Systematic Investigation of the Effects of Multiple SV40 Nuclear Localization Signal Fusion on the Genome Editing Activity of Purified SpCas9

doi:10.3390/bioengineering9020083

	Systematic Investigation of the Effects of Multiple SV40 Nuclear Localization Signal Fusion on the Genome Editing Activity of Purified SpCas9
	Shui, Sailan1 ; Wang, Shaojie1,2,3 ; Liu, Jia1
	2022-02
发表期刊	BIOENGINEERING-BASEL
EISSN	2306-5354
卷号	9 期号:2
发表状态	已发表
DOI	10.3390/bioengineering9020083
摘要	The emergence of CRISPR-Cas9 technology has revolutionized both basic and translational biomedical research. For Cas9 nuclease to exert genome editing activity, nuclear localization signal (NLS) derived from simian virus 40 (SV40) T antigen is commonly installed as genetic fusion to direct the intracellular Cas9 proteins to the nucleus of cells. Notably, previous studies have shown that multiple SV40 NLS fusion can improve the targeting activity of Cas9-derived genome-editing and base-editing tools. In addition, the multi-NLS fusion can increase the intracellular activity of Cas9 in the forms of both constitutive expression and directly delivered Cas9-guide RNA ribonucleoprotein (RNP) complex. However, the relationship between NLS fusion and intracellular Cas9 activity has not been fully understood, including the dependency of activity on the number or organization of NLS fusion. In the present study, we constructed and purified a set of Streptococcus pyogenes Cas9 (SpCas9) variants containing one to four NLS repeats at the N- or C-terminus of the proteins and systematically analyzed the effects of multi-NLS fusion on the activity of SpCas9 RNPs. It was found that multi-NLS fusion could improve the intracellular activity as lipofected or nucleofected Cas9 RNPs. Importantly, multi-NLS fusion could enhance the genome-editing activity of SpCas9 RNPs in primary and stem/progenitor cells and mouse embryos.
关键词	CRISPR-Cas9 Cas9 ribonucleoproteins (RNPs) nuclear localization signal (NLS) genome editing
URL	查看原文
收录类别	SCI ; SCIE
语种	英语
WOS研究方向	Biotechnology & Applied Microbiology ; Engineering
WOS类目	Biotechnology & Applied Microbiology ; Engineering, Biomedical
WOS记录号	WOS:000771659000001
出版者	MDPI
引用统计
文献类型	期刊论文
条目标识符	https://kms.shanghaitech.edu.cn/handle/2MSLDSTB/165001
专题	生命科学与技术学院_硕士生免疫化学研究所_PI研究组_刘佳组
通讯作者	Liu, Jia
作者单位	1.Shanghai Tech Univ, Shanghai Inst Adv Immunochem Studies, Sch Life Sci & Technol, Shanghai 201210, Peoples R China 2.Lab Prot Design & Immunoengn LPDI STI EPFL, CH-1015 Lausanne, Switzerland 3.Swiss Inst Bioinformat SIB, CH-1015 Lausanne, Switzerland
第一作者单位	免疫化学研究所
通讯作者单位	免疫化学研究所
第一作者的第一单位	免疫化学研究所
推荐引用方式 GB/T 7714	Shui, Sailan,Wang, Shaojie,Liu, Jia. Systematic Investigation of the Effects of Multiple SV40 Nuclear Localization Signal Fusion on the Genome Editing Activity of Purified SpCas9[J]. BIOENGINEERING-BASEL,2022,9(2).
APA	Shui, Sailan,Wang, Shaojie,&Liu, Jia.(2022).Systematic Investigation of the Effects of Multiple SV40 Nuclear Localization Signal Fusion on the Genome Editing Activity of Purified SpCas9.BIOENGINEERING-BASEL,9(2).
MLA	Shui, Sailan,et al."Systematic Investigation of the Effects of Multiple SV40 Nuclear Localization Signal Fusion on the Genome Editing Activity of Purified SpCas9".BIOENGINEERING-BASEL 9.2(2022).