The non-redundant role of the Has and Phu heme acquisition systems of Pseudomonas aeruginosa in iron homeostasis

铜绿假单胞菌 Has 和 Phu 血红素获取系统在铁稳态中的非冗余作用

• 批准号: 9469309
• 负责人: Alecia Tennessee Dent
• 金额: $ 2.92万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-12-01 至 2019-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9469309
• 关键词: Acinetobacter; Acinetobacter baumannii; Affinity; Anti-Bacterial Agents; Antibiotic Resistance; Antibiotics; Assimilations; Bacteria; Biochemical; Biophysics; Cell surface; Centers for Disease Control and Prevention (U.S.); Chronic; Clinical; Complement; Coupled; Development; Enterobacter; Enterococcus faecium; Future; Genes; Genetic Transcription; Goals; Heme; Heme Iron; Homeostasis; Human body; Immune; In Vitro; Infection; Iron; Isotope Labeling; Klebsiella pneumoniae; Knock-out; Laboratories; Ligation; Lung; Membrane; Methods; Molecular; Multi-Drug Resistance; Mutation; Neisseria gonorrhoeae; Nutrient; Operon; Pathway interactions; Patients; Production; Property; Proteins; Pseudomonas; Pseudomonas aeruginosa; Receptor Gene; Regulation; Reporting; Resistance; Role; Siderophores; Sigma Factor; Signal Transduction; Site-Directed Mutagenesis; Source; Staphylococcus aureus; System; Techniques; Therapeutic; Time; United States; Virulence; antimicrobial; bacterial genetics; base; cystic fibrosis patients; experimental study; extracellular; heme receptor; in vivo; metabolomics; mortality; novel; pathogen; pathogenic bacteria; promoter; pyochelin; pyoverdin; receptor; resistant strain; sensor; transcriptome; uptake

Project Summary Pathogenic bacteria require iron for their survival and virulence. The opportunistic pathogen Pseudomonas aeruginosa has many mechanisms by which it can acquire iron, including ferric and ferrous iron uptake systems. However, within the host P. aeruginosa adapts to utilize heme as an iron source via the heme assimilation (has) and Pseudomonas heme utilization (phu) systems. The has operon encodes an extracellular hemophore, HasAp that scavenges heme and transfers it to the outer membrane (OM) receptor, HasR which acts as the cell surface signaling (CSS) system. The hemophore and receptor genes are downstream of an Extra Cytoplasmic Function (ECF) sigma and anti-sigma factor (HasI and S, respectively) associated with signal transduction and regulation of heme uptake. We have recently shown that in contrast to the bis-His coordination of HasR, the OM receptor PhuR has a unique His-Tyr coordination, an emerging motif in high affinity heme acquisition systems. 13C-heme isotopic labeling studies in combination with bacterial genetics suggested the PhuR receptor is the high capacity uptake receptor, with the HasR receptor acts primarily as a sensor and regulator of heme utilization. The goal of my proposal will be to determine the contributions of the Has and Phu systems to heme sensing and transport through 1) identifying the downstream targets of the HasAp-HasR cell surface signaling system (CSS) and its extra-cytoplasmic function (ECF) sigma factor HasI; and 2) determine the contributions of the HasR and PhuR receptors to heme uptake and utilization. We will utilize a combination of in vitro biochemical and spectroscopic techniques to characterize the molecular mechanism of heme regulation and uptake in combination with in vivo techniques including bacterial genetics and 13C-heme isotopic labeling coupled to LC-MS/MS to determine heme uptake in knockout and complemented strains. At the conclusion of this project I will have determined the regulatory cascade of the Has heme sensing system and the contributions of the non-redundant Has and Phu systems in heme utilization by P. aeruginosa. This study will provide a platform for the identification of novel antimicrobial therapies.

项目总结