Metallobiochemistry of innate immunity and bacterial physiology

项目摘要 这项研究的主要目的是评估金属螯合人类宿主防御 蛋白质钙卫蛋白（CP）影响金属稳态和细菌病原体的生理学。金属离子 病原体必须从宿主那里获得这些营养物质才能复制， 引起感染。在此过程中，人类先天免疫系统的工作，以限制生物利用度的 包括锰（Mn）、铁（Fe）和锌（Zn）的过渡金属，通过部署CP和其它金属- 隔离感染部位的蛋白质。CP被接受为阻止Mn（II）和Zn（II）从微生物 病原体，我们最近证明，CP坐标铁在还原亚铁氧化态。 用于Fe（II）获取的细菌系统越来越被认为是多种疾病发病机制的关键。 感染状态，包括氧气变得有限的慢性生物膜介导的感染。但没有 已经鉴定了限制Fe（II）的其他宿主防御蛋白;因此研究CP作为Fe（II）- 隔离宿主防御蛋白对于理解慢性感染中宿主-病原体相互作用是重要的。 感染状态。铜绿假单胞菌（Pa）和金黄色葡萄球菌（Sa）是两种人类致病菌 在不同的患者群体中引起慢性多种微生物感染， 囊性纤维化（CF）患者。这种遗传性疾病使个体易患终生肺结核。 感染，其特征是使人衰弱的恶化，从而降低肺功能。值得注意的是，CF肺变得 随着疾病的进展，越来越缺氧，多种证据表明，Fe（II）成为 生物可利用铁的主要形式。CF肺病的进展也与微生物代谢的变化相关。 病因学，Sa是年轻患者中的主要微生物，随后是Pa定植 与肺功能下降有关。导致这一人口变化的潜在生物学仍然很差 然而，最近的研究表明，铁和CP都有助于这一过程。我们假设 CP限制了缺氧环境中Fe（II）的可用性，如CF肺中所发现的，并且这种活性 最终使得Pa在多微生物环境中胜过Sa。在目标1中，我们将评估Fe（II） 螯合的CP，和映射的分布的金属离子在Pa和Sa的文化与CP处理。在目标2中， 我们将检验这样一个假设，即CP将Fe（II）限制在Pa和Sa，从而影响个体的生理机能 以及这两种病原体的共培养动力学。这些调查将使未来的研究， CP和Fe如何驱动CF肺部感染的进展，并可能指导 治疗细菌感染的新的诊断、预防和治疗方法。

PROJECT SUMMARY The primary objective of this research initiative is to evaluate how the metal-sequestering human host-defense protein calprotectin (CP) affects metal homeostasis and physiology of bacterial pathogens. Metal ions are essential nutrients for all organisms, and pathogens must acquire these nutrients from the host to replicate and cause infection. During this process, the human innate immune system works to limit the bioavailability of transition metals including manganese (Mn), iron (Fe), and zinc (Zn) by deploying CP and other metal- sequestering proteins at sites of infection. CP is accepted to withhold Mn(II) and Zn(II) from microbial pathogens, and we recently demonstrated that CP coordinates Fe in the reduced ferrous oxidation state. Bacterial systems for Fe(II) acquisition are increasingly appreciated as critical for pathogenesis in multiple infection states, including chronic, biofilm-mediated infections where oxygen becomes limiting. However, no other host-defense proteins that limit Fe(II) have been identified; thus investigating CP as an Fe(II)- sequestering host-defense protein is important for understanding host-pathogen interactions in chronic infection states. Pseudomonas aeruginosa (Pa) and Staphylococcus aureus (Sa) are two human pathogens that cause chronic polymicrobial infections in diverse patient populations, including lung infections in individuals with cystic fibrosis (CF). This hereditary disease predisposes individuals to life-long pulmonary infections, marked by debilitating exacerbations that reduce lung function. Notably, the CF lung becomes increasingly hypoxic as disease progresses, and multiple lines of evidence indicate that Fe(II) becomes the predominant form of bioavailable Fe. Progression of CF lung disease is also correlated with a shift in microbial etiology, with Sa being the predominant microorganism in younger patients, and subsequent Pa colonization associated with lung function decline. The underlying biology that causes this population shift remains poorly understood; however, recent studies suggest that both Fe and CP contribute to this process. We hypothesize that CP limits Fe(II) availability in hypoxic environments, as found in the CF lung, and that this activity eventually allows Pa to outcompete Sa in polymicrobial environments. In Aim 1, we will evaluate Fe(II) sequestration by CP, and map the distribution of metal ions in Pa and Sa cultures treated with CP. In Aim 2, we will test the hypothesis that CP limits Fe(II) to Pa and Sa, and thereby impacts the individual physiologies and co-culture dynamics of these two pathogens. These investigations will enable future studies that address how CP and Fe drive the progression of CF lung infections, and may guide the design and development of novel diagnostic, preventative, and therapeutic approaches to treat bacterial infections.