Mechanisms and regulation of replication, the cell cycle, gene expression, and horizontal gene transfer in prokaryotes, focusing on Bacillus subtilis

原核生物复制、细胞周期、基因表达和水平基因转移的机制和调控，重点关注枯草芽孢杆菌

• 批准号: 9896667
• 负责人: ALAN D GROSSMAN
• 金额: $ 72.53万
• 依托单位: MASSACHUSETTS INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-04-01 至 2022-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9896667
• 关键词: Antibiotic Resistance; Bacillus subtilis; Bacteria; Cell Cycle Progression; Cell physiology; Cells; Chromosomes; Coupling; DNA biosynthesis; Elements; Environment; Evolution; Frequencies; Gene Expression; Gene Transfer; Generations; Genes; Genome; Goals; Gram-Positive Bacteria; Growth; Homologous Protein; Horizontal Gene Transfer; Human Microbiome; In Vitro; Mediating; Metabolism; Methodology; Mobile Genetic Elements; Molecular; Organism; Pathogenesis; Plasmids; Population; Prevalence; Process; Prokaryotic Cells; Regulation; Replication Initiation; Signal Transduction; Stress; Symbiosis; Tetracycline Resistance; Virus; Work; cdc Genes; chromosome replication; driving force; genetic manipulation; in vivo; insight; interest; microbial; pathogen; prevent; response; trait; transcription factor; uncontrolled cell growth

Our long term goals are to understand many of the molecular mechanisms and regulation underlying basic cellular processes in bacteria. We are particularly interested in the control of DNA replication, cellular responses to perturbations in replication, environmental sensing and signal transduction, and mechanisms controlling horizontal gene transfer. Our organism of choice for these studies is the Gram positive bacterium Bacillus subtilis. There is a wealth of information about B. subtilis. It is easy to grow and manipulate genetically and is related to several pathogens and environmental bacteria that are less tractable experimentally. Horizontal gene transfer (HGT) is the driving force in microbial evolution. It is largely mediated by mobile genetic elements, including viruses, conjugative plasmids, and integrative and conjugative elements (ICEs, also known as conjugative transposons). Conjugative elements are well known agents contributing to the spread of genes for antibiotic resistances, pathogenesis, symbiosis, metabolism, and more. Despite the prevalence and importance of ICEs, there are deficiencies in our understanding of these elements, especially in Gram positive bacteria. Our work will focus on the lifecycle of ICEBs1 in B. subtilis. The ability to experimentally induce ICEBs1 in >90% of cells in a population and achieve relatively high conjugation frequencies will allow us to answer previously difficult or unstudied problems fundamental to the ICE lifecycle. We will also identify and analyze host genes needed for ICE function and study interactions between ICEs and other mobile elements in cells. We will extend our analyses to Tn916, a widespread ICE involved in the spread of tetracycline resistance. Our findings should be relevant to the transfer of genes between bacteria growing in many different environments, including the human microbiome. Our work will also focus on several aspects of chromosome dynamics and gene expression. Cells have multiple mechanisms for regulating the initiation of replication. Cells also have regulatory responses to perturbations in replication, often called checkpoints, which control gene expression and cell cycle progression. Coupling gene expression and cell cycle progression to chromosome replication and integrity helps prevent the generation of cells with defective chromosomes. The coordination of genome duplication with cell cycle progression is important for cellular differentiation and preventing uncontrolled cell growth. Microbial pathogenesis often depends on normal bacterial replication and growth in the host. We will use a variety of approaches and methodologies, both in vivo and in vitro, to characterize: the control of replication initiation; regulators of the replication initiator and transcription factor DnaA; and genes controlled in response to perturbations in replication. Understanding these processes in B. subtilis will provide insights regarding similar processes and homologous proteins in a wide variety of organisms, including many Gram positive pathogens.

我们的长期目标是了解许多基本的分子机制和调控 细菌中的细胞过程。我们对 DNA 复制的控制、细胞 对复制、环境感知和信号转导中的扰动的反应以及机制 控制水平基因转移。我们为这些研究选择的生物体是革兰氏阳性细菌 枯草芽孢杆菌。关于枯草芽孢杆菌有大量的信息。它很容易生长和基因操纵 并且与实验上不易处理的几种病原体和环境细菌有关。 水平基因转移（HGT）是微生物进化的驱动力。很大程度上是通过移动设备介导的 遗传元件，包括病毒、接合质粒以及整合和接合元件（ICE、 也称为接合转座子）。共轭元件是众所周知的有助于 抗生素抗性、发病机制、共生、代谢等基因的传播。 尽管 ICE 很普遍且很重要，但我们对这些的理解还存在缺陷。 元素，尤其是革兰氏阳性细菌中的元素。我们的工作将重点关注枯草芽孢杆菌中 ICEBs1 的生命周期。这 能够在群体中 >90% 的细胞中通过实验诱导 ICEBs1 并实现相对较高的接合 频率将使我们能够回答以前困难或未研究的 ICE 生命周期的基本问题。 我们还将识别和分析 ICE 功能所需的宿主基因，并研究 ICE 和 细胞中的其他移动元件。我们将把我们的分析扩展到 Tn916，这是一种参与传播的广泛 ICE 四环素耐药性。我们的发现应该与生长在细菌之间的基因转移有关 许多不同的环境，包括人类微生物组。 我们的工作还将重点关注染色体动力学和基因表达的几个方面。细胞有 调节复制起始的多种机制。细胞也有调节反应 复制过程中的扰动，通常称为检查点，控制基因表达和细胞周期进程。 将基因表达和细胞周期进程与染色体复制和完整性结合起来有助于防止 产生带有缺陷染色体的细胞。基因组复制与细胞周期的协调 进展对于细胞分化和防止不受控制的细胞生长很重要。微生物 发病机制通常取决于宿主中细菌的正常复制和生长。 我们将使用各种体内和体外方法和方法来表征： 控制复制启动；复制起始子和转录因子 DnaA 的调节因子；和基因 控制以响应复制中的扰动。了解枯草芽孢杆菌中的这些过程将提供 关于多种生物体中相似过程和同源蛋白质的见解，包括许多 革兰氏阳性病原体。