CAREER: The Molecular Basis of Persister Cell and Biofilm Formation by the E. Coli Protein MqsR

职业生涯：大肠杆菌蛋白 MqsR 形成持久细胞和生物膜的分子基础

• 批准号: 0952550
• 负责人: Rebecca Page
• 金额: $ 83.75万
• 依托单位: Brown University
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-04-01 至 2016-03-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0952550&HistoricalAwards=false
• 关键词: CAREER; Molecular; Basis; Persister; Cell

Intellectual Merit:Research on biofilms, bacterial communities familiar to everyone as they coat our teeth at night, has been ongoing for many years. However, even today, the only certain way to remove biofilms is by mechanical force, i.e. a toothbrush. While brushing our teeth is routine, removal of biofilms from ships, pipes and medical devices, and other surfaces, is much more difficult and expensive. The formation of biofilms is one of the major defense and survival mechanisms utilized by bacteria. However, a detailed understanding of how biofilms assemble and are regulated at a molecular level is only rudimentarily understood. The formation of bacterial persisters, a genetically identical sub-population of metabolically quiescent cells that express protein toxins and exhibit multidrug tolerance, is at the core of biofilm formation. However, persistence is also one of the most poorly understood mechanisms used by bacteria to survive environmental stress. Recently, the Escherichia coli protein MqsR (B3022, YgiU) was identified as a key persistence factor, as it is the most highly upregulated gene in persisters. Because its sequence does not resemble that of any characterized protein, its molecular function, i.e. how it is regulated at a molecular level and especially how it drives the formation of the persister phenotype, is currently unknown. Accordingly, the long-term objective of this CAREER project is to elucidate the molecular mechanisms that lead to the persister state, with a focus on understanding the protein activities that lead to persister formation. In addition, a thorough understanding of how these activities are regulated under normal and stressful conditions and how they can be blocked for the development of novel agents that inhibit the formation of the persister cell phenotype will be determined. Specifically, the following questions will be answered: 1) What is the 3-dimensional structure, and thus the function, of the MqsR toxin and how is its toxicity mitigated by its interaction with MqsA (B3021, YgiT)? 2) How do MqsA and the MqsR:MqsA complex differentially regulate E. coli gene transcription? and 3) How does MqsR toxicity lead to biofilm and persister cell formation? Taken together, these studies will define the molecular mechanism of MqsR and provide essential new insights into how MqsR controls bacterial persistence and biofilm formation.Broader impacts:Biofilms, complex communities of bacteria that are highly resistant to antimicrobials and cost the world economy billions of dollars every year, are extraordinarily enriched in persister cells. A molecular understanding of the function and regulation of the proteins that play a key role in persistence, like MqsR, will provide novel targets needed for development of new chemical agents that target biofilms. In this CAREER project, multiple research disciplines, including structural biology, biochemistry, and genetics, will be integrated to provide projects for both undergraduate and graduate students that reveal the interdisciplinary nature of scientific research. The cornerstone of the educational program is a multi-component Protein Science Workshop (PSW) that will provide Community College of Rhode Island (CCRI) students with the opportunity to expand their research experiences. It is composed of lectures and laboratories at Brown University and RI-EPSCoR facilities and will expose the CCRI students to state-of-the-art research and technologies. In addition, each year, one CCRI student will be invited to carry-out their own independent research project. The long-term goal of the collaboration with CCRI, the first of its kind in Rhode Island, is to attract and develop a new generation of scientists from the often forgotten pool of adult students who are returning to school in order to obtain the knowledge and skills that will allow them to pursue higher education and/or entry into the biotechnology workforce.

知识分子的优点：关于生物膜的研究，每个人在晚上涂牙齿时熟悉的细菌群落已经进行了很多年。但是，即使在今天，去除生物膜的唯一某种方法是通过机械力，即牙刷。虽然刷牙是常规的，但从船只，管道和医疗设备以及其他表面上除去生物膜，要困难和昂贵得多。生物膜的形成是细菌使用的主要防御和生存机制之一。然而，只有一定会理解对生物膜如何在分子水平组装和调节的详细理解。生物膜形成的核心是细菌持久的形成，是表达蛋白质毒素并表现出多药耐受性的代谢静态细胞的遗传相同的亚群体的形成。但是，持久性也是细菌用于生存环境压力的最知名度之一。最近，大肠杆菌蛋白MQSR（B3022，YGIU）被确定为关键持久因子，因为它是持久性最高上调的基因。由于其序列与任何特征蛋白质的序列不像其分子功能，即在分子水平上如何调节其序列，尤其是它如何驱动持久表型的形成方式，目前尚不清楚。因此，该职业项目的长期目标是阐明导致持久状态的分子机制，重点是理解导致持久形成的蛋白质活性。此外，将确定将确定抑制持久细胞表型形成的新型药物的开发，对这些活性如何受到调节，以及如何阻止它们。具体而言，将回答以下问题：1）MQSR毒素的3维结构，因此是什么功能，以及它与MQSA（B3021，YGIT）的相互作用如何减轻其毒性？ 2）MQSA和MQSR：MQSA复合物如何差异调节大肠杆菌基因转录？ 3）MQSR毒性如何导致生物膜并持久细胞形成？综上所述，这些研究将定义MQSR的分子机制，并为MQSR如何控制细菌持久性和生物膜形成提供了重要的新见解。Broader的影响：生物膜，复杂的细菌群落，这些群体具有高度抵抗的抗菌剂，并且对世界经济的抗菌抗菌和代价损失了数十亿美元的年限，每年都在典型地富裕地富有繁荣的细胞。像MQSR这样的蛋白质的功能和调节的分子理解，例如MQSR，将提供开发靶向生物膜的新化学剂所需的新靶标。在这个职业项目中，包括结构生物学，生物化学和遗传学在内的多个研究学科，将融入到本科和研究生的项目，以揭示科学研究的跨学科性质。教育计划的基石是一个多组分的蛋白质科学研讨会（PSW），它将为罗德岛社区学院（CCRI）学生提供扩大研究经验的机会。它由布朗大学和RI-EPSCOR设施的讲座和实验室组成，将使CCRI学生接触到最先进的研究和技术。此外，每年都会邀请一名CCRI学生进行自己的独立研究项目。与CCRI合作的长期目标是罗德岛州的第一个同类产品，是从经常被遗忘的成年学生群中吸引和发展新一代的科学家，这些成年学生正在返回学校，以获取知识和技能，以获取使他们能够接受高等教育和/或进入生物技术劳动力的知识和技能。