美国加州大学旧金山分校Alexis C. Komor课题组开发出多路复用正交碱基编辑器（MOBE）系统。相关论文于2024年5月21日发表在《自然—生物技术》杂志上。

研究人员利用RNA aptamer-coat蛋白系统将DNA修饰酶直接招募到向导RNA 上，从而设计出可以正交复用的腺嘌呤碱基编辑器（BE）和胞嘧啶BE。研究人员生成了四种复用正交BE系统，无需富集或选择策略，即可在同一DNA链中实现高达7.1%的精确同时发生编辑率。

在人类细胞中，添加荧光富集策略可将共现编辑率提高到24.8%。这些系统与扩展的原位相邻基序和高保真Cas9变体兼容，在多种类型的细胞中功能良好，与亲代系统相比具有同等或更低的脱靶倾向，并能模拟疾病相关的点突变组合。

据了解，BE可以高效、可编程地安装点突变，同时避免使用双链断裂。同时应用两种或两种以上不同的碱基编辑器（如腺嘌呤碱基编辑器可将A-T碱基对转换为G-C，以及胞嘧啶碱基编辑器，可将C-G碱基对转换为T-A）是不可行的，因为想到RNA的串扰会导致非正交编辑，即所有碱基编辑器都会修改所有目标基因座。

附：英文原文

Title: Development of multiplexed orthogonal base editor (MOBE) systems

Author: Cowan, Quinn T., Gu, Sifeng, Gu, Wanjun, Ranzau, Brodie L., Simonson, Tatum S., Komor, Alexis C.

Issue&Volume: 2024-05-21

Abstract: Base editors (BEs) enable efficient, programmable installation of point mutations while avoiding the use of double-strand breaks. Simultaneous application of two or more different BEs, such as an adenine BE (which converts A·T base pairs to G·C) and a cytosine BE (which converts C·G base pairs to T·A), is not feasible because guide RNA crosstalk results in non-orthogonal editing, with all BEs modifying all target loci. Here we engineer both adenine BEs and cytosine BEs that can be orthogonally multiplexed by using RNA aptamer–coat protein systems to recruit the DNA-modifying enzymes directly to the guide RNAs. We generate four multiplexed orthogonal BE systems that enable rates of precise co-occurring edits of up to 7.1% in the same DNA strand without enrichment or selection strategies. The addition of a fluorescent enrichment strategy increases co-occurring edit rates up to 24.8% in human cells. These systems are compatible with expanded protospacer adjacent motif and high-fidelity Cas9 variants, function well in multiple cell types, have equivalent or reduced off-target propensities compared with their pa

DOI: 10.1038/s41587-024-02240-0

Source: https://www.nature.com/articles/s41587-024-02240-0