Cytoskeletal-Pathogen Interactions in Shigella Infection

志贺氏菌感染中的细胞骨架-病原体相互作用

• 批准号: 6630241
• 负责人: Scott B Snapper
• 金额: $ 14.81万
• 依托单位: MASSACHUSETTS GENERAL HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-09-15 至 2007-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6630241
• 关键词: SDS polyacrylamide gel electrophoresis; Shigella; actins; bacteria infection mechanism; bacterial cytopathogenic effect; bacterial proteins; biological signal transduction; cytoskeleton; genetically modified animals; guanine nucleotide binding protein; guanosinetriphosphatases; host organism interaction; intestinal mucosa; laboratory mouse; membrane proteins; pathologic process; protein structure function; site directed mutagenesis

DESCRIPTION (provided by applicant): Shigella are gram negative enteric pathogens that cause severe diarrheal disease and have been classified as a Category B Biological Agent. Shigella pathogenesis requires bacterial invasion of the colonic epithelium and bacterial spread through the colonic mucosa. Shigella entry into epithelial cells is mediated by effector molecules, secreted through a type III secretion apparatus, that activate Rho family GTPase signaling pathways to induce the formation of cell surface projections and membrane ruffles that engulf the bacteria by macropinocytosis. Both Cdc42 and Rac have been implicated as having a role in the Shigella entry process. Cdc42 is known to activate Rac; it is not clear whether Cdc42 involvement in Shigella entry is mediated exclusively via this link. Moreover, the downstream effectors of Cdc42 and/or Rac activation during Shigella entry are unknown. We have recently confirmed that the major Shigella pathway is Cdc42-dependent. However, we have also demonstrated the existence of a novel Cdc42-independent invasion pathway. Furthermore we have shown that the only known downstream effector of Cdc42 that activates the actin cytoskeleton, N-WASP, is not involved in Shigella entry. Once in the cytoplasm, Shigella moves by active assembly of an actin tail. Actin tail formation is mediated by the Shigella outer membrane protein IcsA, which binds and activates N-WASP. Activated N-WASP stimulates Arp2/3 complex-mediated actin assembly. The molecular mechanism by which IcsA binds and activates N-WASP is poorly understood. Our goals in this proposal are to: 1. Define the specific roles of Cdc42 and Rac in Shigella entry; 2. Identify and characterize the downstream effectors of Rho family activation during Shigella entry; and, 3. Elucidate the mechanism(s) by which Shigella IcsA activates N-WASP and determine whether this mechanism mimics Cdc42 activation of N-WASP These studies will define the specific cellular signaling pathways required for Shigella entry and actin tail formation and will identify downstream pathways of Rho family activation.

描述（由申请人提供）：志贺氏菌是革兰氏阴性肠道病原体，可引起严重腹泻病，已被归类为 B 类生物制剂。志贺氏菌的发病机制需要细菌侵入结肠上皮并通过结肠粘膜传播。志贺氏菌进入上皮细胞是由效应分子介导的，通过 III 型分泌装置分泌，激活 Rho 家族 GTP 酶信号通路，诱导细胞表面突起和膜皱褶的形成，通过巨胞饮作用吞噬细菌。 Cdc42 和 Rac 都被认为在志贺氏菌进入过程中发挥了作用。已知 Cdc42 可以激活 Rac；目前尚不清楚 Cdc42 参与志贺氏菌进入是否仅通过此链接介导。此外，志贺氏菌进入过程中 Cdc42 和/或 Rac 激活的下游效应器尚不清楚。我们最近证实志贺氏菌的主要途径是 Cdc42 依赖性的。然而，我们还证明了一种新的不依赖于 Cdc42 的入侵途径的存在。此外，我们还表明，激活肌动蛋白细胞骨架的唯一已知的 Cdc42 下游效应子 N-WASP 不参与志贺氏菌的进入。一旦进入细胞质，志贺氏菌就会通过肌动蛋白尾部的主动组装来移动。肌动蛋白尾部的形成由志贺氏菌外膜蛋白 IcsA 介导，IcsA 结合并激活 N-WASP。激活的 N-WASP 刺激 Arp2/3 复合物介导的肌动蛋白组装。 IcsA 结合并激活 N-WASP 的分子机制尚不清楚。 我们在此提案中的目标是： 1.明确Cdc42和Rac在志贺氏菌条目中的具体作用； 2. 鉴定和表征志贺氏菌进入过程中Rho家族激活的下游效应子；和， 3. 阐明志贺菌 IcsA 激活 N-WASP 的机制，并确定该机制是否模仿 Cdc42 激活 N-WASP 这些研究将定义志贺氏菌进入和肌动蛋白尾部形成所需的特定细胞信号传导途径，并将确定 Rho 家族激活的下游途径。