Molecular Biology and Virulence of CTX Phage

描述（由申请人提供）：移动的遗传元素深刻地影响了细菌病原体的进化。毒力和抗生素耐药性的许多决定因素是由能够水平传播的质粒和噬菌体等可移动元素携带的。在这个资助周期中，我们研究了CTXF生命周期的几个方面，CTXF是一种丝状噬菌体，编码霍乱毒素，霍乱病原体霍乱弧菌的主要毒力因子。我们描述了介导CTXF整合到霍乱弧菌染色体和控制CTXF基因表达的新机制。我们还扩大了我们的工作范围，探索了SXT的分子生物学的几个方面，SXT是一种霍乱弧菌衍生的整合共轭元件（ICE），编码多种抗生素抗性基因。近年来，SXT及其密切相关的元素在霍乱弧菌临床分离株中高度流行。我们的长期目标是破译使CTXF和SXT传播的分子机制。我们拟从5个方面探讨CTXF和SXT的分子生物学特征，以调控它们的传播和进化。在Aim I中阐明控制CTXF抑制因子RstR表达机制的实验将揭示两个宿主因子，LexA和RpoS如何参与控制CTXF产生的新型遗传回路。在Aims II-V中，我们将探索SXT生命周期各方面的分子基础，这些方面将这种移动元件与噬菌体和质粒区分开来。研究SXT的这些新特征将增强我们对ICEs的理解，这是一组广泛且医学上重要的移动元件，尚未受到深入研究，并将揭示导致伽玛变形菌中SXT相关ICEs流行和多样性的过程。特别是，我们将探讨II)限制其转移到新宿主的SXT结合步骤，III)在供体细胞中维持SXT所需的基因，IV)防止SXT向已存在的宿主冗余传播的机制，以及V)产生新的/杂交SXT相关ice的过程。这些研究的完成将增强我们对水平基因转移控制机制的理解，并产生原核细胞生物学和病原体进化的基本方面的知识。此外，这些研究可能为开发新的抗菌剂和疫苗提供有价值的信息。

DESCRIPTION (provided by applicant): Mobile genetic elements have profoundly influenced the evolution of bacterial pathogens. Many determinants of virulence and antibiotic resistance are borne by mobile elements, such as plasmids and bacteriophages, which are capable of horizontal transmission. During this grant cycle, we studied several aspects of the life cycle of CTXF, a filamentous phage that encodes cholera toxin, the principal virulence factor of the cholera pathogen Vibrio cholerae. We described novel mechanisms that mediate the integration of CTXF into the V. cholerae chromosome and that govern CTXF gene expression. We also expanded the scope of our work to explore several aspects of the molecular biology of SXT, a V. cholerae-derived integrating conjugative element (ICE) that encodes multiple antibiotic resistance genes. In recent years, SXT and closely related elements have become highly prevalent in V. cholerae clinical isolates. Our long-term goal is to decipher the molecular mechanisms that enable the dissemination of CTXF and SXT. We propose to explore 5 aspects of the molecular biology of CTXF and SXT that govern their transmission and evolution. Experiments in Aim I to elucidate mechanisms that control expression of RstR, the CTXF repressor, will reveal how two host factors, LexA and RpoS, contribute to the novel genetic circuitry governing CTXF production. In Aims II-V, we will explore the molecular bases for aspects of the SXT life-cycle that distinguish this mobile element from phages and plasmids. Studying these novel features of SXT will enhance our understanding of ICEs, an extensive and medically important set of mobile elements that have not been subject to intensive study, and will shed light on processes that have contributed to the prevalence and diversity of SXT-related ICEs in the gamma proteobacteria. In particular, we will explore II) steps in SXT conjugation that limit its transfer to new hosts, III) genes required for SXT maintenance in donor cells, IV) mechanisms for preventing redundant transmission of SXT to hosts in which it is already present and V) processes by which new/hybrid SXT-related ICEs are generated. Completion of these studies will enhance our understanding of the mechanisms that control of horizontal gene transfer and yield knowledge of fundamental aspects of prokaryote cell biology and pathogen evolution. Furthermore, these studies may provide valuable information for the creation of new antimicrobial agents and vaccines.