Assembling an understanding of ribosome biosynthesis

描述（由申请人提供）：翻译的基本和必要过程是由核糖体催化的，本提案侧重于核糖体的生物发生。核糖体是复杂的RNPs，其生物发生涉及rRNA的转录、加工、修饰和折叠以及r蛋白和至少40种组装因子的动态结合。长期以来，核糖体一直是了解复杂RNPs功能能力的基准，而揭示30S亚基生物发生机制的工作将同样增加我们对所有细胞RNPs生物发生的理解。我们的长期目标是获得参与大肠杆菌核糖体亚基生物发生的因素和途径的详细了解。由于细菌和真核核糖体的生物发生有许多共同的特征，这些研究的结果无疑将为真核核糖体的生物发生提供新的保守特征和新的见解。此外，由于核糖体生物发生可能在细菌中高度保守，并且与某些细菌的致病性有关，因此发现细菌特异性生物发生事件将产生许多新的可能的抗菌靶点，并可能对医学操作改变致病菌生长的可能方法产生新的见解。我们的方法包括分离体内形成的中间体，随后分析这些颗粒的成分、结构和加工过程。初步结果表明存在三个不同的复合物，至少30个潜在的组装因子，其中三个我们已经确认为组装因子。我们将利用这些数据进一步研究核糖体的生物发生，并确定这一过程中的重要步骤。我们将使用生物化学，遗传学，分子生物学和蛋白质组学来研究核糖体亚基生物发生中间体和参与这些过程的因素，这些数据将用于开发细菌中这一过程重要的相互作用网络。这些研究不仅会产生抗生素筛选和见解的新靶点

DESCRIPTION (provided by applicant): The fundamental and essential process of translation is catalyzed by the ribosome, and this proposal focuses on biogenesis of the ribosome. Ribosomes are intricate RNPs, whose biogenesis involves the transcription, processing, modification and folding of rRNA and the dynamic binding of r-proteins and at least 40 assembly factors. Ribosomes have long been the benchmark for understanding functional capabilities of complex RNPs, and work to unravel the mechanisms of 30S subunit biogenesis will similarly increase our understanding of biogenesis of all cellular RNPs. Our long-term goal is to gain a detailed understanding of the factors and pathways involved in E. coli ribosomal subunit biogenesis. Since bacterial and eukaryotic ribosome biogenesis share several features, results from these studies will undoubtedly lead to new conserved features and new insights into biogenesis of eukaryotic ribosomes. Furthermore, since ribosome biogenesis is likely to be highly conserved among bacteria and is linked to pathogenicity of some bacteria, discoveries of bacterial- specific biogenesis events will yield a number of new possible antimicrobial targets and likely new insights into possible methodologies for medical manipulation to alter growth of pathogenic bacteria. Our approach involves the isolation of in vivo formed intermediates and subsequent analysis of the components, structures, and processing of these particles. Preliminary results have shown the existence of three distinct complexes, at least 30 potential assembly factors, three of which we have already confirmed as assembly factors. We will use these data to further interrogate ribosome biogenesis and to identify important steps in this process. We will use biochemistry, genetics, molecular biology and proteomics to study ribosomal subunit biogenesis intermediates and factors that are involved in these processes and these data will be used to develop a network of interactions important for this process in bacteria. These studies will not only result in novel targets for antibiotic screening and insights into bacterial drug resistance, but will also yield better understanding of the evolution of ribosome biogenesis, and the interplay of biogenesis with fundamental aspects of cell physiology.