SC3: Regulation of polar adhesion during Sinorhizobium meliloti infection

 描述(申请人提供)：拟议的项目将破译控制α-蛋白细菌粘附素产生和生物膜形成的分子机制，α-蛋白细菌是一种生理异质性的群体，包括许多导致严重人类疾病的病原体。生物膜的形成对细菌在不同环境中的生存起着至关重要的作用，并与公众健康密切相关。生物膜中的细胞顽固地附着在 生物和非生物表面的细菌会对抗菌剂产生更强的抵抗力，并导致持续感染。虽然限制生物膜的形成有可能预防和限制细菌感染，但对α-蛋白细菌的生物膜形成及其如何促进宿主定植知之甚少。最近，我们在驯化的实验室菌株Sinorhizobium Meliloti中发现了保守细胞极性因子PodJ的突变。Sinorhizobium Meliloti是一种模式α-蛋白细菌，通过在豆科植物的根组织中定殖并固定分子氮来换取寄主植物的营养，与亲和的豆科植物建立互惠共生关系。纠正突变并将PodJ极性因子恢复为在环境分离的苜蓿链霉菌中发现的版本，使Rm1021实验室菌株能够产生一种名为Holdfast的极性粘附素，并形成强大的生物膜。此外，校正后的等位基因(podJ+)在共生过程中具有竞争优势。这些新的表型从来没有被表征过，因为常用的苜蓿链霉菌株都具有这种突变。这项提议的目的是阐明连接细胞周期依赖的遗传电路 极性因子PodJ对粘合剂Holdfast的调节生产，促进生物膜的形成和增强适合性，特别是在宿主定植期间。实现这一目标包括实现以下四个具体目标：(1)根据细胞周期确定PodJ和Holdfast的亚细胞位置；(2)评估它们对Holdfast合成和生物膜形成的影响的环境条件；(3)评估Holdfast是否增加了对宿主组织的附着；以及(4)确定和确定参与Holdfast生产的细胞因素的作用。研究结果将提供一个更好的模型，说明保守的和物种特有的途径如何有助于α-蛋白细菌的黏附和生物膜的形成。这些知识将为开发抗击生物膜和细菌感染的新方法提供基础。除了完成科学目标之外 如上所述，资助这项建议将提高研究效率，增强研究人员的竞争力，并使学生，特别是那些来自代表性不足的背景的学生，能够接触到令人兴奋的研究领域，从而促进他们对生物医学职业的追求。

 DESCRIPTION (provided by applicant): The proposed project will decipher the molecular mechanisms that control adhesin production and biofilm formation in alpha-proteobacteria, a physiologically heterogeneous group that includes a number of pathogens that cause serious human diseases. Biofilm formation plays a crucial role in the survival of bacteria in diverse environments and is of great relevance to public health. 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 bacterial infections, little is known about biofilm formation by alpha-proteobacteria and how it facilitates host colonization. Recently, we discovered a mutation in the conserved cell polarity factor PodJ in domesticated, laboratory strains of Sinorhizobium meliloti -- a model alpha-proteobacterium that establishes mutualistic symbioses with compatible legumes by colonizing their root tissues and fixing molecular nitrogen in exchange for nutrients from host plants. Correcting the mutation and restoring the PodJ polarity factor to the version found in environmental isolates of S. meliloti enabled the Rm1021 laboratory strain to produce a polar adhesin, called holdfast, and to develop robust biofilms. Furthermore, the corrected allele (podJ+) conferred a competitive advantage during symbiosis. These novel phenotypes have never been characterized because commonly used S. meliloti strains all possess the mutation. The goal of this proposal is to elucidate the genetic circuitry that links the cell cycle-dependent polarity factor PodJ to the regulated production of adhesive holdfast, which promotes biofilm formation and enhances fitness, particularly during host colonization. Achieving this goal involves accomplishing the following four specific aims: (1) determine the subcellular locations of PodJ and holdfast as a function of the cell cycle; (2) evaluate environmental conditions for their effects on holdfast synthesis and biofilm formation; (3) assess if holdfast increases attachment to host tissue; and (4) identify and define the roles of cellular factors that participae in holdfast production. Results from the investigation will provide a better model of how conserved and species-specific pathways can contribute to adhesion and biofilm formation in alpha-proteobacteria. Such knowledge will provide the foundation for developing new methods to combat biofilms and bacterial infections. 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 exposure to an exciting area of research, thus nurturing their pursuits of biomedical careers.