Analysis of Protein Synthesis in Bacterial Persisters

细菌存留物中蛋白质合成的分析

• 批准号: 9017852
• 负责人: DAVID A TIRRELL
• 金额: $ 22.45万
• 依托单位: CALIFORNIA INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-01-01 至 2017-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9017852
• 关键词: Address; Amino Acids; Antibiotic Therapy; Antibiotics; Automobile Driving; Cell Survival; Cells; Chronic; Clinical; Communities; Exhibits; Exposure to; Genes; Growth; Heterogeneity; Infection; Knock-out; Label; Lead; Ligase; Mass Spectrum Analysis; Methionine-tRNA Ligase; Methods; Microbe; Phase; Phenotype; Population; Protein Biosynthesis; Proteins; Proteome; Proteomics; Pseudomonas aeruginosa; Recovery; Resuscitation; Running; Time; Variant; antimicrobial; design; interest; killings; metabolic rate; mutant; pathogen; pathogenic bacteria; prevent; programs; promoter; public health relevance; resistant strain; response; tool

 DESCRIPTION (provided by applicant): We propose to develop methods to enable proteomic analysis of rare persister cells in phenotypically heterogeneous bacterial populations. Isogenic bacterial populations are characterized by phenotypic heterogeneity that includes variations in metabolic rates and responses to antibiotic treatment. Upon exposure to antibiotics, most bacterial cells die. But in many cases, a small subpopulation (usually < 0.1%) persists and, upon relief of antibiotic challenge, resumes growth. These "persister cells" have been observed for a wide variety of microbes treated with many types of antibiotics. The ability of pathogenic bacteria to persist and recover following antimicrobial therapy leads to chronic infections and the emergence of resistant strains. Understanding the mechanisms that enable persistence would constitute an important step toward treating and preventing chronic infections, but because studies of persister cells require analysis of small subpopulations of non-growing (or very slowly growing) cells, characterization of persisters has been difficult. We propose to develop and evaluate a general strategy for selective study of these cells at the proteomic level. Specifically we will use bio-orthogonal non-canonical amino acid tagging (BONCAT) and quantitative mass spectrometry to establish the time-dependent proteomic profiles of persister cells before, during, and upon recovery from antibiotic challenge. We aim to address the following questions: 1. How do persister cells respond to antibiotic challenge? 2. How do persister cells initiate growth following antibiotic challenge? and 3. What makes the persister cell subpopulation different from the antibiotic-susceptible majority? More generally, we will establish bioanalytical methods of broad utility in the study of bacterial persistence, chronic infection, and rare sub-populations in heterogeneous bacterial communities.