Guide RNA Binding Complex

摘要 布鲁氏锥虫物种给可以说是最边缘化的人群带来健康危害和经济困难 世界上的人口数量。一些研究最深入的锥虫，锥体也代表了 许多研究领域，包括抗原变异、宿主-病原体相互作用、发育重新编程 和线粒体生物学。该项目将阐明大分子RNA编辑的机制 底物结合复合体(RESC)稳定并传递线粒体前-mRNAs，并引导RNA进入 U-插入/缺失编辑途径，并协调编辑后的mRNAs的多聚腺苷化和翻译。我们 建立RESC平台作为编辑全酶的RNA结合成分并进行研究 它在编辑反应中的作用，以及超越RNA编辑过程的功能。为此，我们证明了 RESC相关的MERS1焦磷水解酶和KPAP1聚(A)聚合酶靶向Pre-mRNA5‘和3’ 分别结束。重要的是，5‘焦磷酸的去除和3’A-拖尾似乎都是Pre-mRNA的关键 编辑前的稳定。相反，建议将RESC中的特定模块与完成 编辑：编辑后3‘A/U-Tail和与核糖体的mRNA结合。总而言之，现有证据 将~25多肽RESC复合体定位为线粒体RNA加工的多模式联系。 此外，与RESC相关的MERS1复合体、RNA聚合酶(MTRNAP)和3‘ Procesome(MPome)挑战多顺反子上颌环转录和转录的长期模式 初级转录本的核酸内切分。拟议的实验将加深对rna的理解。 通过确定近原子分辨率的RESC结构和个体的RNA结合特异性进行编辑 亚单位。我们将测试一个广泛的功能假设，即离散的RESC模块协调mRNA的完成 具有3‘修改和翻译激活的编辑。最后，我们提出了一个全新的概念 由单个启动子转录并由5个ʹ修饰和形成的单顺反子前mRNA 反义RNA控制3‘-5’降解。通过阐明RESC的结构、RNA结合特性和 更高阶的相互作用，并评估范式转换的“单顺反子假说”，这个程序将 扩大关键寄生虫特定过程的知识，并可能提供新的药物靶点。

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.