PlrSR-dependent Signal Transduction in Bordetella Virulence

摘要/摘要： 百日咳(又名百日咳)是一种重新出现的严重公共卫生问题，据估计发病率 每年有2000万个病例，死亡(主要是婴儿)每年约20万人。最近的涨幅 百日咳在疫苗覆盖率高的国家，如美国，与从 全细胞(WP)到无细胞(AP)百日咳疫苗，并归因于更大的感染宿主 由接种(而不是感染)疫苗的青少年和年轻人组成的个人 孩子们。现在很明显，接种AP疫苗诱导的免疫力不如接种Ap疫苗那么持久 接种可湿性肺炎疫苗，其持久性不如感染百日咳杆菌诱导的免疫力那么持久， 百日咳的主要致病因素。此外，尽管WP和AP疫苗的免疫提供了 对疾病的保护(至少在最初)，它不能防止殖民。新的疫苗 提供消毒、持久的免疫力是必需的。我们发现了一种以前未曾有过的特征 百日咳杆菌及其广泛宿主所需的信号转导系统(PlrSR) 范围病原体支气管败血波尔德氏菌在下呼吸道(LRT)定居并持续存在。我们的 初步数据支持一种模型，在该模型中，PlrSR作为一种激酶对增加的二氧化碳和 低氧条件(反映LRT)，导致PlrR-磷酸(PlrR~P)水平高， 激活基因的表达，包括那些编码高亲和力细胞色素氧化酶的基因。我们的模特 指出在有氧条件下，PLRs主要作为一种磷酸酶发挥作用，而低水平的 PlrR~P是细胞存活所必需的。我们将使用遗传学、分子生物学和全基因组 识别PlrSR调节基因的方法，特别是那些仅在LRT中诱导的基因，并将 确定依赖PlrSR的基因调控的作用以及被调控者编码的因子 毒力基因。使用遗传方法，我们将确定PLRS PDC和PAS的作用 结构域，以及预测的PLRs的激酶和磷酸酶活性，在细菌的能力 在体外和LRT中生长。使用生化方法，我们将确定PLRS是否是氧化还原- 一种敏感的含血红素的蛋白，在低氧条件下作为一种激酶发挥作用，并在 环境空气中的磷酸酶，我们将鉴定PlrR~P结合的DNA序列。我们的结果将 重要的是因为之前未知的依赖于plrSR的毒力因子，以及plrSR系统 它本身将是开发新的成分疫苗和疫苗靶标的极佳候选者 新疗法的发展。我们的结果也将促进我们对NtrYX家族蛋白的理解 (PlrSR是其中的一员，并控制其他病原体的毒力)起作用，它们将 提供对呼吸道病原体一般如何在LRT中生长的洞察。

Summary/Abstract: Pertussis (aka whooping cough) is a serious reemerging public health problem with incidence estimated at 20 million cases annually and deaths (mostly in infants) at ~200,000 annually. Recent rises in pertussis in countries with high vaccine coverage, such as the United States, correlate with a switch from whole cell (wP) to acellular (aP) pertussis vaccines and are attributed to a larger reservoir of infected individuals composed of adolescents and young adults who were vaccinated (rather than infected) as children. It is now apparent that immunity induced by aP vaccination is not as durable as vaccination by wP vaccination, which is not as durable as immunity induced by infection with Bordetella pertussis, the primary causal agent of pertussis. Moreover, while immunization with wP and aP vaccines provides protection against disease (at least initially), it does not protect against colonization. New vaccines that provide sterilizing, long-lasting immunity are needed. We have discovered a previously uncharacterized signal transduction system (PlrSR) that is required for B. pertussis and the closely-related broad host range pathogen Bordetella bronchiseptica to colonize and persist in the lower respiratory tract (LRT). Our preliminary data support a model in which PlrSR functions as a kinase in response to increased CO2 and low oxygen conditions (reflective of the LRT), resulting in high levels of PlrR-phosphate (PlrR~P) that activate expression of genes including those encoding high-affinity cytochrome oxidases. Our model states that under aerobic conditions, PlrS functions primarily as a phosphatase, and that low levels of PlrR~P are essential for cell viability. We will use genetic, molecular biological, and genome-wide approaches to identify PlrSR-regulated genes, especially those induced only in the LRT, and will determine the roles of PlrSR-dependent gene regulation and of the factors encoded by the regulated genes in virulence. Using genetic approaches, we will determine the role of the PlrS PDC and PAS domains, as well as the predicted kinase and phosphatase activities of PlrS, in the ability of the bacteria to grow in vitro and in the LRT. Using biochemical approaches, we will determine if PlrS is a redox- sensitive heme-containing protein that functions as a kinase under low oxygen conditions and a phosphatase in ambient air, and we will identify DNA sequences to which PlrR~P binds. Our results will be significant because previously unknown PlrSR-dependent virulence factors, and the PlrSR system itself, will be excellent candidates for the development of new component vaccines and targets for the development of new therapeutics. Our results will also advance our understanding NtrYX family proteins (of which PlrSR is a member and which control virulence in other pathogens) function and they will provide insight into how respiratory pathogens, in general, grow in the LRT.