Dubey Ashutosh P, Tylec Brianna L, Yi Soon, Tedeschi Frank A, Smith Joseph T, Read Laurie K
Department of Microbiology and Immunology, University at Buffalo Jacobs School of Medicine and Biomedical Sciences, 955 Main Street, Buffalo, NY 14203, USA.
Center for RNA Science and Therapeutics, School of Medicine, Case Western Reserve University, 2109 Adelbert Rd., Cleveland, OH 44106, USA.
Nucleic Acids Res. 2025 Jan 11;53(2). doi: 10.1093/nar/gkae1153.
Uridine insertion/deletion editing of mitochondrial messenger RNAs (mRNAs) in kinetoplastids entails the coordinated action of three complexes. RNA Editing Catalytic Complexes (RECCs) catalyze the enzymatic reactions, while the RNA Editing Substrate Binding Complex (RESC) and RNA Editing Helicase 2 Complex (REH2C) coordinate interactions between RECCs, mRNAs and hundreds of guide RNAs that direct edited sequences. Additionally, numerous auxiliary factors are required for productive editing of specific mRNAs. Here, we elucidate the role of KRBP72, an editing auxiliary factor of the ABC adenosine triphosphatase (ATPase) family that exhibits RNA-binding activity. In procyclic form Trypanosoma brucei, KRBP72 knockdown leads to a pause in editing at the base of a predicted stem loop structure in adenosine triphosphate synthase subunit 6 (A6) mRNA. Enhanced cross-linking and affinity purification revealed KRBP72 binding sites both within and upstream of this stem loop. KRBP72 ATPase activity is essential for its A6 mRNA editing function; however, its RNA-binding activity is dispensable. KRBP72 interacts with most RESC proteins in an RNase-sensitive manner. By contrast, RESC12A associates with KRBP72 in an RNase-insensitive fashion, and RESC12A promotes KRBP72's interaction with RNA. Hence, KRBP72 ATPase activity facilitates progression of editing through a challenging secondary structure, highlighting this protein's crucial role in A6 mRNA editing.
动质体中线粒体信使核糖核酸（mRNA）的尿苷插入/缺失编辑需要三种复合物的协同作用。RNA编辑催化复合物（RECC）催化酶促反应，而RNA编辑底物结合复合物（RESC）和RNA编辑解旋酶2复合物（REH2C）则协调RECC、mRNA和数百种指导RNA之间的相互作用，这些指导RNA指导编辑序列。此外，特定mRNA的有效编辑还需要众多辅助因子。在这里，我们阐明了KRBP72的作用，它是ABC三磷酸腺苷（ATPase）家族的一种编辑辅助因子，具有RNA结合活性。在原循环型布氏锥虫中，敲低KRBP72会导致三磷酸腺苷合酶亚基6（A6）mRNA中预测的茎环结构底部的编辑暂停。增强交联和亲和纯化揭示了该茎环内部和上游的KRBP72结合位点。KRBP72的ATPase活性对其A6 mRNA编辑功能至关重要；然而，其RNA结合活性是可有可无的。KRBP72以对核糖核酸酶敏感的方式与大多数RESC蛋白相互作用。相比之下，RESC12A以对核糖核酸酶不敏感的方式与KRBP72结合，并且RESC12A促进KRBP72与RNA的相互作用。因此，KRBP72的ATPase活性促进编辑通过具有挑战性的二级结构进行，突出了该蛋白在A6 mRNA编辑中的关键作用。
