Sinorhizobium meliloti factors that affect biofilm formation and symbiosis efficiency

项目摘要/摘要 拟议的项目将阐明控制极地粘附素产生的机制，即生物膜 在α-蛋白细菌中的形成和宿主定植，这是一个生理异质性群体，包括一个 重要的人类病原体的数量。尤其是与公共健康相关的生物膜的形成起着至关重要的作用 细菌在不同环境中生存的作用：生物膜中的细胞顽固地附着在生物和非生物上 表面的细菌会对抗菌剂产生更强的抵抗力，并导致持续性感染。 尽管限制生物膜的形成有可能预防和限制微生物疾病，但人们对此知之甚少。 关于α-变形杆菌生物膜的形成以及它如何促进宿主的定植。 我们之前发现了一种与生物膜相关的突变，这种突变在实验室菌株中很常见。 Meliloti，一种与亲和豆科植物形成互惠共生关系的模式α-蛋白细菌，通过 定植根组织和固定氮素，以换取植物寄主的养分。纠正突变 恢复了保守的极性因子PodJ的全长生产，从而能够合成Holdfast(a 极地粘附素)和强健的生物膜的组装，以前从未观察到的表型。我们发现生物膜- 在宿主感染过程中，优势菌株相对于生物膜缺陷菌株具有竞争优势。通过 转座子突变，我们鉴定了一些与生物膜发育有关的基因，其中包括一个 编码保守的转录因子(LdtR)和编码二鸟苷环化酶的另一种 磷酸二酯酶，已知调节c-di-GMP第二信使的水平。这项提案的目标是 是(A)破译生物膜形成和寄主定居期间关键调节者的责任和(B) 建立共生与环境因素关系监测体系。我们计划 通过实现以下三个具体目标来实现这些目标。(1)我们将确定LdtR如何 表达是受调控的，它执行什么细胞功能。(2)我们将评估如何调节c- DI-GMP水平有助于生物膜的形成和宿主感染。(3)我们将评估共生是否会影响 寄主对有毒化合物的反应以及化学胁迫和其他微生物如何影响寄主共生体 互动。调查结果将提供一个更好的模型，说明细胞和外部因素是如何 有助于α-蛋白细菌的黏附、生物膜形成和宿主入侵。 除了实现上述科学目标外，对这项提案的资助将 提高研究人员的研究效率和竞争力，并允许学生，特别是 来自代表性不足的背景的人，以获得研究培训和为生物医学职业做准备。

Project Summary / Abstract The proposed project will elucidate the mechanisms that control polar adhesin production, biofilm formation, and host colonization in alpha-proteobacteria, a physiologically heterogeneous group that includes a number of significant human pathogens. Particularly relevant to public health, biofilm formation plays a crucial role in the survival of bacteria in diverse environments: cells in biofilms attach recalcitrantly to biotic and abiotic surfaces, develop increased resistance to antimicrobial agents, and contribute to persistent infections. Although limiting biofilm formation has the potential to prevent and restrict microbial diseases, little is known about biofilm formation by alpha-proteobacteria and how it facilitates host colonization. We previously discovered a biofilm-associated mutation common in laboratory strains of Sinorhizobium meliloti, a model alpha-proteobacterium that forms mutualistic symbiosis with compatible legumes by colonizing root tissues and fixing nitrogen in exchange for nutrients from plant hosts. Correcting the mutation restored full-length production of the conserved polarity factor PodJ, thus enabling synthesis of the holdfast (a polar adhesin) and assembly of robust biofilms, phenotypes never observed before. We found that biofilm- competent strains possess a competitive advantage over biofilm-deficient strains during host infection. Via transposon mutagenesis, we identified a number of genes involved in biofilm development, including one encoding a conserved transcription factor (LdtR) and another encoding a diguanylate cyclase and phosphodiesterase, known to modulate levels of the c-di-GMP second messenger. The goals of this proposal are (a) to decipher the responsibilities of key regulators during biofilm formation and host colonization and (b) to establish a system for monitoring the relationship between symbiosis and environmental factors. We plan to achieve these goals by accomplishing the following three specific aims. (1) We will determine how LdtR expression is regulated and what cellular functions it performs. (2) We will assess how factors that modulate c- di-GMP levels contribute to biofilm formation and host infection. (3) We will evaluate whether symbiosis affects host response to toxic compounds and how chemical stress and other microbes influence host-symbiont interactions. Results from the investigation will provide a better model of how cellular and external factors can contribute to adhesion, biofilm formation, and host invasion in alpha-proteobacteria. In addition to accomplishing the scientific objectives described above, funding of this proposal will enhance the research productivity and grant competitiveness of the investigator and allow students, particularly those from underrepresented backgrounds, to gain research training and preparation for biomedical careers.