Simpson Rachel M, Bruno Andrew E, Chen Runpu, Lott Kaylen, Tylec Brianna L, Bard Jonathan E, Sun Yijun, Buck Michael J, Read Laurie K
Department of Microbiology and Immunology, University at Buffalo Jacobs School of Medicine and Biomedical Sciences, 3435 Main Street, Buffalo, NY 14214, USA.
Center for Computational Research, University at Buffalo, 701 Ellicott St., Buffalo, NY 14203, USA.
Nucleic Acids Res. 2017 Jul 27;45(13):7965-7983. doi: 10.1093/nar/gkx458.
Uridine insertion/deletion RNA editing is an essential process in kinetoplastid parasites whereby mitochondrial mRNAs are modified through the specific insertion and deletion of uridines to generate functional open reading frames, many of which encode components of the mitochondrial respiratory chain. The roles of numerous non-enzymatic editing factors have remained opaque given the limitations of conventional methods to interrogate the order and mechanism by which editing progresses and thus roles of individual proteins. Here, we examined whole populations of partially edited sequences using high throughput sequencing and a novel bioinformatic platform, the Trypanosome RNA Editing Alignment Tool (TREAT), to elucidate the roles of three proteins in the RNA Editing Mediator Complex (REMC). We determined that the factors examined function in the progression of editing through a gRNA; however, they have distinct roles and REMC is likely heterogeneous in composition. We provide the first evidence that editing can proceed through numerous paths within a single gRNA and that non-linear modifications are essential, generating commonly observed junction regions. Our data support a model in which RNA editing is executed via multiple paths that necessitate successive re-modification of junction regions facilitated, in part, by the REMC variant containing TbRGG2 and MRB8180.
尿苷插入/缺失RNA编辑是动基体寄生虫中的一个重要过程，通过该过程，线粒体mRNA通过尿苷的特异性插入和缺失进行修饰，以产生功能性开放阅读框，其中许多编码线粒体呼吸链的组成部分。鉴于传统方法在探究编辑进展的顺序和机制以及单个蛋白质的作用方面存在局限性，许多非酶编辑因子的作用仍不明确。在这里，我们使用高通量测序和一种新型生物信息学平台——锥虫RNA编辑比对工具（TREAT），研究了部分编辑序列的整个群体，以阐明三种蛋白质在RNA编辑介导复合物（REMC）中的作用。我们确定所研究的因子在通过引导RNA（gRNA）进行编辑的过程中发挥作用；然而，它们具有不同的作用，并且REMC的组成可能是异质的。我们提供了首个证据，表明编辑可以通过单个gRNA内的多条路径进行，并且非线性修饰是必不可少的，会产生常见的连接区域。我们的数据支持一种模型，即RNA编辑通过多条路径执行，这些路径需要对连接区域进行连续的重新修饰，这在一定程度上由含有TbRGG2和MRB8180的REMC变体促进。
