Assembling an understanding of Ribosome Biogenesis

摘要/摘要：核糖体是多组分细胞RNPs中最保守、最具特征的一种。虽然人们对核糖体的结构和功能了解很多，但仍有许多细节有待阐明，特别是关于核糖体组装及其对核糖体功能能力的影响。因此，我们研究大肠杆菌小核糖体亚基（SSUs）的生物合成不仅对理解这些颗粒的组装和功能，而且对理解其他RNPs的动态组装和功能具有重要影响。此外，大肠杆菌产生的总能量中约有40%用于合成核糖体成分。这一观察结果和大肠杆菌生长速率与核糖体生物发生之间的直接关系表明，准确组装和功能核糖体对细胞活力至关重要。我们的长期目标是获得对核糖体生物发生的详细了解，从而揭示这一动态过程对细胞生理学基本方面的影响，如生长调节。本提案详细介绍了大肠杆菌SSU生物发生的研究。这项工作的目的是深入了解构象变化以及在功能SSU组装过程中调节这些变化的因素。为解决这些问题，提出了三个具体目标。首先，将确定16S rRNA内部的构象变化，以及在体外和体内不同组装阶段促进这些变化的特定rRNA核苷酸和核糖体蛋白。其次，利用遗传学和生物化学的结合，将鉴定和表征新的SSU组装中间体。第三，分析有利于SSU装配的辅助因素。将详细探讨在当前资助期内确定的因素，并将确定和描述其他因素。我们在当前资助周期中的进展和我们的初步数据表明，所有三个具体目标都将成功，因此将直接影响对SSU生物发生的理解，并大大扩展对rna -蛋白质相互作用和RNP组装的理解。通过揭示对SSUs的组装和功能至关重要的方面，我们可以确定抗菌作用的新靶点。鉴于抗生素耐药菌株的日益流行，发现这些靶点对人类健康至关重要。

DESCRIPTION (provided by applicant): Summary/Abstract: The ribosome is one the most ubiquitously conserved and best characterized of the multi- component cellular RNPs. While much is known about ribosome structure and function, there are still many details to be elucidated especially regarding ribosome assembly and its impact the functional capacity of ribosomes. Thus, our work to address biosynthesis of E. coli small ribosomal subunits (SSUs) will have a significant impact on understanding not only assembly and function of these particles but also on the dynamic assembly and function of other RNPs. Moreover approximately 40% of the total energy production of E. coli is consumed to synthesize ribosomal components. This observation and the direct correlation between the rates of E. coli growth and ribosome biogenesis indicate that an accurately assembled and functional ribosome is of utmost importance for cell viability. Our long term goal is to gain a detailed understanding of ribosome biogenesis and thus reveal the impact of this dynamic process on fundamental aspects of cell physiology, such as growth regulation. This proposal details studies of E. coli SSU biogenesis. The purpose of this work is to gain an in-depth understanding of conformational changes and the factors that modulate these changes during the course of functional SSU assembly. Three specific aims have been proposed to address these questions. First, conformational changes within 16S rRNA and specific rRNA nucleotides and ribosomal proteins that facilitate these changes during different stages of assembly in vitro and in vivo will be identified. Second, using a combination of genetics and biochemistry, novel SSU assembly intermediates will be identified and characterized. Third, auxiliary factors that facilitate SSU assembly will be analyzed. Factors identified in the current funding period will be explored in detail and additional factors will be identified and characterized. Our progress during the current funding cycle and our preliminary data indicate that all three specific aims will be successful and thus will directly impact understanding of SSU biogenesis and greatly expand understanding of RNA-protein interactions and RNP assembly in general. By revealing aspects that are critical for both assembly and function of SSUs, we may identify novel target sites for antimicrobial action. Given the growing prevalence of antibiotic resistant bacterial strains, the discovery of such targets is critical for human health. PUBLIC HEALTH RELEVANCE: Project Narrative: Known antibiotics share common targets and chemical moieties and given the prevalence of multi-drug resistant bacteria, the identification of additional antibiotics and targets is critical for human health. It has been proposed that bacterial ribosome biogenesis represents promising new ground for development of antimicrobials and thus our studies of ribosome biogenesis in E. coli may result in the identification of novel targets for antibiotic action and may aid in alleviating the problem of bacterial drug resistance as a health concern.