{"title":"I 型 CRISPR-Cas 系统中 HNH 介导的目标 DNA 切割机制","authors":"","doi":"10.1016/j.molcel.2024.06.033","DOIUrl":null,"url":null,"abstract":"<p>The metagenome-derived type I-E and type I-F variant CRISPR-associated complex for antiviral defense (Cascade) complexes, fused with HNH domains, precisely cleave target DNA, representing recently identified genome editing tools. However, the underlying working mechanisms remain unknown. Here, structures of type I-F<sup>HNH</sup> and I-E<sup>HNH</sup> Cascade complexes at different states are reported. In type I-F<sup>HNH</sup> Cascade, Cas8f<sup>HNH</sup> loosely attaches to Cascade head and is adjacent to the 5′ end of the target single-stranded DNA (ssDNA). Formation of the full R-loop drives the Cascade head to move outward, allowing Cas8f<sup>HNH</sup> to detach and rotate ∼150° to accommodate target ssDNA for cleavage. In type I-E<sup>HNH</sup> Cascade, Cas5e<sup>HNH</sup> domain is adjacent to the 5′ end of the target ssDNA. Full crRNA-target pairing drives the lift of the Cascade head, widening the substrate channel for target ssDNA entrance. Altogether, these analyses into both complexes revealed that crRNA-guided positioning of target DNA and target DNA-induced HNH unlocking are two key factors for their site-specific cleavage of target DNA.</p>","PeriodicalId":18950,"journal":{"name":"Molecular Cell","volume":null,"pages":null},"PeriodicalIF":14.5000,"publicationDate":"2024-07-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Mechanisms for HNH-mediated target DNA cleavage in type I CRISPR-Cas systems\",\"authors\":\"\",\"doi\":\"10.1016/j.molcel.2024.06.033\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p>The metagenome-derived type I-E and type I-F variant CRISPR-associated complex for antiviral defense (Cascade) complexes, fused with HNH domains, precisely cleave target DNA, representing recently identified genome editing tools. However, the underlying working mechanisms remain unknown. Here, structures of type I-F<sup>HNH</sup> and I-E<sup>HNH</sup> Cascade complexes at different states are reported. In type I-F<sup>HNH</sup> Cascade, Cas8f<sup>HNH</sup> loosely attaches to Cascade head and is adjacent to the 5′ end of the target single-stranded DNA (ssDNA). Formation of the full R-loop drives the Cascade head to move outward, allowing Cas8f<sup>HNH</sup> to detach and rotate ∼150° to accommodate target ssDNA for cleavage. In type I-E<sup>HNH</sup> Cascade, Cas5e<sup>HNH</sup> domain is adjacent to the 5′ end of the target ssDNA. Full crRNA-target pairing drives the lift of the Cascade head, widening the substrate channel for target ssDNA entrance. Altogether, these analyses into both complexes revealed that crRNA-guided positioning of target DNA and target DNA-induced HNH unlocking are two key factors for their site-specific cleavage of target DNA.</p>\",\"PeriodicalId\":18950,\"journal\":{\"name\":\"Molecular Cell\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":14.5000,\"publicationDate\":\"2024-07-23\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Molecular Cell\",\"FirstCategoryId\":\"99\",\"ListUrlMain\":\"https://doi.org/10.1016/j.molcel.2024.06.033\",\"RegionNum\":1,\"RegionCategory\":\"生物学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q1\",\"JCRName\":\"BIOCHEMISTRY & MOLECULAR BIOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Molecular Cell","FirstCategoryId":"99","ListUrlMain":"https://doi.org/10.1016/j.molcel.2024.06.033","RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"BIOCHEMISTRY & MOLECULAR BIOLOGY","Score":null,"Total":0}
引用次数: 0
摘要
元基因组衍生的 I-E 型和 I-F 型变体 CRISPR 相关抗病毒复合物(Cascade)复合物融合了 HNH 结构域,可精确切割目标 DNA,是最近发现的基因组编辑工具。然而,其潜在的工作机制仍然未知。本文报告了 I-FHNH 型和 I-EHNH 型 Cascade 复合物在不同状态下的结构。在I型-FHNH Cascade中,Cas8fHNH松散地附着在Cascade头部,与目标单链DNA(ssDNA)的5′端相邻。全 R 环的形成会驱动 Cascade 头向外移动,使 Cas8fHNH 脱离并旋转 150°,以适应目标 ssDNA 的切割。在 I 型-EHNH 级联过程中,Cas5eHNH 结构域与靶 ssDNA 的 5′端相邻。crRNA 与目标的完全配对驱动 Cascade 头抬起,拓宽了目标 ssDNA 进入的底物通道。对这两种复合物的分析表明,crRNA 引导的靶 DNA 定位和靶 DNA 诱导的 HNH 解锁是它们定点特异性裂解靶 DNA 的两个关键因素。
Mechanisms for HNH-mediated target DNA cleavage in type I CRISPR-Cas systems
The metagenome-derived type I-E and type I-F variant CRISPR-associated complex for antiviral defense (Cascade) complexes, fused with HNH domains, precisely cleave target DNA, representing recently identified genome editing tools. However, the underlying working mechanisms remain unknown. Here, structures of type I-FHNH and I-EHNH Cascade complexes at different states are reported. In type I-FHNH Cascade, Cas8fHNH loosely attaches to Cascade head and is adjacent to the 5′ end of the target single-stranded DNA (ssDNA). Formation of the full R-loop drives the Cascade head to move outward, allowing Cas8fHNH to detach and rotate ∼150° to accommodate target ssDNA for cleavage. In type I-EHNH Cascade, Cas5eHNH domain is adjacent to the 5′ end of the target ssDNA. Full crRNA-target pairing drives the lift of the Cascade head, widening the substrate channel for target ssDNA entrance. Altogether, these analyses into both complexes revealed that crRNA-guided positioning of target DNA and target DNA-induced HNH unlocking are two key factors for their site-specific cleavage of target DNA.
期刊介绍:
Molecular Cell is a companion to Cell, the leading journal of biology and the highest-impact journal in the world. Launched in December 1997 and published monthly. Molecular Cell is dedicated to publishing cutting-edge research in molecular biology, focusing on fundamental cellular processes. The journal encompasses a wide range of topics, including DNA replication, recombination, and repair; Chromatin biology and genome organization; Transcription; RNA processing and decay; Non-coding RNA function; Translation; Protein folding, modification, and quality control; Signal transduction pathways; Cell cycle and checkpoints; Cell death; Autophagy; Metabolism.