METABOLIC REGULATION OF STAPHYLOCOCCAL PATHOGENESIS

这个子项目是利用由NIH/NCRR资助的中心拨款提供的资源的许多研究子项目之一。子项目和调查员(PI)可能从另一个NIH来源获得了主要资金，因此可能会出现在其他CRISE条目中。列出的机构是针对中心的，而不一定是针对调查员的机构。金黄色葡萄球菌是一种革兰氏阳性病原体，可导致人类和动物的多种疾病。它是医院获得性感染的主要原因，每年给美国造成超过45亿美元的损失。大多数金黄色葡萄球菌毒力因子的表达是由生长阶段决定的，指数生长阶段的特征是产生细胞相关的黏附因子(如纤维连接蛋白结合蛋白)，而指数后生长阶段的特征是产生分泌的毒力因子(如α-毒素)。伴随着进入指数后生长阶段的是三羧酸(TCA)循环活性的增加。乌头酸酶是一种三氯乙酸循环酶，可将柠檬酸转化为异柠檬酸。真核生物具有线粒体和细胞质乌头酸酶活性。细胞质中的乌头酸酶活性是由铁反应蛋白-1(IRP-1)引起的，IRP-1是一种转录后调节铁调节蛋白合成的mRNA结合蛋白。因此，胞浆乌头酸酶是一种双功能蛋白。最近，枯草芽孢杆菌和大肠杆菌的乌头酸酶被证明以一种铁依赖的方式与mRNAs中的序列特异性元件结合。这些观察结果证实，细菌乌头酸酶与真核细胞胞浆乌头酸酶/IRP-1一样，是一种双功能蛋白。唯一的金黄色葡萄球菌乌头酸酶基因(ACNA/CITB)的失活显著减少了毒力因子的合成，改变了宿主与病原体的相互作用，并提高了静止相的存活率。乌头酸酶失活的影响可能是TCA循环中代谢障碍的结果，也可能是调节功能的丧失，或者两者的结合。我们建议确定乌头酸酶失活的哪些影响是由于代谢缺陷，哪些是由于调节功能的丧失。此外，我们将确定乌头酸酶是否会与被确认为潜在受乌头酸酶调控的基因的同源mRNAs结合。细菌是一种单细胞有机体，可以“进食”和分裂；因此，细菌所做的一切都与进食和分裂有关，包括导致人类疾病。对细菌如何调节致病机制以应对营养限制的基本了解，对于开发旨在杀死细菌的新疗法至关重要。该提案中包含的工作是设计抗击细菌感染的新战略的第一步。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Staphylococcus aureus is a Gram-positive pathogen that causes a wide variety of diseases in humans and animals. It is a leading cause of hospital acquired infections, costing the United States more than 4.5 billion dollars annually. The expression of most S. aureus virulence factors is determined by the growth phase.The exponential phase of growth is characterized by the production of cell associated adhesion factors (e.g., fibronectin binding protein) while the post-exponential phase of growth is distinguished by the production of secreted virulence factors (e.g., alpha-toxin). Concomitant with the entry into the post-exponential phase of growth is an increase in tricarboxylic acid (TCA) cycle activity. Aconitase is a TCA cycle enzyme that converts citrate to isocitrate. Eukaryotic organisms have mitochondrial and cytoplasmic aconitase activity. The cytoplasmic aconitase activity is caused by the iron-responsive protein-1 (IRP-1), an mRNA-binding protein that posttranscriptionally regulates the synthesis of iron-regulated proteins. Thus, cytosolic aconitase is a bifunctional protein. Recently, it has been demonstrated that aconitase from Bacillus subtilis and Escherichia coli bind to sequence specific-elements in mRNAs in an iron-dependent fashion. These observations established bacterial aconitase, like eukaryotic cytosolic aconitase/IRP-1, as a bifunctional protein. Inactivation of the sole S. aureus aconitase gene (acnA/citB), significantly decreases virulence factor synthesis, alters host-pathogen interaction, and enhances stationary phase survival. The affects of aconitase inactivation can be the result of a metabolic block in the TCA cycle, the loss of regulatory function, or a combination of the two. We propose to determine which affects of aconitase inactivation are due to a metabolic defect and which are due to the loss of regulatory function. Additionally, we will determine if aconitase will bind to the cognate mRNAs of genes identified as potentially regulated by aconitase. [Lay Summary] Bacteria are single celled organisms that "eat" and divide; therefore, everything bacteria do is linked to eating and dividing, including causing disease in humans. A fundamental understanding of how bacteria regulate pathogenesis in response to nutrient limitation is critical to developing new treatments designed to kill bacteria. The work contained within this proposal is a first step in designing new strategies to combat bacterial infections.