Additional, diverse CRISPR systems CRISPR systems have been revolutionizing molecular biology. Mining the metagenomic database, Yan et al. systematically discovered additional subtypes of type V CRISPR-Cas systems. The additional Cas12 effectors displayed a range of activities, including target and collateral cleavage of single-stranded RNA and DNA, as well as double-stranded DNA nicking and cleavage. These diverse nuclease activities suggest how an ancient transposase may have evolved into various type V effectors and expand the nucleic acid detection and genome-editing toolbox. Science, this issue p. 88 Functionalities of newly identified type V CRISPR-Cas12 effectors include RNA cleavage and double-stranded DNA nicking. Type V CRISPR-Cas systems are distinguished by a single RNA-guided RuvC domain-containing effector, Cas12. Although effectors of subtypes V-A (Cas12a) and V-B (Cas12b) have been studied in detail, the distinct domain architectures and diverged RuvC sequences of uncharacterized Cas12 proteins suggest unexplored functional diversity. Here, we identify and characterize Cas12c, -g, -h, and -i. Cas12c, -h, and -i demonstrate RNA-guided double-stranded DNA (dsDNA) interference activity. Cas12i exhibits markedly different efficiencies of CRISPR RNA spacer complementary and noncomplementary strand cleavage resulting in predominant dsDNA nicking. Cas12g is an RNA-guided ribonuclease (RNase) with collateral RNase and single-strand DNase activities. Our study reveals the functional diversity emerging along different routes of type V CRISPR-Cas evolution and expands the CRISPR toolbox.

中文翻译：

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功能多样的 V 型 CRISPR-Cas 系统


另外，多样化的 CRISPR 系统 CRISPR 系统已经彻底改变了分子生物学。挖掘宏基因组数据库，Yan 等人。系统地发现了 V 型 CRISPR-Cas 系统的其他亚型。额外的 Cas12 效应子表现出一系列活性，包括单链 RNA 和 DNA 的靶标和旁路切割，以及双链 DNA 切口和切割。这些不同的核酸酶活性表明古老的转座酶可能如何进化成各种V型效应子并扩展核酸检测和基因组编辑工具箱。科学，本期第 14 页。 88 新鉴定的 V 型 CRISPR-Cas12 效应子的功能包括 RNA 切割和双链 DNA 切口。 V 型 CRISPR-Cas 系统的特点是包含单个 RNA 引导的 RuvC 结构域效应子 Cas12。尽管对 VA (Cas12a) 和 VB (Cas12b) 亚型的效应子进行了详细研究，但未表征的 Cas12 蛋白的不同结构域结构和不同的 RuvC 序列表明尚未探索的功能多样性。在这里，我们识别并表征了 Cas12c、-g、-h 和 -i。 Cas12c、-h 和 -i 表现出 RNA 引导的双链 DNA (dsDNA) 干扰活性。 Cas12i 表现出明显不同的 CRISPR RNA 间隔互补和非互补链切割效率，导致主要的 dsDNA 切口。 Cas12g 是一种 RNA 引导的核糖核酸酶 (RNase)，具有附带 RNase 和单链 DNase 活性。我们的研究揭示了 V 型 CRISPR-Cas 进化不同途径中出现的功能多样性，并扩展了 CRISPR 工具箱。