Enterococcal Pathogenesis:Role of Cytolysin

描述(由申请人提供)：肠球菌已成为耐多药医院感染的主要原因。然而，它们作为从昆虫到人类的一切生物的共生微生物已经存在了4亿多年。我们之前的工作以及其他人的工作表明，使医院分离的粪肠球菌毒力更强的因素之一是一种被称为“细胞溶素”的毒素。粪肠球菌溶细胞素具有抗菌特性，也是一种对人体细胞的溶细胞毒素。我们先前证明，这种毒素使感染导致的死亡风险增加了5倍，它是在致病性岛上传递的，并且它的表达受到一种名为“遥感”的新过程的调节，该过程允许细菌检测附近目标细胞的存在，并产生高水平的毒素。虽然我们已经证明粪肠球菌溶细胞素毒素在结构和调控上是新的，但我们还不知道它如何影响人类感染的严重性，或者在使粪肠球菌感染率高达100%的感染模型中。 剧毒要大几倍。我们也不清楚“遥感”是如何在 转录水平。因此，我们建议确定毒素的抗菌特性是否允许粪肠球菌进入胃肠道微生物区系，是否会导致细菌组成的变化和由于抗菌活性而在结肠内过度生长，是否通过对哺乳动物细胞的溶细胞活性损害结肠的屏障功能，或者是否通过阻碍先天性免疫系统将有机体从感染部位清除而加剧感染。为了了解它的表达是如何调控的，我们还建议剖析新的两组分调控系统，该系统已被证明是其抑制和抑制所必需的。这将通过绘制启动细胞溶素操纵子转录的精确点，并确定这些起始点与操纵子调节区结构特征之间的关系来完成。我们将确定导致高水平毒素表达的诱导信号是否来自细胞外部，或者t是否内化并直接导致操纵子抑制。最后，我们将确定每个调控蛋白是如何相互联系的，以及与溶细胞素操纵子控制区内的结构特征之间的联系。这些结果将为了解细胞溶血素如何加剧粪肠球菌感染以及 控制其表达所需的信息。

DESCRIPTION (provided by applicant): Enterococci have emerged as leading causes of multidrug resistant hospital infection. Yet they have existed as commensal microbes of everything from insects to man for over 400 million years. Our previous work, as well as the work of others, has shown that one of the factors that makes hospital isolate of Enterococcus faecalis more virulent is a toxin termed "cytolysin." The E. faecalis cytolysin has antibacterial properties, as well as being a cytolytic toxin for human cells. We previously showed that this toxin contributed a 5-fold increased risk of death from infection, that it was conveyed on a pathogenicity island along with several additional virulence traits, and that its expression is regulated by a novel process termed "telesensing" that allows the bacterium to detect the presence of target cells in the vicinity, and in respose produce high levels of the toxin. Although we have shown that the E. faecalis cytolysin toxi is novel in structure and regulation, we do not yet know how it contributes to the severity of infection in man, or in models of infection where it renders E. faecalis infection as much as 100 times more acutely toxic. We also do not know exactly how "telesensing" works at the level of transcription. Therefore, we propose to determine whether the antibacterial property of the toxin allows E. faecalis entry into the GI tract microbiota, whether it there causes changes in the bacterial composition and overgrows within the colon because of the antibacterial activity, whether it compromises the barrier function of the colon through its cytolytic activity for mammalian cells, or whether it exacerbates infection by impeding the innate immune system from clearing the organism from a site of infection. To understand how its expression is regulated, we also propose to dissect the novel two component regulatory system which has been shown to be necessary for its repression and depression. This will be done by mapping the precise points where transcription of the cytolysin operon is initiated, and determining the relationship between those start sites and structural features of the regulatory region of the operon. We will determine whether the inducing signal that results in high level toxin expression is communicated from the outside of the cell, or whether t is internalized and directly causes operon depression. Finally, we will determine exactl how each of the regulatory proteins relate to each other, and relate to structural features within the control region of the cytolysin operon. These results will provide needed information for knowing how the cytolysin exacerbates E. faecalis infection, as well as the information needed for controlling its expression.