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Kinetoplastid RNA editing

动质体RNA编辑

基本信息

批准号：
10200087
负责人：
Laurie K. Read
金额：
$ 35.04万
依托单位：
STATE UNIVERSITY OF NEW YORK AT BUFFALO
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-07-13 至 2024-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10200087
关键词：
Address Affinity Chromatography Antibodies Binding Bioenergetics Bioinformatics Blood Circulation Co-Immunoprecipitations Complex Data Defect Development Developmental Process Discrimination Essential Genes Foundations Future Generations Goals Growth Guide RNA Heterogeneity High-Throughput Nucleotide Sequencing Holoenzymes Human Immunoprecipitation In Vitro Insect Vectors Insecta Knowledge Laboratories Left Length Life Life Cycle Stages Mass Spectrum Analysis Mediator of activation protein Medical Messenger RNA Mitochondria Mitochondrial Proteins Modeling Molecular Nucleotides Organism Parasites Pattern Play Poly A Positioning Attribute Process Proteins Publishing RNA RNA Binding RNA Editing RNA Interference Regulation Regulator Genes Resolution Ribonucleoproteins Role Site System Tail Technology Testing Trypanosoma Trypanosoma brucei brucei Uridine Virulence Work bioinformatics tool drug development enzyme mechanism innovation insertion/deletion mutation insight knock-down mitochondrial messenger RNA mutant novel protein complex protein function protein protein interaction recruit scaffold tool trafficking transcriptome

项目摘要

RNA editing in kinetoplastid parasites entails massive remodeling of mitochondrial mRNAs by posttranscriptional insertion and deletion of uridine (U) residues to generate mRNAs encoding proteins involved in parasite bioenergetics. U insertion/deletion RNA editing is unique to kinetoplastids and essential for T. brucei survival and virulence. The RNA editing holoenzyme comprises two subcomplexes: RECC (RNA Editing Core Complex) and RESC (RNA Editing Substrate Binding Complex). Trans-acting guide RNAs (gRNAs) direct U insertion/deletion, acting sequentially such that editing proceeds 3' to 5' along an mRNA. RECC contains the editing enzymes, and the mechanisms by which RECC catalyzes U insertion/deletion at a single editing site have been extensively studied. More recently, RESC has emerged as the scaffold for editing and a coordinator of interactions between mRNAs, gRNAs, and RECC, although the mechanisms by which it accomplishes these tasks are essentially unknown. Adding even more complexity, editing of specific mRNAs is differentially regulated between human bloodstream form (BF) and insect vector procyclic form (PF) parasites. The mechanism(s) of developmental regulation, and the precise point(s) in the life cycle at which they are effected, remain entirely mysterious. The goals of this application are to (1) delineate the roles of distinct components of RESC and a novel RESC-related complex in gRNA trafficking and utilization, and (2) to define aspects of the editing process that serve as control points for stage specific regulation during the T. brucei life cycle. To accomplish these goals, we developed an innovative and powerful bioinformatic tool (HTS/TREAT) that permits us to now examine the mitochondrial transcriptome at single nucleotide resolution, and group partially edited mRNA intermediates in ways that illuminate specific aspects of gRNA usage. Combining this bioinformatic approach with RNA immunoprecipitation and protein-protein interaction studies, we will define the mechanisms by which specific RESC proteins function, correlate these to RESC heterogeneity and dynamic interactions, and elucidate the steps in the editing process that are developmentally controlled. We will also take advantage of recent technology that permits generation of substantial numbers of insect metacyclic form (MF) T. brucei, a largely unexplored intermediate between PF and BF. Using HTS/TREAT, we will define the patterns of RNA editing in MFs, and reveal whether BF RNA editing patterns for some or all mRNAs emerge in MFs, providing insight into the temporal control of RNA editing during T. brucei development. The proposed studies address several important gaps in knowledge regarding the essential kinetoplastid RNA editing process. Moreover, this work may help lay the foundation for future drug development and provide important insights into ribonucleoprotein complex function in higher organisms.

动质体寄生虫中的RNA编辑导致线粒体mRNAs的大规模重塑，尿苷（U）残基的转录后插入和缺失以产生编码蛋白质的mRNA 与寄生虫生物能量学有关U插入/缺失RNA编辑是动质体所特有的，并且对于 T.布鲁氏菌存活率和毒力。RNA编辑全酶包括两个亚复合物：RECC（RNA 编辑核心复合物（RNA Editing Core Complex）和RESC（RNA Editing Substrate Binding Complex）。反式作用向导RNA （gRNA）指导U插入/缺失，顺序地起作用，使得编辑沿着mRNA从3'沿着5'进行。 RECC含有编辑酶，以及RECC催化U插入/缺失的机制。单个编辑位点已被广泛研究。最近，RESC已经成为编辑的支架以及mRNA、gRNA和RECC之间相互作用的协调者，尽管它的机制是完成这些任务基本上是未知的。更复杂的是，特定mRNA的编辑是在人类血流形式（BF）和昆虫媒介原环形式（PF）寄生虫之间差异调节。发育调节的机制，以及它们在生命周期中的精确点影响，保持完全神秘。本应用程序的目标是（1）描述不同的 RESC的组成部分和一种新的RESC相关复合物在gRNA运输和利用，和（2）定义编辑过程的各个方面，作为T.布鲁氏菌病周期为了实现这些目标，我们开发了一个创新的和强大的生物信息学工具（HTS/TREAT）这使得我们现在可以在单核苷酸分辨率下检查线粒体转录组，部分编辑的mRNA中间体，以阐明gRNA使用的特定方面。组合该生物信息学方法与RNA免疫沉淀和蛋白质-蛋白质相互作用的研究，我们将定义特异性RESC蛋白发挥功能的机制，将这些与RESC异质性和动态互动，并阐明在编辑过程中的步骤，是发展控制。我们还将利用允许产生大量昆虫元环形式的最新技术 (MF)T.布氏杆菌，PF和BF之间的一个基本上未开发的中间体。使用HTS/TREAT，我们将定义在MF中的RNA编辑模式，并揭示是否BF RNA编辑模式的一些或所有的mRNA出现在 MF，提供了对T.布氏发展拟议研究解决了关于基本动质体RNA编辑知识的几个重要空白，过程此外，这项工作可能有助于为未来的药物开发奠定基础，深入了解核糖核蛋白复合体在高等生物中的功能。

项目成果

期刊论文数量（6）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Developmental dynamics of mitochondrial mRNA abundance and editing reveal roles for temperature and the differentiation-repressive kinase RDK1 in cytochrome oxidase subunit II mRNA editing.

DOI：
10.1128/mbio.01854-23
发表时间：
2023-10-31
期刊：
mBio
影响因子：
6.4
作者：
通讯作者：

High throughput sequencing revolution reveals conserved fundamentals of U-indel editing.

高通量测序革命揭示了 U-indel 编辑的保守基础。

DOI：
10.1002/wrna.1487
发表时间：
2018
期刊：
Wiley interdisciplinary reviews. RNA
影响因子：
0
作者：
Zimmer,SaraL;Simpson,RachelM;Read,LaurieK
通讯作者：
Read,LaurieK

Trypanosome RNAEditing Substrate Binding Complex integrity and function depends on the upstream action of RESC10.

DOI：
10.1093/nar/gkab129
发表时间：
2021-04-06
期刊：
Nucleic acids research
影响因子：
14.9
作者：
Dubey AP;Tylec BL;McAdams NM;Sortino K;Read LK
通讯作者：
Read LK

Intrinsic and regulated properties of minimally edited trypanosome mRNAs.

最低限度编辑的锥虫 mRNA 的内在和调控特性。

DOI：
10.1093/nar/gkz012
发表时间：
2019
期刊：
Nucleic acids research
影响因子：
14.9
作者：
Tylec,BriannaL;Simpson,RachelM;Kirby,LauraE;Chen,Runpu;Sun,Yijun;Koslowsky,DonnaJ;Read,LaurieK
通讯作者：
Read,LaurieK

Developmental regulation of edited CYb and COIII mitochondrial mRNAs is achieved by distinct mechanisms in Trypanosoma brucei.

布氏锥虫中编辑的 CYb 和 COIII 线粒体 mRNA 的发育调节是通过不同的机制实现的。

DOI：
10.1093/nar/gkaa641
发表时间：
2020
期刊：
Nucleic acids research
影响因子：
14.9
作者：
SmithJr,JosephT;Doleželová,Eva;Tylec,Brianna;Bard,JonathanE;Chen,Runpu;Sun,Yijun;Zíková,Alena;Read,LaurieK
通讯作者：
Read,LaurieK