Elucidating the A. baumannii response to calprotectin-mediated zinc starvation

描述(申请人提供)：鲍曼不动杆菌是呼吸机相关肺炎患者最常见的分离细菌之一，可导致多种感染，特别是在重症监护病房的高度易感人群中。鲍曼不动杆菌对抗生素的耐药性迅速发展，导致我们有效治疗感染的能力显著下降。因此，针对替代预防措施和治疗的研究有了相当大的扩展。了解鲍曼不动杆菌的致病机制将提高我们成功进行治疗干预的能力，并可能识别可以作为开发新的抗菌剂或疫苗的靶向的细菌因子。钙保护素(CP)是一种宿主蛋白，存在于炎症部位，在那里它具有促炎活性。CP还通过隔离必需金属锌(锌)和锰(锰)而显示出抗微生物活性。CP介导的锌螯合作用抑制鲍曼不动杆菌的生长，在鲍曼不动杆菌肺炎期间，CP的丢失促进了细菌的增殖和传播。众所周知，细菌 通过特殊获取系统的表达、代谢过程的改变和金属蛋白质组的改变来感知和响应营养饥饿。缺乏这些能力的细菌通常无法确定成功的感染。因此，鲍曼不动杆菌必须采取策略在宿主体内获取锌。在鉴定鲍曼不动杆菌适应CP介导的锌饥饿所需的细菌因子的实验中，我们发现了一种鲍曼不动杆菌的锌获取系统，我们将其命名为ZnuABC。这个系统由一个锌响应调节器控制，我们将其命名为Zur。此外，在对Zur和ZnuABC编码基因上游基因组区域的电子分析中，ABC发现了一个被称为Zur盒的保守序列，Zur与其结合以抑制下游基因的表达。对鲍曼不动杆菌基因组中这一共识序列的搜索发现了可能受Zur和锌调控的其他基因。我们推测，在感染后，CP产生了缺锌的环境，鲍曼不动杆菌通过上调锌获取系统和其他锌反应基因做出反应，这些基因对致病至关重要。这一提议解决了我们在以下三个综合具体目标中的假设。目的1：探讨CP介导的锌饥饿在鲍曼不动杆菌肺炎中的作用。目的2：确定鲍曼不动杆菌对锌限制条件的反应。目的：阐明两个锌调控基因在鲍曼不动杆菌锌稳态中的作用。总之，这些研究将提高我们对细菌性肺炎期间宿主营养免疫的理解，并确定鲍曼不动杆菌是如何颠覆这种宿主防御而致病的。此外，这些机制的特征将为开发改进的治疗方法奠定基础，以应对目前治疗鲍曼不动杆菌感染的疗效迅速下降的情况。

DESCRIPTION (provided by applicant): Acinetobacter baumannii is one of the most frequently isolated bacteria from patients with ventilator-associated pneumonia and causes a diverse number of infections, particularly in highly susceptible individuals in intensive care unit. The rapid development of antibiotic resistance by A. baumannii has led to a significant decrease in our ability to effectively treat infections. Consequently, there has been a considerable expansion in research directed towards alternative preventative measures and treatments. Understanding the mechanisms of A. baumannii pathogenesis will improve our capacity for successful therapeutic intervention and may identify bacterial factors that can be targeted for development of new antimicrobials or vaccines. Calprotectin (CP) is a host protein present at sites of inflammation where it displays pro-inflammatory activity. CP also exhibits antimicrobial activity through sequestration of the essential metals zinc (Zn) and manganese (Mn). CP- mediated Zn chelation inhibits A. baumannii growth, and during A. baumannii pneumonia, the loss of CP enhances bacterial proliferation and dissemination. It is well-established that bacteria sense and respond to nutrient starvation via expression of specialized acquisition systems, modification of metabolic processes, and alterations to the metalloproteome. Bacteria that lack these capabilities are generally unable to establish a successful infection. Therefore, A. baumannii must employ strategies to acquire Zn within the host. In experiments to identify bacterial factors required for A. baumannii adaptation to CP-mediated Zn starvation, we discovered an A. baumannii Zn acquisition system that we have named ZnuABC. This system is under the control of a Zn-responsive regulator, which we have named Zur. Further in silico analysis of the genomic region upstream of the genes encoding Zur and ZnuABC uncovered a conserved sequence called a zur box, to which Zur binds to repress downstream gene expression. A search for this consensus sequence within the A. baumannii genome identified additional genes that may be regulated by Zur and Zn. We hypothesize that following infection, CP generates a Zn-starved environment, and A. baumannii responds through the up- regulation of a Zn acquisition system and other Zn-responsive genes that are essential for pathogenesis. This proposal addresses our hypothesis in the following series of three integrated Specific Aims. Aim 1: Determine the role of CP-mediated Zn starvation to A. baumannii pneumonia. Aim 2: Define the A. baumannii response to Zn-limiting conditions. Aim 3: Elucidate the functions of two Zn-regulated genes in A. baumannii Zn homeostasis. Together these studies will improve our understanding of host nutritional immunity during bacterial pneumonia and define how A. baumannii subverts this host defense to cause disease. Furthermore, characterization of these mechanisms will lay the groundwork for the development of improved therapeutics in the face of the rapidly decreasing efficacy of the current treatments for A. baumannii infection.