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Repurposing type I–F CRISPR–Cas system as a transcriptional activation tool in human cells

来源:Nature

Abstract

Class 2 CRISPR–Cas proteins have been widely developed as genome editing and transcriptional regulating tools. Class 1 type I CRISPR–Cas constitutes ~60% of all the CRISPR–Cas systems. However, only type I–B and I–E systems have been used to control mammalian gene expression and for genome editing. Here we demonstrate the feasibility of using type I–F system to regulate human gene expression. By fusing transcription activation domain to Pseudomonas aeruginosa type I–F Cas proteins, we activate gene transcription in human cells. In most cases, type I–F system is more efficient than other CRISPR-based systems. Transcription activation is enhanced by elongating the crRNA. In addition, we achieve multiplexed gene activation with a crRNA array. Furthermore, type I–F system activates target genes specifically without off-target transcription activation. These data demonstrate the robustness and programmability of type I–F CRISPR–Cas in human cells.


Introduction


Clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated (cas) genes-based defence systems protect bacteria and archaea against phage and other foreign genetic elements1,2,3. Since the identification of increasing number of cas genes, the CRISPR–Cas systems have been classified into two Classes (Class 1 and Class 2) and six types (Type I–VI)4 based on the different arrangements of cas genes and the subunits of effector complexes5,6,7. Class 2 CRISPR–Cas systems, the best-studied system with single effector protein (e.g., Cas9, Cas12, or Cas13) for foreign DNA or RNA interference, are subdivided into Type II (Cas9), Type V (Cas12), and Type VI (Cas13). In the past few years, Class 2 CRISPR–Cas systems have revolutionized both basic and clinical researches, enabling more rapid, precise, and robust genome editing and modifications in cultured cells and animals8,9,10,11,12,13,14,15,16,17. However, there were only a few applications of Class 1 CRISPR–Cas (Type I, Type III and Type IV) system.

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Fig. 1: Targeted transcription activation by type I–F PaeCascade.

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参考文献

Repurposing type I–F CRISPR–Cas system as a transcriptional activation tool in human cells

Yuxi Chen, Jiaqi Liu, Shengyao Zhi, Qi Zheng, Wenbin Ma, Junjiu Huang, Yizhi Liu, Dan Liu, Puping Liang & Zhou Songyang


文章分类: 科技论文CAS蛋白
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