Using Salmonella Pathogenesis and Cell Biology as a Discovery Tool

摘要 致病性革兰氏阴性细菌的内膜含有不同于 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，当可溶性宿主的先天免疫破坏保护性的外膜屏障，增加 获取化学品的途径。探索a)在最少宿主的细胞培养中干扰致病的分子 细胞损伤，以及b)在使外膜通透的肉汤条件下扰乱内膜 有可能揭示细菌的毒力是否会被扰乱内部的化合物短路 膜，但在标准的肉汤条件下不能抑制细菌生长。巨噬细胞内的细胞生物学- 基于一种名为Safire的方法似乎可以识别对细菌内膜有负面影响的化合物 同时保留宿主细胞膜。在本申请中，我们建议研究Safire鉴定的化合物， D66，它似乎破坏细菌内膜而不裂解它们。已出版和未出版 初步数据表明，当脂多糖层是 受损或外排泵受损，情况与巨噬细胞吞噬小体一致。然而， D66在不破坏内膜的浓度下迅速破坏内膜电压 脂双层，表明该化合物不会引起细菌溶解，但会更多地干扰细胞膜 巧妙地。我们假设D66在触发压力的过程中破坏细菌细胞膜 反应途径，但不会引起脂质双层的快速物理破坏。我们提出了细胞生物学 以及动物感染实验来验证这一假说。

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 Published and unpublished preliminary data together indicate that D66 accesses bacterial inner membranes when the LPS layer is damaged or efflux pumps are compromised, conditions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.