新加坡南洋理工大学Meng How Tan课题组发现，一种循环置换的CasRx平台可用于高效、特异性RNA编辑。相关论文于2024年10月9日发表于国际顶尖学术期刊《自然—生物技术》杂志上。

研究人员通过合理的蛋白质工程开发了一种优化的RNA编辑平台，即基于CasRx的可编程RNA编辑技术（xPERT）。研究人员展示了通过循环置换CasRx K940L突变体的拓扑重排，构建出一个用于连接脱氨酶结构域的稳固支架。

研究人员将此工具与REPAIR系统进行了对比，结果表明xPERT在靶向活性上与REPAIRv1相当，但具有与REPAIRv2类似的低脱靶编辑特性。xPERT平台可用于改变RNA序列信息，不会带来基因组损伤，能够实现细胞的暂时性变化并定制蛋白功能。

据了解，无活性的Cas13同源物通过其C端与突变的人类ADAR2脱氨酶结构域融合，实现了特定转录本中的可编程腺苷到肌苷（A-to-I）RNA编辑。尽管前景广阔，现有的RNA编辑工具通常在效率与特异性之间存在权衡问题，且脱靶编辑仍未解决。

附：英文原文

Title: A circularly permuted CasRx platform for efficient, site-specific RNA editing

Author: Wang, Yuanming, Liu, Kaiwen Ivy, Liu, Mengying Mandy, Ooi, Kean Hean, Nguyen, Tram Anh, Chee, Jiunn En, Teo, Shun Xiang Danny, He, Shan, Tay, Jie Wen Douglas, Teo, Seok Yee, Liew, Kai Shin, Ge, Xiao Yu, Ng, Zhi Jian, Avagyan, Hasmik, Liu, Hao, Yi, Zirong, Chang, Keziah, Kok, Eng Piew Louis, Chen, Runjia, Yau, Chun En, Koh, Jun Wei, Wan, Yue, Tan, Meng How

Issue&Volume: 2024-10-09

Abstract: Inactive Cas13 orthologs have been fused to a mutant human ADAR2 deaminase domain at the C terminus to enable programmable adenosine-to-inosine (A-to-I) RNA editing in selected transcripts. Although promising, existing RNA-editing tools generally suffer from a trade-off between efficacy and specificity, and off-target editing remains an unsolved problem. Here we describe the development of an optimized RNA-editing platform by rational protein engineering, CasRx-based Programmable Editing of RNA Technology (xPERT). We demonstrate that the topological rearrangement of a CasRx K940L mutant by circular permutation results in a robust scaffold for the tethering of a deaminase domain. We benchmark our tool against the REPAIR system and show that xPERT exhibits strong on-target activity like REPAIRv1 but low off-target editing like REPAIRv2. Our xPERT platform can be used to alter RNA sequence information without risking genome damage, effect temporary cellular changes and customize protein function.

DOI: 10.1038/s41587-024-02430-w

Source: https://www.nature.com/articles/s41587-024-02430-w