不同物种MPG蛋白介导水稻A-to-K碱基编辑  

MPG proteins from different species mediate A-to-K base editing in rice

作　　者：吴雪梅 任斌 李欣格 旷永洁 张大伟[1] 周焕斌 WU Xue-Mei;REN Bin;LI Xin-Ge;KUANG Yong-Jie;ZHANG Da-Wei;ZHOU Huan-Bin(Key Laboratory of Bio-Resource and Eco-Environment of Ministry of Education,College of Life Science,Sichuan University,Chengdu 610065,China;State Key Laboratory for Biology of Plant Diseases and Insect Pests,Institute of Plant Protection,Chinese Academy of Agricultural Sciences,Beijing 100193,China;Scientific Observing and Experimental Station of Crop Pests in Guilin,Ministry of Agriculture and Rural Affairs,Guilin 541399,China;State Key Laboratory of North China Crop Improvement and Regulation,College of Plant Protection,Hebei Agricultural University,Baoding 071000,China)

机构地区：[1]四川大学生命科学学院生物资源与生态环境教育部重点实验室,成都610065 [2]中国农业科学院植物保护研究所植物病虫害综合治理全国重点实验室,北京100193 [3]农业农村部桂林作物有害生物科学观测实验站,桂林541399 [4]河北农业大学植物保护学院华北作物改良与调控国家重点实验室,保定071000

出　　处：《四川大学学报(自然科学版)》2025年第1期198-206,共9页Journal of Sichuan University(Natural Science Edition)

基　　金：科技创新2030-重大项目(2023ZD04074);国家重点研发计划(2023YFD1202905);植物病虫害综合治理全国重点实验室开放基金(SKLOF202402)。

摘　　要：为进一步优化植物腺嘌呤碱基颠换编辑工具,对不同来源的N-甲基嘌呤DNA糖基化酶(MPG,或称AAG)介导的植物腺嘌呤碱基颠换编辑工具pAKBE进行碱基编辑活性测试,获得具腺嘌呤碱基颠换编辑活性的MPG.通过农杆菌介导的水稻稳定遗传转化实验,首先探究鼠源mAAG的多种突变体在水稻中的A-to-K(K=G/T)编辑能力并逐步得到最佳变体mAAGv3-EF(A-to-T的编辑效率为9.37%),再将mAAG中的变异位点引入人源hMPG形成hMPGv3-EF,结果发现该突变体的编辑效率未有明显提升(A-to-T的编辑效率为10.00%).进一步还获得水稻内源OsMPG及其突变体OsMPG-N169S并证实它们具有A-to-K编辑能力.综合分析发现三种来源的N-甲基嘌呤DNA糖基化酶介导的植物腺嘌呤碱基颠换编辑工具均具有A-to-K编辑活性,这为未来植物腺嘌呤碱基颠换编辑的深入优化工作奠定一定基础.In order to further optimize plant A-to-K base editors(pAKBEs),which mediated by Nmethylpurine DNA glycosylase(MPG,also known as AAG)from different sources was tested.Initially,the A-to-G/T/C editing ability of various mutants of mAAG in rice were analyzed and the mAAGv3-EF mutant exhibited the highest A-to-T efficiency of 9.37%.Subsequently,the R165E and Y179F mutations from mAAG-EF were introduced into hMPGv3,resulting in the creation of hMPGv3-EF.However,the experimental results showed that hMPGv3-EF did not significantly improve pAKBE efficiency in rice,with the highest A-to-T editing efficiency reaching 10.00%.Finally,the A-to-T editing potential of rice endogenous OsMPG and its mutant,OsMPG-S,was demonstrated.Both OsMPG and OsMPG-S-mediated pAKBEs successfully generated A-to-T base editing.Further analysis revealed that all tested N-methylpurnine DNA glycosylases-mediated pAKBEs exhibited A-to-K editing efficiency.The discovery of distinct editing abilities in rBE118a~f may provide a solid foundation for future research.

关 键 词：N-甲基嘌呤DNA糖基化酶 植物腺嘌呤碱基颠换编辑器 基因组编辑 水稻 

分 类 号：Q78[生物学—分子生物学]