A novel preribosomal complex in trypanosomes

锥虫中的一种新型前核糖体复合物

• 批准号: 8247798
• 负责人: Noreen Williams
• 金额: $ 30.94万
• 依托单位: STATE UNIVERSITY OF NEW YORK AT BUFFALO
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-04-01 至 2014-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8247798
• 关键词: African Trypanosomiasis; Antibiotics; Bacteria; Binding; Binding Proteins; Biogenesis; Biological Assay; Cell Nucleolus; Cell Nucleus; Chagas Disease; Chemicals; Complex; Computer Simulation; Data; Databases; Disease; Drug Delivery Systems; Energy Transfer; Fluorescence; Gel; Goals; Homologous Gene; Human; Immune; In Vitro; Intervention; Ketolides; Leishmania; Leishmaniasis; Macrolides; Maintenance; Maps; Mediating; Methods; Molecular; Mutation; Nucleoplasm; Organism; Parasites; Pathway interactions; Play; Process; Protein Binding Domain; Protein Fragment; Proteins; RNA-Binding Proteins; Research; Resolution; Ribosomal Biogenesis Pathway; Ribosomal RNA; Role; Route; Screening procedure; Signal Transduction; Small RNA; Staging; Structure; System; Technology; Testing; Transcription Factor TFIIIA; Trypanosoma; Trypanosoma brucei brucei; Trypanosoma cruzi; Work; base; chemotherapy; domain mapping; in vivo; migration; novel; particle; public health relevance; research study; ribosomal protein L5; trafficking

DESCRIPTION (provided by applicant): We have shown that two trypanosome-specific RNA binding proteins, P34 and P37, play an essential role in the ribosomal biogenesis pathway in T. brucei. This represents a novel finding since the ribosomal pathway is highly conserved. To exploit this observation for the ultimate goal of targeting the T. brucei ribosomal pathway in chemotherapy, we must understand the mechanism by which these proteins act in the pathway. In recent work, we have shown that P34 and P37 interact with both L5 and 5S rRNA, the highly conserved components of the preribosomal particle. Loss of P34/P37 leads to a decrease in 5S rRNA levels and other effects in ribosomal formation. The objective of this proposal is to determine the critical residues involved in this interaction and develop a method to identify molecules that disrupt this interaction. Our hypothesis is that the molecular interactions of these proteins will provide a valid target for intervention in ribosomal assembly in T. brucei. The specific aims of the project are to: 1. Determine the critical molecular determinants on L5 and P34/P37 that are required for participation in the T. brucei preribosomal complex. 2. Determine the critical regions on 5S rRNA that interact with L5 and P34/P37 in the T. brucei preribosomal complex. 3. Determine whether the interaction of P34/P37 with L5 and 5S rRNA is required for trafficking of the complex to the nucleolus. 4. Develop a FRET (fluorescence energy transfer) based screen for the disruption of the interaction between P34/P37, L5, and 5S rRNA in the preribosomal particle. Differences in ribosomal structure and function between bacteria and the infected human host have provided targets for macrolide and ketolide antibiotics. Since this preribosomal particle is unique to kinetoplastids and play an essential role in the ribosomal structure and biogenesis in these organisms, mechanistic studies with this complex will allow us to define a strategic target for chemical intervention. PUBLIC HEALTH RELEVANCE: The protozoan parasites, the Kinetoplastids, include the organisms Trypanosoma brucei, Trypanosoma cruzi and Leishmania, which cause serious diseases in much of the world (African sleeping sickness, Chagas' disease and leishmaniasis, respectively). This study will examine a unique preribosomal complex occurring in these parasites with the goal of developing an assay to identify drugs that target the parasite but not host complex.

描述（由申请人提供）：我们已经证明两种锥虫特异性RNA结合蛋白P34和P37在布氏锥虫的核糖体生物发生途径中发挥重要作用。这是一个新的发现，因为核糖体途径是高度保守的。为了利用这一观察结果实现化疗中靶向布氏锥虫核糖体途径的最终目标，我们必须了解这些蛋白质在该途径中发挥作用的机制。在最近的工作中，我们发现 P34 和 P37 与 L5 和 5S rRNA（前核糖体颗粒的高度保守成分）相互作用。 P34/P37 的缺失会导致 5S rRNA 水平降低以及对核糖体形成的其他影响。该提案的目的是确定参与这种相互作用的关键残基，并开发一种方法来识别破坏这种相互作用的分子。我们的假设是，这些蛋白质的分子相互作用将为干预布氏锥虫核糖体组装提供有效的靶标。该项目的具体目标是： 1. 确定参与布氏锥虫前核糖体复合体所需的 L5 和 P34/P37 的关键分子决定因素。 2. 确定 5S rRNA 上与 T. brucei 前核糖体复合物中的 L5 和 P34/P37 相互作用的关键区域。 3. 确定复合物转运至核仁是否需要 P34/P37 与 L5 和 5S rRNA 的相互作用。 4. 开发基于 FRET（荧光能量转移）的筛选，用于破坏前核糖体颗粒中 P34/P37、L5 和 5S rRNA 之间的相互作用。细菌和受感染的人类宿主之间核糖体结构和功能的差异为大环内酯和酮内酯抗生素提供了靶标。由于这种前核糖体颗粒是动质体所独有的，并且在这些生物体的核糖体结构和生物发生中发挥着重要作用，因此对该复合物的机制研究将使我们能够确定化学干预的战略目标。 公共卫生相关性：原生动物寄生虫动质体包括布氏锥虫、克氏锥虫和利什曼原虫，它们在世界大部分地区引起严重疾病（分别为非洲昏睡病、南美锥虫病和利什曼病）。这项研究将检查这些寄生虫中存在的独特的前核糖体复合物，目的是开发一种测定方法来识别针对寄生虫而不是宿主复合物的药物。