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Nucleolus localization of SpyCas9 affects its stability and interferes with host protein translation in mammalian cells

文献类型：期刊论文

作者Tan, Renke4; Du, Wenhao3; Liu, Yiyang4; Cong, Xiaoji2; Bai, Meirong1; Jiang, Chenxiao4; Li, Zengxia4; Tan, Minjia2; Ma, Dengke K.1; Huang, Qiang3
刊名GENES & DISEASES
出版日期2022-05-01
卷号9期号:3页码:731-740
关键词CRISPR/Cas9 Gene editing Global transcription Nucleolus detention signal Translation
ISSN号2352-4820
DOI10.1016/j.gendis.2020.09.003
通讯作者Huang, Qiang(huangqiang@fudan.edu.cn) ; Jiang, Wei(jiangw@fudan.edu.cn) ; Dang, Yongjun(yongjundang@fudan.edu.cn)
英文摘要The CRISPR/Cas9 system, originally derived from the prokaryotic adaptive immune system, has been developed as efficient genome editing tools. It enables precise gene manipulation on chromosomal DNA through the specific binding of programmable sgRNA to target DNA, and the Cas9 protein, which has endonuclease activity, will cut a double strand break at specific locus. However, Cas9 is a foreign protein in mammalian cells, and the potential risks associated with its introduction into mammalian cells are not fully understood. In this study, we performed pull-down and mass spectrometry (MS) analysis of Streptococcus pyogenes Cas9 (SpyCas9) interacting proteins in HEK293T cells and showed that the majority of Cas9-associated proteins identified by MS were localized in the nucleolus. Interestingly, we further discovered that the Cas9 protein contains a sequence encoding a nucleolus detention signal (NoDS). Compared with wild-type (WT) Cas9, NoDS-mutated variants of Cas9 (mCas9) are less stable, although their gene editing activity is minimally affected. Overexpression of WT Cas9, but not mCas9, causes general effects on transcription and protein translation in the host cell. Overall, identification of NoDS in Cas9 will improve the understanding of Cas9's biological function in vivo, and the removal of NoDS in Cas9 may enhance its safety for future clinical use. Copyright (C) 2020, Chongqing Medical University. Production and hosting by Elsevier B.V.
WOS关键词CRISPR-CAS9 ; CRISPR/CAS9 ; NUCLEASES ; P53 ; SCREENS ; CAR
资助项目National Key Research and Development Program of China[2018YFC0310900] ; National Natural Science Foundation of China[31270830] ; National Natural Science Foundation of China[21572038] ; National Natural Science Foundation of China[21877016] ; National Natural Science Foundation of China[31671386] ; National Natural Science Foundation of China[81972621] ; Development Fund for Shanghai Talents ; State Key Laboratory of Bioorganic and Natural Products Chemistry, Fund of State Key Laboratory of Drug Research, Chinese Academy of Science[SIMM1601KF-08] ; Fudan University Graduate Student Research Grant
WOS研究方向Biochemistry & Molecular Biology ; Genetics & Heredity
语种英语
WOS记录号WOS:000793155300013
出版者ELSEVIER
源URL[http://119.78.100.183/handle/2S10ELR8/301050]  
专题新药研究国家重点实验室
通讯作者Huang, Qiang; Jiang, Wei; Dang, Yongjun
作者单位1.Univ Calif San Francisco, Cardiovasc Res Inst, San Francisco, CA 94158 USA
2.Chinese Acad Sci, Drug Discovery & Design Ctr, Shanghai Inst Mat Med, State Key Lab Drug Res, Shanghai 201203, Peoples R China
3.Fudan Univ, Sch Life Sci, State Key Lab Genet Engn, Shanghai 200043, Peoples R China
4.Fudan Univ, Shanghai Med Coll, Key Lab Metab & Mol Med, Minist Educ,Dept Biochem & Mol Biol,Sch Basic Med, Shanghai 200032, Peoples R China
推荐引用方式
GB/T 7714		 Tan, Renke,Du, Wenhao,Liu, Yiyang,et al. Nucleolus localization of SpyCas9 affects its stability and interferes with host protein translation in mammalian cells[J]. GENES & DISEASES,2022,9(3):731-740.
APA Tan, Renke.,Du, Wenhao.,Liu, Yiyang.,Cong, Xiaoji.,Bai, Meirong.,...&Dang, Yongjun.(2022).Nucleolus localization of SpyCas9 affects its stability and interferes with host protein translation in mammalian cells.GENES & DISEASES,9(3),731-740.
MLA Tan, Renke,et al."Nucleolus localization of SpyCas9 affects its stability and interferes with host protein translation in mammalian cells".GENES & DISEASES 9.3(2022):731-740.

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来源：上海药物研究所

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