Two pathways for calcium signaling and virulence regulation in P. aeruginosa

铜绿假单胞菌中钙信号传导和毒力调节的两条途径

• 批准号: 10459268
• 负责人: Marianna Patrauchan
• 金额: $ 18.55万
• 依托单位: OKLAHOMA STATE UNIVERSITY STILLWATER
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-07-01 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10459268
• 关键词: Acute; Affinity; Alginates; Antibiotics; Artificial Implants; Bacteria; Bacterial Infections; Binding; Binding Proteins; Bioinformatics; Calcium Binding; Calcium Signaling; Calcium ion; Cause of Death; Cell physiology; Cells; Centers for Disease Control and Prevention (U.S.); Chronic; Chronic Obstructive Pulmonary Disease; Communicable Diseases; Core Facility; Cyprinus carpio; Cystic Fibrosis; Cystic Fibrosis Transmembrane Conductance Regulator; Defect; Dependence; Development; EF Hand Motifs; Endocarditis; Environment; Eukaryota; Eukaryotic Cell; Gene Expression; Gene Expression Regulation; Genetic; Genetic Transcription; Homeostasis; Immune response; Immunocompromised Host; In Vitro; Infection; Integration Host Factors; Invaded; Knowledge; Lead; Life; Liquid substance; Lung; Measures; Microbial Biofilms; Molecular; Molecular Conformation; Mutation; Nosocomial pneumonia; Oklahoma; Organism; Oxidative Stress; Pathogenicity; Pathway interactions; Patients; Peptide Hydrolases; Phagocytosis; Physiological; Physiological Adaptation; Play; Production; Prognosis; Prokaryotic Cells; Proteins; Pseudomonas; Pseudomonas Infections; Pseudomonas aeruginosa; Pyocyanine; Regulation; Research; Resistance; Respiratory Center; Respiratory Disease; Respiratory Tract Infections; Rest; Role; Route; Second Messenger Systems; Signal Transduction; Signal Transduction Pathway; System; Testing; Time; Transducers; Virulence; Virulence Factors; World Health Organization; acute infection; base; cell motility; chronic infection; combat; cystic fibrosis patients; healthcare-associated infections; human pathogen; in vivo; individual patient; innovation; novel; novel strategies; pathogen; priority pathogen; protein function; pyoverdin; response; rhamnolipid; sensor; wound

Abstract Pseudomonas aeruginosa is an opportunistic human pathogen that causes severe, life threatening infections in patients with cystic fibrosis (CF), endocarditis, wounds, artificial implants, and in healthcare-associated infections. The versatility of P. aeruginosa pathogenicity is associated with an outstanding physiological adaptability of the organism and its ability to modulate host responses, due in part to a tightly coordinated regulation of gene expression. Therefore, to gain control over currently untreatable Pseudomonas infections, it is critically important to generate new knowledge of the regulatory circuits coordinating the pathogen virulence in response to host factors. Calcium ion (Ca2+) is an essential intracellular messenger in eukaryotic cells, regulating vital cellular processes. It accumulates in pulmonary fluids of CF patients and in mitral annulus of endocarditis patients. Alterations in the host Ca2+ homeostasis may serve as a trigger for enhanced virulence of invading pathogens. In support, we showed that Ca2+ positively regulates biofilm formation, swarming, and production of several virulence factors in P. aeruginosa. However, the molecular mechanisms of such regulation are not known. It is also not known whether intracellular Ca2+ plays role as a second messenger in prokaryotes as it does in eukaryotes. Understanding the mechanisms of Ca2+ regulation, signaling and homeostasis will provide novel means for controlling P. aeruginosa viability, virulence, and interactions with the host. Earlier, we identified two putative Ca2+-binding proteins EfhP and CarP, mutations in which cause multiple Ca2+-dependent defects in virulence and infectivity. EfhP contains two EF-hand motives, known to bind Ca2+ and relay Ca2+ signal through conformational changes. CarP is predicted to form a beta-propeller and has a putative phytase domain. Based on the bioinformatics and preliminary studies, we hypothesize that EfhP and CarP provide different routes of Ca2+ signal transduction regulating virulence and host- pathogen interactions in response to Ca2+ in a host. To test this, we propose to determine the cellular localization and identify binding partners and signal-transducing pathways regulated by the two proteins. We will also characterize the role of EfhP and CarP in P. aeruginosa interactions with a host, and define their involvement in the development of acute and chronic infections. By utilizing the expertise of three OCRID core facilities, we will unravel the mechanisms of Ca2+ signaling and its role in regulating the ability of P. aeruginosa to cause infections at the molecular, cellular, and organismal level. This research is highly innovative as for the first time it will experimentally demonstrate Ca2+ signaling in bacteria, identify the components of Ca2+ signal transduction pathways, and define the role of Ca2+ signaling in P. aeruginosa pathogenicity in vivo.

摘要 铜绿假单胞菌是一种机会性的人类病原体，其导致严重的、危及生命的感染， 囊性纤维化（CF）、心内膜炎、伤口、人工植入物和医疗保健相关患者 感染.铜绿假单胞菌致病性的多功能性与其显著的生理学特性相关。 生物体的适应性及其调节宿主反应的能力，部分原因是由于紧密协调的 基因表达的调控。因此，为了控制目前无法治疗的假单胞菌感染， 对于产生协调病原体毒力的调节回路的新知识至关重要 以应对宿主因素。钙离子（Ca 2+）是真核细胞中一种重要的胞内信使， 调节重要的细胞过程。它积聚在CF患者的肺液中和CF患者的二尖瓣环中。 心内膜炎患者。宿主Ca 2+稳态的改变可能是增强毒力的触发因素 入侵的病原体。作为支持，我们发现Ca 2+正调控生物膜的形成，群集， 铜绿假单胞菌中几种毒力因子的产生。然而，这种分子机制 规则是未知的。细胞内Ca ~（2+）是否作为第二信使， 就像真核生物一样。了解Ca 2+调节、信号传导和 稳态将提供新的手段来控制铜绿假单胞菌的活力，毒力和相互作用， 主持人早些时候，我们鉴定了两种假定的Ca 2+结合蛋白EfhP和CarP， 在毒力和感染性方面存在多种依赖钙离子的缺陷。EfhP包含两个已知的EF手动机， 结合Ca ~（2+）并通过构象变化传递Ca ~（2+）信号。据预测，CarP将形成β螺旋桨， 具有推定的植酸酶结构域。基于生物信息学和初步研究，我们假设， EfhP和CarP提供不同的Ca 2+信号转导途径，调节毒力和宿主- 病原体相互作用响应于宿主中的Ca 2+。为了验证这一点，我们建议确定细胞 定位和鉴定由两种蛋白质调节的结合伴侣和信号转导途径。我们 还将描述EfhP和CarP在铜绿假单胞菌与宿主相互作用中的作用，并定义其 参与急性和慢性感染的发展。通过利用OCRID三个核心的专门知识， 本研究旨在阐明Ca 2+信号转导机制及其在铜绿假单胞菌细胞增殖能力调控中的作用 在分子、细胞和有机体水平上引起感染。这项研究具有很高的创新性， 第一次将实验证明细菌中的Ca 2+信号，识别Ca 2+信号的成分 转导途径，并定义了Ca 2+信号转导在铜绿假单胞菌体内致病性中的作用。