Engineered circular guide RNAs enhance miniature CRISPR/Cas12f-based gene activation and adenine base editing

IF 15.7 1区综合性期刊 Q1 MULTIDISCIPLINARY SCIENCES Nature Communications Pub Date : 2025-03-28 DOI:10.1038/s41467-025-58367-4

Xin Zhang, Mengrao Li, Kechen Chen, Yuchen Liu, Jiawei Liu, Jiahong Wang, Hongxin Huang, Yanqun Zhang, Tao Huang, Shufeng Ma, Kaitong Liao, Jiayi Zhou, Mei Wang, Ying Lin, Zhili Rong

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Abstract

CRISPR system has been widely used due to its precision and versatility in gene editing. Un1Cas12f1 from uncultured archaeon (hereafter referred to as Cas12f), known for its compact size (529 aa), exhibits obvious delivery advantage for gene editing in vitro and in vivo. However, its activity remains suboptimal. In this study, we engineer circular guide RNA (cgRNA) for Cas12f and significantly improve the efficiency of gene activation about 1.9–19.2-fold. When combined with a phase separation system, the activation efficiency is further increased about 2.3–3.9-fold. In addition, cgRNA enhances the editing efficiency and narrows the editing window of adenine base editing about 1.2–2.5-fold. Importantly, this optimization strategy also boosts the Cas12f-induced gene activation efficiency in mouse liver. Therefore, we demonstrate that cgRNA is able to enhance Cas12f-based gene activation and adenine base editing, which holds great potential for gene therapy.

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工程化的环状引导rna增强了基于CRISPR/ cas12f的微型基因激活和腺嘌呤碱基编辑

CRISPR系统因其在基因编辑中的精确性和通用性而得到了广泛的应用。来自未培养古菌的Un1Cas12f1（以下简称Cas12f）以其紧凑的尺寸（529 aa）而著名，在体外和体内都具有明显的基因编辑传递优势。然而，它的活动仍然不是最优的。在本研究中，我们为Cas12f设计了环状引导RNA (cgRNA)，显著提高了基因激活效率约1.9 - 19.2倍。当与相分离系统结合时，活化效率进一步提高约2.3 - 3.9倍。此外，cgRNA提高了编辑效率，将腺嘌呤碱基编辑的编辑窗口缩小了约1.2 - 2.5倍。重要的是，该优化策略还提高了cas12f诱导的基因在小鼠肝脏中的激活效率。因此，我们证明cgRNA能够增强基于cas12f的基因激活和腺嘌呤碱基编辑，这在基因治疗中具有很大的潜力。

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来源期刊

Nature Communications Biological Science Disciplines-

CiteScore

24.90

自引率

2.40%

发文量

6928

审稿时长

3.7 months

期刊介绍： Nature Communications, an open-access journal, publishes high-quality research spanning all areas of the natural sciences. Papers featured in the journal showcase significant advances relevant to specialists in each respective field. With a 2-year impact factor of 16.6 (2022) and a median time of 8 days from submission to the first editorial decision, Nature Communications is committed to rapid dissemination of research findings. As a multidisciplinary journal, it welcomes contributions from biological, health, physical, chemical, Earth, social, mathematical, applied, and engineering sciences, aiming to highlight important breakthroughs within each domain.