TadA orthologs enable both cytosine and adenine editing of base editors

Shuqian Zhang; Bo Yuan; Jixin Cao; Liting Song; Jinlong Chen; Jiayi Qiu; Zilong Qiu; Xing-Ming Zhao; Jingqi Chen; Tian-Lin Cheng

doi:10.1038/s41467-023-36003-3

TadA orthologs enable both cytosine and adenine editing of base editors

Nat Commun. 2023 Jan 26;14(1):414. doi: 10.1038/s41467-023-36003-3.

Authors

Shuqian Zhang^#^{1

2}, Bo Yuan^#³, Jixin Cao⁴, Liting Song⁴, Jinlong Chen¹, Jiayi Qiu¹, Zilong Qiu^{3

5

6}, Xing-Ming Zhao^{4

7

8}, Jingqi Chen^{4

7

8}, Tian-Lin Cheng⁹

Affiliations

¹ Institute for Translational Brain Research, State Key Laboratory of Medical Neurobiology, MOE Frontiers Center for Brain Science, Institute of Pediatrics, National Children's Medical Center, Children's Hospital, Fudan University, Shanghai, China.
² Department of Pediatrics, Qilu Hospital of Shandong University, Ji'nan, 250012, China.
³ Institute of Neuroscience, State Key Laboratory of Neuroscience, CAS Center for Excellence in Brain Science and Intelligence Technology, Chinese Academy of Sciences, Shanghai, 200031, China.
⁴ Institute of Science and Technology for Brain-Inspired Intelligence, Fudan University, Shanghai, China.
⁵ National Clinical Research Center for Aging and Medicine, Huashan Hopsital, Fudan University, Shanghai, 200032, China.
⁶ Songjiang Hospital, Songjiang Institute, Shanghai Jiao Tong University School of Medicine, Shanghai, China.
⁷ State Key Laboratory of Medical Neurobiology, Institutes of Brain Science, Fudan University, Shanghai, China.
⁸ MOE Key Laboratory of Computational Neuroscience and Brain-Inspired Intelligence, and MOE Frontiers Center for Brain Science, Fudan University, Shanghai, China.
⁹ Institute for Translational Brain Research, State Key Laboratory of Medical Neurobiology, MOE Frontiers Center for Brain Science, Institute of Pediatrics, National Children's Medical Center, Children's Hospital, Fudan University, Shanghai, China. chengtianlin@fudan.edu.cn.

^# Contributed equally.

Abstract

Cytidine and adenosine deaminases are required for cytosine and adenine editing of base editors respectively, and no single deaminase could enable concurrent and comparable cytosine and adenine editing. Additionally, distinct properties of cytidine and adenosine deaminases lead to various types of off-target effects, including Cas9-indendepent DNA off-target effects for cytosine base editors (CBEs) and RNA off-target effects particularly severe for adenine base editors (ABEs). Here we demonstrate that 25 TadA orthologs could be engineered to generate functional ABEs, CBEs or ACBEs via single or double mutations, which display minimized Cas9-independent DNA off-target effects and genotoxicity, with orthologs B5ZCW4, Q57LE3, E8WVH3, Q13XZ4 and B3PCY2 as promising candidates for further engineering. Furthermore, RNA off-target effects of TadA ortholog-derived base editors could be further reduced or even eliminated by additional single mutation. Taken together, our work expands the base editing toolkits, and also provides important clues for the potential evolutionary process of deaminases.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Adenine
Adenosine / genetics
CRISPR-Cas Systems / genetics
Cytosine*
DNA
Gene Editing*
RNA

Substances

Cytosine
Adenine
DNA
RNA
Adenosine