当前位置: X-MOL 学术Nat. Chem. Biol. › 论文详情
Our official English website, www.x-mol.net, welcomes your feedback! (Note: you will need to create a separate account there.)
Engineering a precise adenine base editor with minimal bystander editing
Nature Chemical Biology ( IF 12.9 ) Pub Date : 2022-10-13 , DOI: 10.1038/s41589-022-01163-8
Liang Chen 1 , Shun Zhang 1 , Niannian Xue 1 , Mengjia Hong 1 , Xiaohui Zhang 1 , Dan Zhang 1 , Jing Yang 1 , Sijia Bai 1 , Yifan Huang 1 , Haowei Meng 2 , Hao Wu 2 , Changming Luan 1 , Biyun Zhu 1 , Gaomeng Ru 1 , Hongyi Gao 1 , Liping Zhong 3 , Meizhen Liu 1 , Mingyao Liu 1 , Yiyun Cheng 1 , Chengqi Yi 2 , Liren Wang 1 , Yongxiang Zhao 3 , Gaojie Song 1 , Dali Li 1
Affiliation  

Adenine base editors (ABEs) catalyze A-to-G transitions showing broad applications, but their bystander mutations and off-target editing effects raise safety concerns. Through structure-guided engineering, we found ABE8e with an N108Q mutation reduced both adenine and cytosine bystander editing, and introduction of an additional L145T mutation (ABE9), further refined the editing window to 1–2 nucleotides with eliminated cytosine editing. Importantly, ABE9 induced very minimal RNA and undetectable Cas9-independent DNA off-target effects, which mainly installed desired single A-to-G conversion in mouse and rat embryos to efficiently generate disease models. Moreover, ABE9 accurately edited the A5 position of the protospacer sequence in pathogenic homopolymeric adenosine sites (up to 342.5-fold precision over ABE8e) and was further confirmed through a library of guide RNA–target sequence pairs. Owing to the minimized editing window, ABE9 could further broaden the targeting scope for precise correction of pathogenic single-nucleotide variants when fused to Cas9 variants with expanded protospacer adjacent motif compatibility. bpNLS, bipartite nuclear localization signals.



中文翻译:


以最少的旁观者编辑设计精确的腺嘌呤碱基编辑器



腺嘌呤碱基编辑器 (ABE) 催化 A 到 G 的转变显示出广泛的应用,但它们的旁观者突变和脱靶编辑效应引起了安全问题。通过结构引导工程,我们发现具有 N108Q 突变的 ABE8e 减少了腺嘌呤和胞嘧啶旁观者编辑,并引入了额外的 L145T 突变 (ABE9),进一步将编辑窗口细化为 1-2 个核苷酸,消除了胞嘧啶编辑。重要的是,ABE9 诱导了非常少量的 RNA 和不可检测的独立于 Cas9 的 DNA 脱靶效应,这主要是在小鼠和大鼠胚胎中安装所需的单个 A 到 G 的转换,以有效地生成疾病模型。此外,ABE9 准确地编辑了致病性同聚腺苷位点中原型间隔序列的 A 5位点(精度比 ABE8e 高达 342.5 倍),并通过引导 RNA-靶序列对文库得到进一步证实。由于编辑窗口最小化,当与具有扩展的原型间隔子相邻基序兼容性的 Cas9 变体融合时,ABE9 可以进一步扩大靶向范围,以精确校正致病性单核苷酸变体。 bpNLS,二分核定位信号。

更新日期:2022-10-14
down
wechat
bug