文献类型： 外文期刊

作者： He, Yao ¹ ; Liu, Shishi ² ; Chen, Long ⁴ ; Pu, Dongkai ² ; Zhong, Zhaohui ² ; Xu, Tang ¹ ; Ren, Qiurong ² ; Dong, Chuan ² ; Wang, Yawei ² ; Wang, Danning ² ; Zheng, Xuelian ¹ ; Guo, Fengbiao ⁶ ; Zhang, Tao ⁴ ; Qi, Yiping ⁹ ; Zhang, Yong ¹ ;

作者机构： 1.Southwest Univ, Integrat Sci Ctr Germplasm Creat Western China Cho, Sch Life Sci, Chongqing Key Lab Tree Germplasm Innovat & Utiliza, Chongqing 400715, Peoples R China

2.Univ Elect Sci & Technol China, Ctr Informat Biol, Sch Life Sci & Technol, Dept Biotechnol, Chengdu 610054, Peoples R China

3.Sichuan Acad Agr Sci, Sichuan Inst Edible Fungi, Chengdu 610066, Peoples R China

4.Yangzhou Univ, Coll Agr, Jiangsu Key Lab Crop Genom & Mol Breeding, Zhongshan Biol Breeding Lab,Key Lab Plant Funct Ge, Yangzhou 225009, Peoples R China

5.Yangzhou Univ, Jiangsu Coinnovat Ctr Modern Prod Technol Grain Cr, Jiangsu Key Lab Crop Genet & Physiol, Yangzhou 225009, Peoples R China

6.Wuhan Univ, Dept Resp & Crit Care Med, Zhongnan Hosp, Wuhan 430071, Peoples R China

7.Wuhan Univ, Key Lab Combinatorial Biosynth & Drug Discovery, Minist Educ, Wuhan 430072, Peoples R China

8.Wuhan Univ, Sch Pharmaceut Sci, Wuhan 430072, Peoples R China

9.Univ Maryland, Dept Plant Sci & Landscape Architecture, College Pk, MD 20742 USA

10.Univ Maryland, Inst Biosci & Biotechnol Res, Rockville, MD 20850 USA

关键词： anti-CRISPR; Cas12a; AcrVA1; off-target effects; fine-tuning genome editing; inducible and tissue specific genome editing; CRISPRa; synthetic logic circuit

期刊名称：SCIENCE CHINA-LIFE SCIENCES （ 影响因子：8.0； 五年影响因子：7.3 ）

ISSN： 1674-7305

年卷期： 2024 年

页码：

收录情况： SCI

摘要： CRISPR-Cas12a genome engineering systems have been widely used in plant research and crop breeding. To date, the performance and use of anti-CRISPR-Cas12a systems have not been fully established in plants. Here, we conduct in silico analysis to identify putative anti-CRISPR systems for Cas12a. These putative anti-CRISPR proteins, along with known anti-CRISPR proteins, are assessed for their ability to inhibit Cas12a cleavage activity in vivo and in planta. Among all anti-CRISPR proteins tested, AcrVA1 shows robust inhibition of Mb2Cas12a and LbCas12a in E. coli. Further tests show that AcrVA1 inhibits LbCas12a mediated genome editing in rice protoplasts and stable transgenic lines. Impressively, co-expression of AcrVA1 mitigates off-target effects by CRISPR-LbCas12a, as revealed by whole genome sequencing. In addition, transgenic plants expressing AcrVA1 exhibit different levels of inhibition to LbCas12a mediated genome editing, representing a novel way of fine-tuning genome editing efficiency. By controlling temporal and spatial expression of AcrVA1, we show that inducible and tissue specific genome editing can be achieved in plants. Furthermore, we demonstrate that AcrVA1 also inhibits LbCas12a-based CRISPR activation (CRISPRa) and based on this principle we build logic gates to turn on and off target genes in plant cells. Together, we have established an efficient anti-CRISPR-Cas12a system in plants and demonstrate its versatile applications in mitigating off-target effects, fine-tuning genome editing efficiency, achieving spatial-temporal control of genome editing, and generating synthetic logic gates for controlling target gene expression in plant cells.