EMBL Grenoble, 71 Avenue des Martyrs, 38042 Grenoble, France.
EMBL Heidelberg, Structural and Computational Biology Unit, Meyerhofstraße 1, 69117 Heidelberg, Germany.
Nucleic Acids Res. 2023 May 22;51(9):4602-4612. doi: 10.1093/nar/gkad217.
Kinetoplastid parasites, such as trypanosomes or leishmania, rely on RNA-templated RNA editing to mature mitochondrial cryptic pre-mRNAs into functional protein-coding transcripts. Processive pan-editing of multiple editing blocks within a single transcript is dependent on the 20-subunit RNA editing substrate binding complex (RESC) that serves as a platform to orchestrate the interactions between pre-mRNA, guide RNAs (gRNAs), the catalytic RNA editing complex (RECC), and a set of RNA helicases. Due to the lack of molecular structures and biochemical studies with purified components, neither the spacio-temporal interplay of these factors nor the selection mechanism for the different RNA components is understood. Here we report the cryo-EM structure of Trypanosoma brucei RESC1-RESC2, a central hub module of the RESC complex. The structure reveals that RESC1 and RESC2 form an obligatory domain-swapped dimer. Although the tertiary structures of both subunits closely resemble each other, only RESC2 selectively binds 5'-triphosphate-nucleosides, a defining characteristic of gRNAs. We therefore propose RESC2 as the protective 5'-end binding site for gRNAs within the RESC complex. Overall, our structure provides a starting point for the study of the assembly and function of larger RNA-bound kinetoplast RNA editing modules and might aid in the design of anti-parasite drugs.
动基体原生动物寄生虫，如锥虫或利什曼原虫，依赖 RNA 模板的 RNA 编辑将线粒体隐匿的前体 mRNA 成熟为有功能的蛋白编码转录本。单个转录本中多个编辑块的连续编辑依赖于 20 亚基 RNA 编辑底物结合复合物 (RESC)，它作为一个平台来协调前体 mRNA、指导 RNA (gRNA)、催化 RNA 编辑复合物 (RECC) 和一组 RNA 解旋酶之间的相互作用。由于缺乏与纯化成分的分子结构和生化研究，这些因素的时空相互作用以及不同 RNA 成分的选择机制尚不清楚。在这里，我们报告了布鲁氏锥虫 RESC1-RESC2 的 cryo-EM 结构，这是 RESC 复合物的中心枢纽模块。该结构揭示了 RESC1 和 RESC2 形成必需的结构域交换二聚体。尽管两个亚基的三级结构非常相似，但只有 RESC2 选择性地结合 5'-三磷酸核苷，这是 gRNA 的一个定义特征。因此，我们提出 RESC2 是 RESC 复合物中 gRNA 的保护性 5'-末端结合位点。总的来说，我们的结构为更大的 RNA 结合的动基体 RNA 编辑模块的组装和功能研究提供了一个起点，并可能有助于设计抗寄生虫药物。
