RNA-DNA hybrid binding domain broadens the editing window of base editors.

IF 12.1 1区医学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Molecular Therapy Pub Date : 2025-06-14 DOI:10.1016/j.ymthe.2025.06.024

Yue Yang,Zihao Fu,Shengcheng Deng,Guanglan Wu,Chuanle Wang,Xiao Luo,Rui Kang,Yuxi Chen,Chengxiang Peng,Pengfei Zhang,Kaixin Cui,Fen Wan,Junhua Wang,Qin Zhou,Wei Chen,Yuanyan Xiong,Wenbin Ma,Zhou Songyang,Puping Liang

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引用次数: 0

Abstract

Adenine base editors (ABEs) and cytosine base editors (CBEs) are prominent tools for precise genome editing but are hindered by limited editing activity at positions proximal to the protospacer adjacent motif (PAM). This study investigates the potential of enhancing base editors editing activity by fusing them with RNA-DNA hybrid binding domains (RHBDs). Specifically, fusing ABE8e with the RHBD of Homo sapiens RNaseH1 (RHBD1) significantly increased A-to-G editing efficiency in the PAM-proximal region (A9-A15) by up to 3.5-fold, while reducing off-target cytosine editing. Additionally, RHBD1 is compatible with ABEmax, BE4max, and dual base editor (eA&C-BEmax), enhancing their editing activity at the PAM-proximal bases. Notably, RHBD1-fused BE4max led to a 3.1-fold improvement in C-to-T editing efficiency at PAM-proximal region (C9-C12). Furthermore, we demonstrated that RHBD1-fused ABE8e could effectively edit disease-related single nucleotide variations (SNVs) in human cells, and validated its efficacy in adult mouse liver. These findings highlight the significance of the RHBD in expanding editing window and the applicability of base editors for gene therapy and disease modeling.

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RNA-DNA杂交结合域拓宽了碱基编辑器的编辑窗口。

腺嘌呤碱基编辑器（ABEs）和胞嘧啶碱基编辑器（CBEs）是精确基因组编辑的重要工具，但由于原间隔器邻近基序（PAM）附近的编辑活性有限而受到阻碍。本研究探讨了通过将碱基编辑器与RNA-DNA杂交结合域（rhbd）融合来增强碱基编辑器编辑活性的潜力。具体而言，将ABE8e与现代人RNaseH1 （RHBD1）的RHBD融合后，pam -近端区域（A9-A15）的A-to-G编辑效率显著提高了3.5倍，同时减少了脱靶胞嘧啶编辑。此外，RHBD1与ABEmax、BE4max和双碱基编辑器（eA&C-BEmax）兼容，增强了它们在pam -近端碱基上的编辑活性。值得注意的是，rhbd1融合的BE4max使pam -近端区C-to-T编辑效率提高了3.1倍（C9-C12）。此外，我们证明了rhbd1融合的ABE8e可以有效地编辑人类细胞中疾病相关的单核苷酸变异（snv），并验证了其在成年小鼠肝脏中的有效性。这些发现突出了RHBD在扩展编辑窗口方面的重要性，以及碱基编辑器在基因治疗和疾病建模方面的适用性。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Molecular Therapy 医学-生物工程与应用微生物

CiteScore

19.20

自引率

3.20%

发文量

357

审稿时长

3 months

期刊介绍： Molecular Therapy is the leading journal for research in gene transfer, vector development, stem cell manipulation, and therapeutic interventions. It covers a broad spectrum of topics including genetic and acquired disease correction, vaccine development, pre-clinical validation, safety/efficacy studies, and clinical trials. With a focus on advancing genetics, medicine, and biotechnology, Molecular Therapy publishes peer-reviewed research, reviews, and commentaries to showcase the latest advancements in the field. With an impressive impact factor of 12.4 in 2022, it continues to attract top-tier contributions.