Glycosylase base editors enable C-to-A and C-to-G base changes.

Nature Biotechnology
Dongdong ZhaoXueli Zhang

Abstract

Current base editors (BEs) catalyze only base transitions (C to T and A to G) and cannot produce base transversions. Here we present BEs that cause C-to-A transversions in Escherichia coli and C-to-G transversions in mammalian cells. These glycosylase base editors (GBEs) consist of a Cas9 nickase, a cytidine deaminase and a uracil-DNA glycosylase (Ung). Ung excises the U base created by the deaminase, forming an apurinic/apyrimidinic (AP) site that initiates the DNA repair process. In E. coli, we used activation-induced cytidine deaminase (AID) to construct AID-nCas9-Ung and found that it converts C to A with an average editing specificity of 93.8% ± 4.8% and editing efficiency of 87.2% ± 6.9%. For use in mammalian cells, we replaced AID with rat APOBEC1 (APOBEC-nCas9-Ung). We tested APOBEC-nCas9-Ung at 30 endogenous sites, and we observed C-to-G conversions with a high editing specificity at the sixth position of the protospacer between 29.7% and 92.2% and an editing efficiency between 5.3% and 53.0%. APOBEC-nCas9-Ung supplements the current adenine and cytidine BEs (ABE and CBE, respectively) and could be used to target G/C disease-causing mutations.

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Citations

Sep 5, 2020·Microbial Cell Factories·Zhenquan LiuDawei Zhang
Oct 21, 2020·Nature Reviews. Drug Discovery·Elizabeth M PortoGene W Yeo
Feb 2, 2021·Frontiers in Plant Science·Satya Swathi Nadakuduti, Felix Enciso-Rodríguez
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Feb 3, 2021·Plant Biotechnology Journal·Ayako Nishizawa-Yokoi, Seiichi Toki
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Methods Mentioned

BETA
deamination
PCR

Software Mentioned

PRIME
HiSeq Control Software ( HCS ) RTA
BWA
GENEWIZ
OFFinder
Cas
GenScript

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