫生相关性：呼吸道上皮细胞中的钙依赖信号肺炎，即肺部炎症细胞的聚集，是发病率和死亡率的主要原因。这个项目试图了解中性粒细胞被招募到呼吸道以应对细菌感染的基本过程。

DESCRIPTION (provided by applicant): The airway epithelium initiates defenses against inhaled pulmonary pathogens by signaling to recruit phagocytic cells to the site of infection. This signaling is accomplished by toll-like receptors that are activated by conserved bacterial components, stimulate MAPKs and NF-?B signaling cascades to elicit expression of chemokines such as IL-8 that recruit PMNs into the airway. For PMNs to transmigrate across the tight junctions of the epithelial barrier, several membrane spanning proteins of the tight and adherens junctions must be modified. We propose that TLR signaling initiated by TLR2 and possibly TLR5 activates the Ca2+ dependent protease calpain, which targets the epithelial junctional proteins occludin and E-cadherin to facilitate paracellular passage of PMNs into the airway. Similarly, LPS activation of TLR4 induces cleavage of junctional proteins by Ca2+-independent proteases, such as TACE to facilitate PMN transmigration. These same junctional components are also the targets of bacterial virulence factors, the type III secreted toxins of P. aeruginosa, which further modify the epithelial barrier to facilitate bacterial invasion. The combined effects of GTPase inhibition and ADP ribosylation, the activities of P. aeruginosa toxins, alters the localization and functions of junctional proteins critical for the maintenance of the epithelial cytoskeleton and its barrier function. In the experiments proposed we will characterize the signaling process that facilitates PMN migration through the epithelial barrier and how the same targets can be exploited by bacterial pathogens. PUBLIC HEALTH RELEVANCE: Calcium dependent signaling in airway epithelial cells Pneumonia, the accumulation of inflammatory cells in the lung, is a major cause of morbidity and mortality. This project seeks to understand the fundamental process through which polymorphonuclear leukocytes are recruited into the airway in response to bacterial infection.