Experimental and theoretical investigations of gene regulation by chromosomal topological domains in E. coli

细菌是地球上最简单的生命形式之一。尽管它们相对简单，但每个细菌细胞仍然是一个复杂的生命系统，能够对环境的变化做出精确的反应。在过去的70年里，已经发现了描述这种调节如何发生的细节，但是我们的理解仍然存在差距。新的研究揭示了细菌细胞利用染色体DNA结构全局控制基因调控的新机制的证据。这种机制长期以来被认为是细菌生命形式所缺失的。研究人员将使用综合实验和计算建模方法探索这种以前未知的调节机制。项目目标的成功完成将有助于理解细菌系统中基因调控的进化。作为该项目更广泛影响的一部分，研究人员将参与研究生的跨学科培训，为高中生提供研究经验，并为巴尔的摩地区内城社区的小学生提供暑期课程。该项目的目的是研究染色体拓扑结构域在调节大肠杆菌基因表达中的作用。该项目将结合单细胞成像、生化分析和这些拓扑结构域的理论建模，以揭示它们对基因表达的全局调控作用。细菌遗传信息处理的经典教条只关注通过蛋白质（如RNA聚合酶和转录因子）识别DNA和/或RNA序列的机制来调节基因的转录活性。近年来越来越多的研究表明，染色体DNA的超卷曲状态是影响细菌基因表达的另一个基本因素。染色体DNA的拓扑结构在个体结构域之间的扩散被禁止，在基因调控中必须发挥重要作用。然而，关于大肠杆菌中染色体拓扑结构域是如何组织的，以及它们如何影响基因表达的知识仍然难以捉摸。在这个项目中，两位首席研究员将结合他们在大肠杆菌染色体组织的实验分析和细菌基因调控网络的理论建模方面的专业知识，以解决三个具体目标：(1)研究是否以及如何使用单细胞mRNA和蛋白质生产成像方法协调同一拓扑结构域中多个基因的表达；(2)基因组尺度上染色体拓扑结构域的基因调控模式；(3)确定染色体拓扑组织与基因表达谱的相关性。提出的工作结果将导致拓扑知情基因调控网络模型的发展。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

Bacteria are one of the simplest forms of life on Earth. Despite their relative simplicity, each single bacterial cell is still a sophisticated living system capable of precisely responding to changes in its environment. Over the last 70 years details describing of how this regulation occurs have been uncovered, however there is still gaps in our understanding. New research is uncovering evidence of a novel mechanism employed by bacterial cells to globally control the regulation of genes using the structure of chromosomal DNA. This mechanism was long thought to be missing from bacterial life forms. Investigators will explore this previously unknown regulatory mechanism using an integrated experimental and computational modeling approach. Successful completion of the project goals will contribute to the understanding of the evolution of gene regulation in bacterial systems. As part of the broader impacts of the project, investigators will engage in interdisciplinary training of graduate students, research experience for high school students, and summer programs for elementary students within inner city communities in the Baltimore area.The objective of the project is to investigate the role of chromosomal topological domains in regulating gene expression in E. coli. The project will combine single-cell imaging, biochemical analysis, and theoretical modeling of these topological domains to uncover their global regulatory effect on gene expression. The classic dogma of bacterial genetic information processing focuses on the regulation of a gene's transcriptional activity only through the mechanism of DNA and/or RNA sequence recognition by proteins such as RNA polymerase and transcription factors. In recent years an increasing number of studies have shown that the supercoiling state of chromosomal DNA is another fundamental factor impacting gene expression in bacteria. The topological organization of chromosomal DNA into individual domains, between which the diffusion of supercoiling is prohibited, must play an important role in gene regulation. However, knowledge of how chromosomal topological domains are organized in E. coli, and how they impact gene expression, remains elusive. In this project the two principal investigators will combine their expertise in the experimental analyses of E. coli chromosomal organization and theoretical modeling of bacterial gene regulatory networks to address three specific aims: (1) Investigate whether and how the expression of multiple genes within the same topological domain is coordinated using single cell mRNA and protein production imaging methods; (2) Model gene regulation by chromosomal topological domains at the genome scale; and (3) Determine the correlation between chromosomal topological organization and gene expression profile. Results of the proposed work will result in the development of a topologically-informed gene regulatory network model.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.