The role of GraRS in resistance to cationic antimicrobial peptides in S. aureus

描述(申请人提供)：金黄色葡萄球菌是一种高度适应人类的病原体。这种适应的一个结果是它能够产生抗药性阳离子抗菌肽(CAPS)，这是宿主天然免疫的重要组成部分。在仔细研究金黄色葡萄球菌基因组时，我们发现了四个两组分的调控系统，每个系统都与ABC转运蛋白系统相邻。其中一个系统，GraRS，调节邻近的VraFG外排系统，包括ATPase和膜通透酶。GRAR或vraG的突变导致了对CAP如RP-1(来自血小板的PMP-1的同系物)和HNP-1(来自中性粒细胞的HNP-1)、吲哚西丁和LL37的耐药性的降低。作为一种血管内病原体，金黄色葡萄球菌可能会遇到来自血小板和中性粒细胞(RP-1和HNP-1)的帽，这是我们在本提案中将重点关注的。根据诱导研究和微阵列分析，金黄色葡萄球菌对选择性CAP的暴露也导致了graRS和vraFG以及其他与CAP抗性相关的基因的诱导，包括mprF、dltABCD和另一组ABC转运蛋白基因vraDE。然而，这些基因在金黄色葡萄球菌GRARS介导的CAP抗性中的确切作用尚未确定(目标I)。此外，与任何帽子都能诱导GraRS和vraFG表达的表皮葡萄球菌不同，金黄色葡萄球菌的诱导更具选择性，只对少数帽子有反应。根据金黄色葡萄球菌和表皮葡萄球菌的激酶传感器GRAS序列的差异，我们建议定义GRAS中的确切残基，这些残基介导了下游基因诱导的特定帽子的识别(AIM II)。除了细菌因素，我们还对合成帽的结构以及它们结合和诱导GraRS表达的能力感兴趣(目标III)。最后，这些发现在GraRS中的体内相关性将在相关的感染性心内膜炎兔模型中进行探索。综上所述，这些研究将引出金黄色葡萄球菌对相关宿主帽的先天抵抗力的科学基础，从而提供一个平台，以制定策略来减轻对阳离子抗菌肽的抵抗力。

DESCRIPTION (provided by applicant): S. aureus is a highly adaptive human pathogen. One result of this adaptation is its ability to develop resistance cationic antimicrobial peptides (CAPs), which are an important part of innate immunity in the hosts. In perusing the S. aureus genomes, we found four two component regulatory systems each of lie adjacent to an ABC transporter system. One of the system, GraRS, regulates the adjacent VraFG efflux system comprising an ATPase and a membrane permease. Mutations in graR or vraG have led to a reduction in resistance to CAPs such as RP-1 (a congener to PMP-1 from platelets) and hNP-1 (from neutrophils), indolicidin and LL37. As an intravascular pathogen, S. aureus likely encounters CAPs from platelets and neutrophils (RP-1 and hNP-1), which we will focus on in this proposal. The exposure of S. aureus to selective CAP has also led to induction of graRS and vraFG and other genes relevant CAP resistance including mprF, dltABCD and also another set of ABC transporter genes vraDE, based on induction studies and microarray analysis. However, the exact contribution of these genes to CAP resistance mediated by GraRS in S. aureus is not defined (Aim I). In addition, in contrast to S. epidermidis where the expression of graRS and vraFG can be induced by any CAPs, the induction in S. aureus is more selective, responding to only a few CAPs. Based on the difference in the sequence of the kinase sensor GraS between S. aureus and S. epidermidis, we propose to define the exact residues within GraS that mediate this recognition of specific CAPs for induction of downstream genes (Aim II). Besides the bacterial factors, we are also interested in the structure of synthetic CAPs and their ability to bind and induce expression of graRS (Aim III). Finally, the in vivo correlate of these finding in GraRS will be explored in a relevant rabbit model of infective endocarditis. Together, these studies will elicit the scientific basis of innate resistance to relevant host CAPs in S. aureus, thus providing a platform to devise strategies to alleviate resistance to cationic antimicrobial peptides.