Bacterial cation exporters: a new signaling paradigm for bacterial virulence

描述(申请人提供)：肺炎链球菌是全球发病率和死亡率的主要原因。肺炎球菌能够在人体内的许多部位定居和复制，包括鼻咽(定植)、肺部(肺炎)、血液(脓毒症)和脑(脑膜炎)。了解肺炎球菌疾病的发病机制的关键是了解细菌遇到的信号以及对这些信号的特定转录反应。一个未被探索的环境是肺炎球菌进入鼻咽并转移到血液中时遇到的高浓度、潜在有毒的钙和其他金属。这一应用将通过研究环境阳离子对全球基因表达的特定作用来阐明肺炎链球菌对二价阳离子的转录反应，并表征两个新发现的阳离子外排系统，这两个阳离子外排系统是宿主发病所必需的。在这项研究中，我们将通过基因芯片和qRT-PCR鉴定肺炎链球菌对钙、锰和锌的特异性转录反应。这项研究不仅提供了一个独特的视角来确定全球对细胞外阳离子感应的反应，而且由于这些转运蛋白中的突变积累了高水平的同源阳离子，因此可以确定细胞内阳离子浓度如何影响细胞信号通路。这些数据将被用来进一步了解肺炎链球菌对宿主中遇到的各种信号的具体反应。转运蛋白本身将受到广泛的分子特征的影响，包括通过对临床菌株进行测序来确定保守结构域，然后根据序列同源性产生突变以确定金属特异性和蛋白质功能的关键残基。通过在分子水平上表征这些转运蛋白而获得的知识将有助于我们理解细菌中的金属运输。公共卫生相关性：以前对病原菌的大多数研究都集中在金属的获得上，相比之下，我们对病原菌的金属外流的理解仍然是一个有趣的问题。此外，由于这些转运蛋白是宿主致病所必需的，它们提供了一个新的抗菌靶点。通过了解赋予金属离子选择性和功能的残基，这些研究将为病原菌用来输出人类宿主中丰富的阳离子的策略提供更多的洞察力。

DESCRIPTION (provided by applicant): Streptococcus pneumoniae is a major cause of morbidity and mortality worldwide. The pneumococcus is able to colonize and replicate in a number of sites in the human host including the nasopharynx (colonization), lungs (pneumoniae), blood (sepsis, and the brain (meningitis). Crucial to understanding the pathogenesis of pneumococcal disease is understanding the signals encountered by the bacteria and the specific transcriptional response to these signals. An unexplored setting is the high, potentially toxic, concentrations of calcium and other metals encountered as the pneumococcus enters the nasopharynx and translocates to the bloodstream. This application will elucidate the transcriptional response of S. pneumoniae to divalent cations by investigating the specific roles of environmental cations on global gene expression as well as characterize two newly discovered cation efflux systems that are required for host pathogenesis. In this study we will characterize the specific transcriptional responses of S. pneumoniae to calcium, manganese, and zinc by both microarray and qRT-PCR. This study provides a unique perspective to not only determine the global response to extracellular cation sensing, but since mutants in these transporters accumulate high levels of their cognate cation, allows for the determination of how intracellular cation concentration can influence cell signaling pathways. These data will be used to further understand the specific responses of S. pneumoniae to the various signals encountered in the host. The transporters themselves will be subject to extensive molecular characterization including determining conserved domains via sequencing clinical strain and then generating mutants based on sequence homology to ascertain critical residues for metal specificity and protein function. The knowledge gained by characterizing these transporters at a molecular level will aid in our understanding of metal transport in bacteria. PUBLIC HEALTH RELEVANCE: A majority of previous studies in pathogenic bacteria has focused on metal acquisition, in contrast our understanding of metal efflux by pathogenic bacteria remains an intriguing question. Furthermore, as these transporters are required for host pathogenesis, they provide a novel antibacterial target. By understanding the residues conferring metal ion selectivity and function, these studies will provide greater insight into the strategies utilized by pathogenic bacteria to export cations found in abundance in the human host.