Efficient and multiplex gene upregulation in plants through CRISPR-Cas-mediated knockin of enhancers  

作　　者：Qi Yao Rundong Shen Yang Shao Yifu Tian Peijin Han Xuening Zhang Jian-Kang Zhu Yuming Lu 

机构地区：[1]Shanghai Collaborative Innovation Center of Agri-Seeds,Joint Center for Single-Cell Biology,School of Agriculture and Biology,Shanghai Jiao Tong University,Shanghai 200240,China [2]Shanghai Center for Plant Stress Biology,Center for Excellence in Molecular Plant Sciences,Chinese Academy of Sciences,Shanghai 201602,China [3]University of the Chinese Academy of Sciences,Beijing,China [4]College of Plant Science and Technology,Huazhong Agricultural University,Wuhan 430070,China [5]institute of Advanced Biotechnology and School of Medicine,Southern University of Science and Technology,Shenzhen 518055,China

出　　处：《Molecular Plant》2024年第9期1472-1483,共12页分子植物（英文版）

基　　金：supported by the National Key R&D Program of China(2021YFD1201300 to Y.L.and 2021YFA1300404 to J.-K.Z.);the Shanghai Agricultural Science and Technology Innovation Program of China(K2023001 to Y.L.);the National Natural Science Foundation of China(32070396 to Y.L.and 32188102 to J.-K.Z.).

摘　　要：Geneupregulation through genome editing is important for plant research and breeding.Targeted insertion of short transcriptional enhancers(STEs)into gene promoters may offer a universal solution akin to transgene-mediated overexpression while avoiding the drawbacks associated with transgenesis.Here,we introduce an“in locus activation”technique in rice that leverages well-characterized STEs for refined,heritable,and multiplexed gene upregulation.To address the scarcity of potent enhancers,we developed a large-scale mining approach and discovered a suite of STEs that are capable of enhancing gene expression in rice protoplasts.The in locus integration of these STEs into eight rice genes resulted in substantial tran-scriptional upregulation in the edited plants,with up to 869.1-fold increases in their transcript levels.Em-ploying a variety of STEs,we achieved delicate control of gene expression,enabling the fine-tuning of key phenotypic traits such as plant height.Our approach also enabled efficient multiplexed gene upregu-lation,with up to four genes activated simultaneously,significantly enhancing the nicotinamide mononucleotide metabolic pathway.Importantly,heritability studies from the To to T3 generations confirmed the stable and heritable nature of STE-driven gene activation.Collectively,our work demon-strates that coupled with STE mining,leveraging genome editing for in locus activation and gene upregu-lation holds great promise to be widely adopted in fundamental plant research and crop breeding.

关 键 词：genome editing short transcriptional enhancers gene upregulation crop breeding 

分 类 号：Q94[生物学—植物学]