Fusion of a rice endogenous N-methylpurine DNA glycosylase to a plant adenine base transition editor ABE8e enables A-toK base editing in rice plants  

作　　者：Yucai Li Shaoya Li Chenfei Li Chen Zhang Lei Yan Jingying Li Yubing He Yan Guo Lanqin Xia 

机构地区：[1]Institute of Crop Sciences(ICS),Chinese Academy of Agricultural Sciences(CAAS),Beijing 100081,China [2]State Key Laboratory of Plant Physiology and Biochemistry,College of Biological Sciences,China Agricultural University,Beijing 100193,China [3]Hainan Yazhou Bay Seed Laboratory/National Nanfan Research Institute(Sanya),CAAS,Sanya 572024,China

出　　处：《aBIOTECH》2024年第2期127-139,共13页生物技术通报（英文版）

基　　金：funded by the National Natural Science Foundation of China(Grant No.32188102 to L.X),Hainan Yazhou Bay Seed Lab(Grant No.B23CJ0208 to L.X);the Central Public-interest Scientific Institution Basal Research Fund(Grant No.ZDXM2308 to L.X);National Engineering Research Centre of Crop Molecular Breeding.

摘　　要：Engineering of a new type of plant base editor for simultaneous adenine transition and transversion within the editing window will greatly expand the scope and potential of base editing in directed evolution and crop improvement.Here,we isolated a rice endogenous hypoxanthine excision protein,N-methylpurine DNA glycosylase(OsMPG),and engineered two plant A-to-K(K=G or T)base editors,rAKBE01 and rAKBE02,for simultaneous adenine transition and transversion base editing in rice by fusing OsMPG or its mutant mOsMPG to a plant adenine transition base editor,ABE8e.We further coupled either OsMPG or mOsMPG with a transactivation factor VP64 to generate rAKBE03 and rAKBE04,respectively.Testing these four rAKBEs,at five endogenous loci in rice protoplasts,indicated that rAKBE03 and rAKBE04 enabled higher levels of A-to-G base transitions when compared to ABE8e and ABE8e-VP64.Furthermore,whereas rAKBE01 only enabled A-to-C/T editing at one endogenous locus,in comparison with rAKBE02 and rAKBE03,rAKBE04 could significantly improve the A-to-C/T base transversion efficiencies by up to 6.57-and 1.75-fold in the rice protoplasts,respectively.Moreover,although no stable lines with A-to-C transversion were induced by rAKBE01 and rAKBE04,rAKBE04 could enable simultaneous A-to-G and A-to-T transition and transversion base editing,at all the five target loci,with the efficiencies of A-to-G transition and A-to-T transversion editing ranging from 70.97 to 92.31%and 1.67 to 4.84%in rice stable lines,respectively.Together,these rAKBEs enable different portfolios of editing products and,thus,now expands the potential of base editing in diverse application scenario for crop improvement.

关 键 词：Rice(Oryza sativa L) Rice N-methylpurine DNA glycosylase(OsMPG) A-to-K base editor(AKBE) Transactivation module VP64 

分 类 号：S511[农业科学—作物学]