Engineering CjCas9 for Efficient Base Editing and Prime Editing

doi:10.1089/crispr.2024.0018

	Engineering CjCas9 for Efficient Base Editing and Prime Editing
	Liu SY(刘思远)1 ; Zhao YD(赵英迪)1 ; Mo QQ(莫淇钦)1 ; Sun YD(孙亚东)2 ; Ma HH(马涵慧)1
	2024-11-01
发表期刊	CRISPR JOURNAL (IF:3.7[JCR-2023],4.2[5-Year])
ISSN	2573-1599
EISSN	2573-1602
卷号	7 期号:6
发表状态	已发表
DOI	10.1089/crispr.2024.0018
摘要	The CRISPR-Cas9 system has been applied for clinical applications of gene therapy. Most CRISPR-based gene therapies are derived from Streptococcus pyogenes Cas9, which is challenging to package into a single adeno-associated virus vector and limits its clinical applications. Campylobacter jejuni Cas9 (CjCas9) is one of the smallest Cas9 proteins. CjCas9-mediated base editing (CjBE) efficiency varies across genomic sites, while CjCas9-mediated prime editing (CjPE) efficiency is less than 5% on average. Here we developed enhanced cytosine base editors (enCjCBEs) and adenine base editors (enCjABEs) by engineered CjCas9P47K. We demonstrated the robust C-to-T conversion (70% on average) by enCjCBE or A-to-G conversion (76% on average) by enCjABE. Meanwhile, we applied the CjCas9P47K variant to generate enhanced CjPE (enCjPE), which increases the editing efficiency 17-fold at the PRNP site over wild-type CjPE. Fusing nonspecific DNA binding protein Sso7d to enCjCas9 and MS2 stem-loop RNA aptamer to the 3-terminal of cognate pegRNA resulted in 12% editing efficiency on average with a 24-fold increase over wild-type CjPE, and we termed it SsenCjPE. The SsenCjPE can also be combined with hMLH1dn to further increase the editing efficiency and MMLV RTase Delta RnH to reduce size. Finally, we introduced an additional mutation D829R into SsenCjPE and generated SsenCjPE-M2 with a 61-fold increase of PE efficiency over wild-type at the PRNP site. In summary, enCjBEs, SsenCjPEs, or SsenCjPE-M2 are compact Cas9-derived BE or prime editors in biological research or biomedical applications.
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收录类别	SCI
语种	英语
WOS研究方向	Genetics & Heredity
WOS类目	Genetics & Heredity
WOS记录号	WOS:001356632400001
出版者	MARY ANN LIEBERT, INC
文献类型	期刊论文
条目标识符	https://kms.shanghaitech.edu.cn/handle/2MSLDSTB/449039
专题	生命科学与技术学院_博士生生命科学与技术学院_PI研究组_马涵慧组生命科学与技术学院_PI研究组_孙亚东组
共同第一作者	Zhao YD(赵英迪)
通讯作者	Ma HH(马涵慧)
作者单位	1.上海科技大学生命科学与技术学院基因编辑中心 2.上海科技大学生命科学与技术学院
第一作者单位	生命科学与技术学院
通讯作者单位	生命科学与技术学院
第一作者的第一单位	生命科学与技术学院
推荐引用方式 GB/T 7714	Liu SY,Zhao YD,Mo QQ,et al. Engineering CjCas9 for Efficient Base Editing and Prime Editing[J]. CRISPR JOURNAL,2024,7(6).
APA	Liu SY,Zhao YD,Mo QQ,Sun YD,&Ma HH.(2024).Engineering CjCas9 for Efficient Base Editing and Prime Editing.CRISPR JOURNAL,7(6).
MLA	Liu SY,et al."Engineering CjCas9 for Efficient Base Editing and Prime Editing".CRISPR JOURNAL 7.6(2024).