Using Salmonella Pathogenesis and Cell Biology as a Discovery Tool

使用沙门氏菌发病机制和细胞生物学作为发现工具

基本信息

批准号：
10665946
负责人：
Corrella S Detweiler
金额：
$ 23.48万
依托单位：
UNIVERSITY OF COLORADO
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-03-01 至 2025-02-28
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10665946
关键词：
Address Animals Bacteria Bacterial Infections Binding Biological CRISPR interference Cell Culture Techniques Cell Membrane Permeability Cell membrane Cells Cellular biology Chemicals Cholesterol Colony-forming units Cytolysis Data Dose Escherichia coli Female Future Genes Genetic Gram-Negative Bacteria Growth Infection Interruption Ions Lead Lipid Bilayers Lipids Liver Macrophage Macrophage Cell Biology Mammalian Cell Membrane Membrane Proteins Methods Microscopy Modification Mus Natural Immunity Nature Pathogenesis Pathogenicity Pathway interactions Peptides Permeability Phagosomes Polymyxin B Process Propidium Diiodide Proteins Protons Publishing Resistance Salmonella Salmonella typhimurium Spleen Testing Toxic effect Vesicle Virulence Work biological adaptation to stress cell injury efflux pump experimental study high reward high risk in vivo inhibitor innovation knock-down male mutant nanomolar negative affect novel strategies pathogen sex small molecule tool transcriptomics voltage

项目摘要

SUMMARY The inner membrane of pathogenic Gram-negative bacteria contains lipids and proteins that are distinct from those of mammalian cell membranes. Inner membranes therefore could be attacked by small molecules during infection, when soluble host innate immunity damages the protective outer membrane barrier, increasing access to chemicals. Exploration of molecules that a) interrupt pathogenesis in cell culture with minimal host cell damage, and b) perturb inner membranes under broth conditions that permeabilize the outer membrane has the potential to reveal whether bacterial virulence can be short-circuited by compounds that disturb inner membranes but do not inhibit bacterial growth under standard broth conditions. An in-macrophage cell biology- based method called SAFIRE appears to identify compounds that negatively affect bacterial inner membranes while sparing host cell membranes. Within this application we propose to study a SAFIRE-identified compound, D66, which appears to damage bacterial inner membranes without lysing them. Published and unpublished preliminary data together indicate that D66 accesses bacterial inner membranes when the LPS layer is damaged or efflux pumps are compromised, conditions consistent with the macrophage phagosome. However, D66 disrupts inner membrane voltage rapidly and at concentrations that do not destroy the inner membrane lipid bilayer, indicating that the compound is not causing bacterial lysis but is disturbing the membrane more subtly. We hypothesize that D66 damages the bacterial cell membrane in a process that triggers stress response pathways but does not cause rapid physical disruption of the lipid bilayer. We propose cell biological and animal infection experiments to test this hypothesis.

概括致病革兰氏阴性细菌的内膜含有脂质和蛋白质，与众不同哺乳动物细胞膜的那些。因此，内膜可能会受到小分子的攻击感染时，当可溶性宿主先天免疫会损害保护性外膜屏障时，增加进入化学物质。探索分子的a）在细胞培养中中断最小宿主的细胞培养的发病机理细胞损伤和b）在肉汤条件下扰动内膜，使外膜渗透有可能揭示细菌毒力是否可以被干扰内部的化合物短路膜，但不抑制标准肉汤条件下的细菌生长。巨噬细胞生物学 - 基于称为Safire的方法似乎识别对细菌内膜负面影响的化合物同时保留宿主细胞膜。在此应用程序中，我们建议研究一种被鉴定的化合物， D66，它似乎不会损害细菌性内膜而不会裂解它们。出版和未出版初步数据一起表明D66在LPS层是时访问细菌内膜受损或外排泵被损害，条件与巨噬细胞吞噬体一致。然而， D66迅速破坏内膜电压，并以不会破坏内膜的浓度脂质双层，表明该化合物并未引起细菌裂解，而是在膜上干扰膜巧妙。我们假设D66在触发压力的过程中损害细菌细胞膜响应途径，但不会引起脂质双层的快速物理破坏。我们提出细胞生物学和动物感染实验以检验该假设。