Molecular mechanisms and benefits of phenotypic heterogeneity in Sinorhizobium meliloti populations

苜蓿中华根瘤菌群体表型异质性的分子机制和益处

• 批准号: 218318381
• 负责人: Professorin Dr. Anke Becker
• 金额: --
• 依托单位: Institut für Mikrobiologie und Molekularbiologie
• 依托单位国家: 德国
• 项目类别: Priority Programmes
• 财政年份: 2012
• 资助国家: 德国
• 起止时间: 2011-12-31 至 2020-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/218318381?language=en
• 关键词: Molecular; mechanisms; benefits; phenotypic; heterogeneity

Phenotypic heterogeneity is prevalent in the microbial world, especially in biofilms. We describe a case using colonies of the alpha-proteobacterium bacterium Sinorhizobium meliloti. A sub-population strongly expresses genes specific for exopolysaccharide (EPS) production, while their genetically identical neighbors exhibit either very little or no expression. Although the genetic pathways controlling EPS are relatively well understood, such as quorum sensing, little is known about how multiple phenotypes with respect to EPS production arise. To understand this phenomenon, we have used an interdisciplinary approach, integrating mathematical modeling, genetic engineering, live cell imaging, and data processing analyses. The approach relies upon a triple-reporter construct which is capable of carrying three different promoters, each fused to a dedicated fluorescent reporter gene. This provides a method whereby the growth of a colony from a single cell can be monitored, and each individual can be systematically classified with regards to its ancestral lineage and its contributor status. We classify each individual into one of three classes: non-contributor, weak contributor, and strong contributor.On the basis of this platform, we outline a research program to address the question of how heterogeneity arises in the regulatory circuitry controlling EPS production. The program focuses on several aspects of regulatory circuits essential for bi-stability. One is the presence of mechanisms capable of generating a heterogenic signal (e.g., hemi-methylation of promoter regions, and the stress response), and another is mechanisms capable of heterogenic signal amplification (e.g., positive feedback loops). Both features have been found in the regulatory circuitry controlling EPS production in S. meliloti. These will be integrated in discriminatory theoretical models which are testable in the laboratory. Another important question is the biological impact of phenotypic heterogeneity. To answer this, we make use of our recent work which compares a mutant (displaying a non-contributor phenotype) to the wild type. The mutant grew faster and was more motile, while the wild type was better at survival of harsh physical conditions. In this program, we propose a method to obtain populations that are selected for a single class by fusing the promoters controlling EPS production to positive and negative selection cassettes. This will allow the conditional enrichment of single-class populations without further genetic perturbation and allow characterization of the contributor and non-contributor classes. Finally, we ask the question of how the cooperative/contributor phenotypes of S. meliloti can be favored in the context of evolution and develop a model addressing this question.

表型异质性在微生物世界中普遍存在，特别是在生物膜中。我们描述了一种情况下使用的α-变形杆菌属细菌苜蓿中华根瘤菌菌落。一个亚群强烈表达胞外多糖（EPS）生产特异性基因，而它们的遗传上相同的邻居表现出非常少或没有表达。虽然控制EPS的遗传途径相对较好地理解，如群体感应，但对EPS产生的多种表型如何产生知之甚少。为了理解这一现象，我们使用了跨学科的方法，整合了数学建模，遗传工程，活细胞成像和数据处理分析。该方法依赖于三重报告构建体，其能够携带三个不同的启动子，每个启动子融合到专用的荧光报告基因。这提供了一种方法，通过该方法可以监测来自单个细胞的集落的生长，并且每个个体可以关于其祖先谱系及其贡献者状态进行系统分类。我们将每个人分为三类：非贡献者，弱贡献者，强contributor.On这个平台的基础上，我们概述了一个研究计划，以解决如何异质性出现在控制EPS生产的监管电路的问题。该计划侧重于对双稳态至关重要的监管电路的几个方面。一个是存在能够产生异源信号的机制（例如，启动子区域的半甲基化，和应激反应），另一个是能够异源信号放大的机制（例如，正反馈回路）。这两个特点已被发现在调控电路控制EPS生产在S。苜蓿草。这些将被纳入可在实验室测试的歧视性理论模型。另一个重要的问题是表型异质性的生物学影响。为了回答这个问题，我们利用我们最近的工作，比较突变体（显示非贡献者表型）的野生型。突变体生长得更快，更有活力，而野生型在恶劣的物理条件下生存得更好。在这个程序中，我们提出了一种方法，以获得人口选择为一个单一的类融合的启动子控制EPS生产的积极和消极的选择盒。这将允许有条件地富集单类群体而不进一步遗传扰动，并允许表征贡献者和非贡献者类。最后，我们问的问题是如何合作/贡献表型的S。草木犀可以在进化的背景下，并制定一个解决这个问题的模型。