Development of TALE‐adenine base editors in plants

Abstract

SummaryBase editors enable precise nucleotide changes at targeted genomic loci without requiring double‐stranded DNA breaks or repair templates. TALE‐adenine base editors (TALE‐ABEs) are genome editing tools, composed of a DNA‐binding domain from transcription activator‐like effectors (TALEs), an engineered adenosine deaminase (TadA8e), and a cytosine deaminase domain (DddA), that allow A•T‐to‐G•C editing in human mitochondrial DNA. However, the editing ability of TALE‐ABEs in plants apart from chloroplast DNA has not been described, so far, and the functional role how DddA enhances TadA8e is still unclear. We tested a series of TALE‐ABEs with different deaminase fusion architectures in Nicotiana benthamiana and rice. The results indicate that the double‐stranded DNA‐specific cytosine deaminase DddA can boost the activities of single‐stranded DNA‐specific deaminases (TadA8e or APOBEC3A) on double‐stranded DNA. We analysed A•T‐to‐G•C editing efficiencies in a β‐glucuronidase reporter system and showed precise adenine editing in genomic regions with high product purity in rice protoplasts. Furthermore, we have successfully regenerated rice plants with A•T‐to‐G•C mutations in the chloroplast genome using TALE‐ABE. Consequently, TALE‐adenine base editors provide alternatives for crop improvement and gene therapy by editing nuclear or organellar genomes.