Guide RNA Binding Complex

引导RNA结合复合物

• 批准号: 9352420
• 负责人: Ruslan Afasizhev
• 金额: $ 51.56万
• 依托单位: BOSTON UNIVERSITY MEDICAL CAMPUS
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-05-10 至 2022-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9352420
• 关键词: Antigenic Variation; Antisense RNA; Architecture; Area; Binding; Binding Sites; Biology; Characteristics; Chemicals; Complex; Cryoelectron Microscopy; Crystallization; DNA; DNA-Directed RNA Polymerase; Data; Development; Diphosphates; Ensure; Event; Excision; Exonuclease; Funding; Gap Junctions; Genes; Genetic; Genetic Transcription; Genome; Goals; Guide RNA; Health Hazards; Holoenzymes; Hybrids; Hydrolysis; Individual; Investigation; Kinetoplast DNA; Knowledge; Maps; Mass Spectrum Analysis; Mediator of activation protein; Medical; Messenger RNA; Methods; Mitochondria; Mitochondrial RNA; Modality; Modeling; Modification; Molecular; Parasites; Pathway interactions; Pattern; Polyadenylation; Polynucleotide Adenylyltransferase; Population; Positioning Attribute; Pre-mRNA Polyadenylation Factor; Process; Property; Proteins; RNA; RNA Binding; RNA Editing; RNA Processing; Reaction; Recruitment Activity; Research; Resolution; Ribosomes; Role; Specificity; Structure; System; Tail; Testing; Transcript; Transcription Initiation Site; Translational Activation; Translations; Tropical Disease; Trypanosoma; Trypanosoma brucei brucei; base; crosslink; endonuclease; experimental study; genome-wide analysis; health economics; in vivo; insertion/deletion mutation; mRNA Precursor; mRNA Stability; multimodality; new therapeutic target; novel; pathogen; polypeptide; programs; promoter; reconstitution; sensor; transcriptome; tripolyphosphate

ABSTRACT Trypanosoma brucei species inflict health hazards and economic hardship on arguably the most marginalized populations in the world. Some of the best-studied Excavata, trypanosomes also represent important models in many areas of research, including antigenic variation, host-pathogen interaction, developmental reprogramming and mitochondrial biology. This project will elucidate mechanisms by which macromolecular RNA editing substrate binding complex (RESC) stabilizes and delivers mitochondrial pre-mRNAs and guide RNAs into the U-insertion/deletion editing pathway, and coordinates polyadenylation and translation of edited mRNAs. We establish the RESC platform as the RNA binding constituent of the editing holoenzyme and seek to investigate its role in editing reactions, and functions beyond the RNA editing process. To this end, we demonstrate that RESC-associated MERS1 pyrophosphohydrolase and KPAP1 poly(A) polymerase target pre-mRNA 5′ and 3′ ends, respectively. Importantly, both 5′ pyrophosphate removal and 3′ A-tailing appear to be critical for pre-mRNA stabilization prior to editing. Conversely, specific module within RESC is suggested to couple the completion of editing with post-editing 3′ A/U-tailing and mRNA binding to the ribosome. Collectively, the existing evidence positions the ~25 polypeptide RESC complex as the multimodal nexus of mitochondrial RNA processing. Furthermore, initial investigation of RESC-associated MERS1 complex, RNA polymerase (MTRNAP), and the 3′ processome (MPsome) challenges the long-standing model of multicistronic maxicircle transcription and endonucleolytic partitioning of primary transcripts. The proposed experiments will deepen understanding of RNA editing by determining the RESC structure at near-atomic resolution and RNA binding specificities of individual subunits. We will test a broad functional hypothesis that discrete RESC modules coordinate completion of mRNA editing with 3′ modification and translational activation. Finally, we put forward a fundamentally novel concept of monocistronic pre-mRNAs that are transcribed from individual promoters and shaped by 5ʹ modification and antisense RNA-controlled 3′-5′ degradation. By elucidating the RESC structure, RNA binding properties, and higher-order interactions, and evaluating the paradigm-shifting “monocistronic hypothesis,” this program will expand the knowledge of critical parasite-specific processes and may provide new drug targets.

抽象的 布鲁氏锥虫物种对最边缘化的危害和经济困难造成了健康危害和经济困难 世界上的人口。锥虫中一些最良性的excavata也代表了重要模型 许多研究领域，包括抗原变异，宿主 - 病原体相互作用，发育重编程 和线粒体生物学。该项目将阐明大分子RNA编辑的机制 底物结合复合物（RESC）稳定并递送线粒体前MRNA，并将RNA引导到 U-插入/缺失编辑途径，并坐在编辑mRNA的聚腺苷酸化和翻译。我们 建立救援平台作为RNA结合构成编辑全酶的构成，并试图调查 它在编辑反应中的作用以及超出RNA编辑过程的功能。为此，我们证明了 救援相关的MERS1焦磷酸氢化酶和KPAP1聚（A）聚合酶靶标靶标前MRNA 5'和3' 结束分别。重要的是，5'焦磷酸的去除和3'a尾的外观对于前MRNA至关重要 在编辑之前进行稳定。相反，建议在RESC中进行特定的模块，以将 编辑后编辑3'a/u-tailing和mRNA与核糖体结合。共同证据 将〜25多肽救援复合物定位为线粒体RNA加工的多模式联系。 此外，对救援相关的MERS1复合物，RNA聚合酶（MTRNAP）和3'的初步研究 流程（MPSOME）挑战多发性分布性最大转录的长期模型和 主要成绩单的内核分解分区。提出的实验将加深对RNA的理解 通过在接近原子分辨率和个人的RNA结合规格下确定救援结构来编辑 亚基。我们将检验一个广泛的功能假设，即离散救援模块坐标mRNA的完成 用3'修改和翻译激活进行编辑。最后，我们提出了一个根本新颖的概念 从单个启动子转录并根据5°修改和形状的单概主义者前MRNA和 反义RNA控制的3'-5'降解。通过阐明救援结构，RNA结合特性和 高阶相互作用，并评估范式转移的“单科式假设”，该程序将 扩大关键寄生虫特异性过程的知识，并可能提供新的药物靶标。