Discovery and Functional Characterization of Diverse Class 2 CRISPR-Cas Systems.
Discovery and Functional Characterization of Diverse Class 2 CRISPR-Cas Systems.
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DOI：
10.1016/j.molcel.2015.10.008
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发表时间：
2015-11-05
期刊：
影响因子：
16
通讯作者：
Koonin EV
中科院分区：
文献类型：
--
作者：
Shmakov S;Abudayyeh OO;Makarova KS;Wolf YI;Gootenberg JS;Semenova E;Minakhin L;Joung J;Konermann S;Severinov K;Zhang F;Koonin EV
微生物 CRISPR-Cas 系统分为 1 类（具有多亚基效应复合物）和 2 类（具有单蛋白效应子）。目前，已知的 2 类效应器只有 Cas9 和 Cpf1 两种。我们在此描述了三种不同的 2 类 CRISPR-Cas 系统。两个已识别系统 C2c1 和 C2c3 的效应器包含与 Cpf1 远亲相关的 RuvC 样核酸内切酶结构域。第三个系统 C2c2 包含一个具有两个预测 HEPN RNase 结构域的效应子。 C2c1 产生成熟的 CRISPR RNA (crRNA) 取决于 tracrRNA，而 C2c2 crRNA 的成熟则不依赖于 tracrRNA。我们发现 C2c1 系统可以以类似于 Cpf1 的 5'-PAM 依赖性方式介导 DNA 干扰。然而，与 Cpf1（一种单 RNA 引导的核酸酶）不同，C2c1 依赖 crRNA 和 tracrRNA 进行 DNA 切割。最后，比较分析表明，2 类 CRISPR-Cas 系统通过 1 类适应模块与从不同移动元件获得的效应蛋白的重组而多次进化。
Microbial CRISPR-Cas systems are divided into Class 1, with multisubunit effector complexes, and Class 2, with single protein effectors. Currently, only two Class 2 effectors, Cas9 and Cpf1, are known. We describe here three distinct Class 2 CRISPR-Cas systems. The effectors of two of the identified systems, C2c1 and C2c3, contain RuvC-like endonuclease domains distantly related to Cpf1. The third system, C2c2, contains an effector with two predicted HEPN RNase domains. Whereas production of mature CRISPR RNA (crRNA) by C2c1 depends on tracrRNA, C2c2 crRNA maturation is tracrRNA-independent. We found that C2c1 systems can mediate DNA interference in a 5’-PAM-dependent fashion analogous to Cpf1. However, unlike Cpf1, which is a single-RNA-guided nuclease, C2c1 depends on both crRNA and tracrRNA for DNA cleavage. Finally, comparative analysis indicates that Class 2 CRISPR-Cas systems evolved on multiple occasions through recombination of Class 1 adaptation modules with effector proteins acquired from distinct mobile elements.