BACTERIAL INVASION OF EUKARYOTIC TISSUES

真核组织的细菌入侵

• 批准号: 2568902
• 负责人: D J KOPECKO
• 金额: --
• 依托单位国家: 美国
• 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/2568902
• 关键词: bacterial cytopathogenic effect; bacterial genetics; confocal scanning microscopy; enteric bacteria; eukaryote; fluorescence microscopy; gastrointestinal epithelium; gastrointestinal infection; messenger RNA; molecular cloning; molecular pathology; pathologic process; prokaryote; receptor binding; tissue /cell culture; transmission electron microscopy; video microscopy; virulence

Many bacterial pathogens of the intestinal tract have as an early and necessary step in initiation of disease the ability to penetrate gut epithelial cells. My lab's work in the early 1980's established that Shigella's genetic machinery to trigger invasion is encoded on a large virulence-associated plasmid. More recent studies have shown that the invasion ability of other enteric bacteria is chromosomally encoded. This project is aimed at understanding the prokaryotic and eukaryotic requirements for bacterial internalization, with the ultimate aim being a thorough molecular definition of the events involved in bacterial invasion of eukaryotic tissues. Current experimental approaches involve the use of assays measuring bacterial entry into cultured lines of human epithelial cells of various tissue origins. Biochemical inhibitors of prokaryotic structure/function or of eukaryotic cell processes are employed in these tissue culture invasion assays to examine the requirements for bacterial uptake. Direct visualization of bacterial entry is measured via transmission electron microscopy, video microscopy, fluorescent microscopy, and confocal microscopy. In addition to the mechanistic approaches described above, genetic techniques are employed to clone the responsible bacterial genes. Eukaryotic receptors for bacterial ligands and specific eukaryotic cell responses to bacterial invasion are measured via inhibitor competition assays, ligand binding assays, and mRNA analyses of infected eukaryotic cells. The information gained from each of these approaches is integrated to provide a mechanistic understanding of bacterial entry for each pathway studied.

许多肠道细菌病原体在早期和 疾病发生的必要步骤——穿透肠道的能力 上皮细胞。 我的实验室在 1980 年代初的工作证实了这一点 志贺氏菌触发入侵的遗传机制被编码在一个大的基因上 毒力相关质粒。最近的研究表明， 其他肠道细菌的侵袭能力是由染色体编码的。这 该项目旨在了解原核生物和真核生物 细菌内化的要求，最终目标是 对细菌相关事件的全面分子定义 侵入真核组织。目前的实验方法包括 使用检测方法测量细菌进入人类培养系的情况 各种组织来源的上皮细胞。生化抑制剂 原核结构/功能或真核细胞过程是 在这些组织培养侵袭试验中使用来检查 细菌吸收的要求。细菌的直接可视化 通过透射电子显微镜、视频显微镜测量进入， 荧光显微镜和共焦显微镜。 除了 上述机械方法，采用遗传技术 克隆负责的细菌基因。真核受体 细菌配体和对细菌的特异性真核细胞反应 通过抑制剂竞争测定、配体结合来测量入侵 感染真核细胞的测定和 mRNA 分析。信息 从这些方法中的每一种中获得的成果被整合起来以提供 对所研究的每个途径的细菌进入的机制的理解。