Mathieu Sébastien, Lesch Elena, Garcia Shahinez, Graindorge Stéfanie, Schallenberg-Rüdinger Mareike, Hammani Kamel
Institut de Biologie Moléculaire des Plantes, Centre National de la Recherche Scientifique (CNRS), Université de Strasbourg, 12 rue du Général Zimmer, 67084 Strasbourg, France.
Institut für Zelluläre und Molekulare Botanik, Abteilung Molekulare Evolution, Universität Bonn, 53115 Bonn, Germany.
Nucleic Acids Res. 2025 Apr 10;53(7). doi: 10.1093/nar/gkaf279.
In plant mitochondria and chloroplasts, cytidine-to-uridine RNA editing is necessary for the production of functional proteins. While natural PLS-type PPR proteins are specialized in this process, synthetic PPR proteins offer significant potential for targeted RNA editing. In this study, we engineered chimeric editing factors by fusing synthetic P-type PPR guides with the DYW cytidine deaminase domain of a moss mitochondrial editing factor, PPR56. These designer PPR editors (dPPRe) elicited efficient and precise de novo RNA editing in Escherichia coli as well as in the chloroplasts and mitochondria of Nicotiana benthamiana. Chloroplast transcriptome-wide analysis of the most efficient dPPRe revealed minimal off-target effects, with only three nontarget C sites edited due to sequence similarity with the intended target. This study introduces a novel and precise method for RNA base editing in plant organelles, paving the way for new approaches in gene regulation applicable to plants and potentially other organisms.
在植物线粒体和叶绿体中，胞嘧啶到尿嘧啶的RNA编辑对于功能性蛋白质的产生是必要的。虽然天然的PLS型PPR蛋白专门参与这一过程，但合成PPR蛋白在靶向RNA编辑方面具有巨大潜力。在本研究中，我们通过将合成的P型PPR引导序列与苔藓线粒体编辑因子PPR56的DYW胞嘧啶脱氨酶结构域融合，构建了嵌合编辑因子。这些设计的PPR编辑器（dPPRe）在大肠杆菌以及本氏烟草的叶绿体和线粒体中引发了高效且精确的从头RNA编辑。对最有效的dPPRe进行叶绿体转录组范围分析发现，脱靶效应极小，由于与预期靶标序列相似，仅三个非靶标C位点被编辑。本研究介绍了一种用于植物细胞器RNA碱基编辑的新颖且精确的方法，为适用于植物及潜在其他生物体的基因调控新方法铺平了道路。
