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AcrIIC4通过防止R循环形成来抑制II-C类型Cas9

Wei Sun^1,2, Zhi Cheng^1,2,3, Jiuyu Wang^1,2

¹Key Laboratory of RNA Biology, Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China.

Proceedings of the National Academy of Sciences of the United States of America

|July 26, 2023

概括

反CRISPR蛋白AcrIIC4通过阻止DNA解和R循环形成来抑制CRISPR-Cas9. 这项结构研究揭示了AcrIIC4如何阻止Cas9核酶激活,为CRISPR-Cas免疫规避提供了洞察力.

科学领域:

分子生物学分子生物学
结构生物学结构生物学
微生物遗传学微生物遗传学

背景情况:

抗CRISPR (Acr) 蛋白质是菌体编码的CRISPR-Cas适应性免疫的抑制剂.
AcrIIC4是II-C型CRISPR-Cas9系统的宽频抑制剂,其机制未被描述.
了解Acr机制对于破译微生物防御系统和开发生物技术工具至关重要.

研究的目的:

为了阐明由AcrIIC4.4通过AcrIIC9 (HpaCas9) 抑制Hemophilus parainfluenzae Cas9 (HpaCas9) 的机制.
确定AcrIIC4与HpaCas9-sgRNA-DNA复合体相互作用的结构基础.
为了揭示AcrIIC4如何阻止Cas9核酶活性.

主要方法:

采用X射线晶体学,单独确定AcrIIC4的结构,并与HpaCas9-sgRNA和向DNA复合.
进行了生物化学测试,以评估AcrIIC4结合对Cas9活性的功能影响.

主要成果:

AcrIIC4 结合 HpaCas9.9 的 REC1 和 REC2 域之间的裂.
AcrIIC4结合限制了REC2域的移动性,防止了目标双链DNA解.
形成全长导向RNA:目标DNA异重复和随后的R循环被阻止,抑制了Cas9核酶的激活.

结论:

AcrIIC4采用一种独特的机制,通过防止R循环形成来抑制Cas9,这是DNA裂变的关键步骤.
这种抑制发生,尽管允许初始DNA与Cas9效应体复合体结合.
这些发现为一种新的抗CRISPR策略及其对CRISPR-Cas免疫力的影响提供了结构性的见解.

关键词:

这就是CRISPR-Cas系统.在DNA解过程中,这是一个R环.在 REC 叶片中,REC 叶片这是一种抗CRISPR蛋白质.

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