德国图宾根大学Thorsten Stafforst研究团队，利用摆动增强的环状CLUSTER引导RNA进行体内RNA碱基编辑。2024年7月12日，国际知名学术期刊《自然—生物技术》在线发表了这一成果。

研究人员发现，在主导旁观者编辑的5′-UAN三联体中，G•U摆动碱基对能够有效减轻脱靶事件，同时保持高效的目标编辑。该策略可普遍适用于现有的腺苷到肌苷（A-to-I）RNA碱基编辑系统，并补充了其他抑制方法，如G•A错配和尿苷（U）减少。

将摆动碱基配对与环化的CLUSTER方法结合使用，研究人员在细胞培养中对Mecp2转录本的相关疾病突变，实现了高精度和高效率的编辑（高达87%）。通过病毒介导的引导RNA单独递送，在小鼠Rett综合征模型的中枢神经系统中实现了功能性MeCP2蛋白的恢复，编辑效率高达19%，并在体内实现了优异的旁观者控制。

据悉，通过使用定制的引导RNA招募内源性RNA腺苷脱氨酶（ADAR），进行A-to-I的RNA碱基编辑，在RNA水平上安全操纵遗传信息具有很大前景。然而，编辑的精确性和效率常因旁系脱靶编辑而受到影响。

附：英文原文

Title: Precise in vivo RNA base editing with a wobble-enhanced circular CLUSTER guide RNA

Author: Reautschnig, Philipp, Fruhner, Carolin, Wahn, Nicolai, Wiegand, Charlotte P., Kragness, Sabrina, Yung, John F., Hofacker, Daniel T., Fisk, Jenna, Eidelman, Michelle, Waffenschmidt, Nils, Feige, Maximilian, Pfeiffer, Laura S., Schulz, Annika E., Fll, Yvonne, Levanon, Erez Y., Mandel, Gail, Stafforst, Thorsten

Issue&Volume: 2024-07-12

Abstract: Recruiting the endogenous editing enzyme adenosine deaminase acting on RNA (ADAR) with tailored guide RNAs for adenosine-to-inosine (A-to-I) RNA base editing is promising for safely manipulating genetic information at the RNA level. However, the precision and efficiency of editing are often compromised by bystander off-target editing. Here, we find that in 5′-UAN triplets, which dominate bystander editing, GU wobble base pairs effectively mitigate off-target events while maintaining high on-target efficiency. This strategy is universally applicable to existing A-to-I RNA base-editing systems and complements other suppression methods such as GA mismatches and uridine (U) depletion. Combining wobble base pairing with a circularized format of the CLUSTER approach achieves highly precise and efficient editing (up to 87%) of a disease-relevant mutation in the Mecp2 transcript in cell culture. Virus-mediated delivery of the guide RNA alone realizes functional MeCP2 protein restoration in the central nervous system of a murine Rett syndrome model with editing yields of up to 19% and excellent bystander control in vivo.

DOI: 10.1038/s41587-024-02313-0

Source: https://www.nature.com/articles/s41587-024-02313-0