New genes involved in cellular responses to quinolone treatment

参与细胞对喹诺酮治疗反应的新基因

• 批准号: 7467361
• 负责人: Xilin Zhao
• 金额: $ 19.13万
• 依托单位: UNIV OF MED/DENT OF NJ-NJ MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-07-15 至 2010-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7467361
• 关键词: Affect; Amino Acid Sequence; Antitoxins; Apoptosis; Attention; Bacteria; Basic Science; Binding Sites; Biochemical; Biochemical Reaction; Biochemistry; Biological Assay; Cell Death; Cells; Cessation of life; Chloramphenicol; Complex; Condition; Cyclic AMP-Dependent Protein Kinases; DNA; DNA Gyrase; DNA Repair; Defect; Depressed mood; Development; Down-Regulation; Drug Formulations; Drug resistance; Enhancers; Escherichia coli; Excision; Exposure to; Gene Expression; Gene Expression Profiling; Genes; Genetic; Genomics; Goals; Government; Growth; Heat-Shock Response; High temperature of physical object; Hydrogen Peroxide; Infection; Investigation; Knock-out; Lactams; Lead; Libraries; Link; Maps; Measures; Mediating; Molecular Profiling; Mutation; Nalidixic Acid; Oxidative Stress; Pathway interactions; Patients; Persons; Pharmaceutical Preparations; Phosphorylation; Phosphotransferases; Physiological; Predisposition; Process; Promoter Regions; Property; Protein Kinase; Protein Overexpression; Proteins; Purpose; Quinolones; Recovery; Regulation; Resistance; Role; Sequence Analysis; Site; Stress; System; Tetracycline; Tetracyclines; Thinking; Time; Toxin; Work; antimicrobial; antimicrobial drug; bacterial resistance; bactericide; biological adaptation to stress; cell killing; design; env Gene Products; florfenicol; genetic regulatory protein; high throughput screening; improved; inhibitor/antagonist; interest; killings; kinase inhibitor; mutant; programs; protein purification; response; small molecule; stressor; ultraviolet irradiation; uptake; Affect; Amino Acid Sequence; Antitoxins; Apoptosis; Attention; Bacteria; Basic Science; Binding Sites; Biochemical; Biochemical Reaction; Biochemistry; Biological Assay; Cell Death; Cells; Cessation of life; Chloramphenicol; Complex; Condition; Cyclic AMP-Dependent Protein Kinases; DNA; DNA Gyrase; DNA Repair; Defect; Depressed mood; Development; Down-Regulation; Drug Formulations; Drug resistance; Enhancers; Escherichia coli; Excision; Exposure to; Gene Expression; Gene Expression Profiling; Genes; Genetic; Genomics; Goals; Government; Growth; Heat-Shock Response; High temperature of physical object; Hydrogen Peroxide; Infection; Investigation; Knock-out; Lactams; Lead; Libraries; Link; Maps; Measures; Mediating; Molecular Profiling; Mutation; Nalidixic Acid; Oxidative Stress; Pathway interactions; Patients; Persons; Pharmaceutical Preparations; Phosphorylation; Phosphotransferases; Physiological; Predisposition; Process; Promoter Regions; Property; Protein Kinase; Protein Overexpression; Proteins; Purpose; Quinolones; Recovery; Regulation; Resistance; Role; Sequence Analysis; Site; Stress; System; Tetracycline; Tetracyclines; Thinking; Time; Toxin; Work; antimicrobial; antimicrobial drug; bacterial resistance; bactericide; biological adaptation to stress; cell killing; design; env Gene Products; florfenicol; genetic regulatory protein; high throughput screening; improved; inhibitor/antagonist; interest; killings; kinase inhibitor; mutant; programs; protein purification; response; small molecule; stressor; ultraviolet irradiation; uptake

DESCRIPTION (provided by applicant): The long-term goal of this program is to understand bacterial stress responses in enough detail to eventually design small-molecule inhibitors of stress response as antimicrobial potentiators. The present R21 application describes a basic-science, proof-of-principle project to study a new protein kinase that when inactivated decreases survival of E. coli cells treated with a variety of antimicrobials, hydrogen peroxide, and high temperature. A deficiency of the kinase reduces bacterial survival to quinolone treatment by 10- to 100-fold and causes the bacteriostatic compound chloramphenicol to become bactericidal. It also dramatically lowers the ability of nalidixic acid to induce new resistant mutants. The kinase deficiency is suppressed by deletion of a toxin-antitoxin gene pair thought to contribute both to protection from stress and to bacterial apoptosis. This genetic interaction with toxin-antitoxin systems leads to the hypothesis that the kinase normally limits toxin activity; in the absence of the kinase, toxins kill cells during stressful conditions, thereby enhancing the action of many antimicrobials at the same time. The kinase gene is also implicated in the Cpx envelope protein stress response pathway by having two CpxR binding sites upstream of its promoter region, establishing another link of the kinase to stress responses. Both the genetics and biochemistry of this stress-response kinase will be studied. The upstream regulation of the kinase gene will be studied through effects of mutations in the Cpx and other related two-component regulatory systems, and downstream effects will be studied through genetic interactions with the toxin-antitoxin systems. To obtain a framework for the role of the kinase in stress response networks, gene expression profiling will also be carried out with a variety of stresses in the presence/absence of the kinase activity. The kinase has been purified. As a part of its further characterization, the enzymatic reaction conditions will be optimized, the autophosphorylation site(s) will be determined, and proteins it normally phosphorylates will be identified. The proposed work constitutes an early characterization of regulatory networks involved in bacterial stress response, persistence/tolerance, and apoptosis. Such studies may eventually lead to ways for making many antimicrobials more effective by interfering with bacterial stress responses. Bacterial resistance, tolerance, and persistence to antimicrobial treatment is a growing threat for our ability to cure infections. Protective genes involved in bacterial stress responses help bacteria evade and survive antimicrobial treatment. These protective stress response networks will be studied with the long-term goal of developing small-molecule inhibitors for antimicrobial enhancement.

描述（由申请人提供）：该程序的长期目标是了解足够细节的细菌应力反应，以最终将应力反应的小分子抑制剂设计为抗菌剂量增强剂。目前的R21应用描述了一个基本科学，原则项目，用于研究一种新的蛋白激酶，当灭活时，该蛋白质激酶会降低用多种抗菌剂，过氧化氢和高温处理的大肠杆菌细胞的存活率。激酶的缺乏可将细菌的生存率降低到喹诺酮类治疗，从而使1-100倍，并导致抑菌化合物化合物氯霉素变为杀菌性。它还大大降低了奈替酸诱导新的抗性突变体的能力。通过缺失毒素 - 抗毒素基因对抑制激酶缺乏症，这既有助于保护胁迫和细菌凋亡。这种与毒素 - 抗毒素系统的遗传相互作用导致了以下假设：激酶通常会限制毒素活性。在没有激酶的情况下，毒素在压力条件下杀死细胞，从而同时增强许多抗菌剂的作用。激酶基因还通过在其启动子区域上游的两个CPXR结合位点具有CPX包膜蛋白应力响应途径与CPX蛋白质应激响应途径有关，从而建立了激酶与应激反应的另一种联系。将研究这种应激反应激酶的遗传学和生物化学。激酶基因的上游调节将通过CPX和其他相关的两组分组系统中突变的影响进行研究，并将通过与毒素 - 抗毒素系统的遗传相互作用来研究下游效应。为了获得激酶在应力反应网络中的作用的框架，在存在/不存在激酶活性的情况下，还将以多种应力进行基因表达分析。激酶已被纯化。作为其进一步表征的一部分，将优化酶促反应条件，确定自磷酸化位点，并确定通常磷酸化的蛋白质。提出的工作构成了与细菌应力反应，持久性/耐受性和凋亡有关的调节网络的早期表征。这样的研究最终可能导致通过干扰细菌应激反应来使许多抗菌药物更有效的方法。 细菌耐药性，耐受性和对抗菌治疗的持久性是我们治愈感染能力的日益威胁。参与细菌胁迫反应的保护基因有助于细菌逃避和存活抗菌治疗。将研究这些保护性应力反应网络的长期目标，即开发用于抗菌增强的小分子抑制剂。