密歇根大学张燕小组宣布他们研究出Cas9感知CRISPR RNA丰度以调控CRISPR间隔片段获取。2025年9月3日出版的《自然》发表了这项成果。

该课题组发现了Cas9的第一个独立于其双RNA伴侣的生物学作用。在crRNA和/或tracrRNA耗尽后，apoCas9奈瑟菌刺激间隔物获取效率。在生理上，Cas9在具有短CRISPR阵列的细胞中感知低水平的crRNA——例如那些正在进行阵列新生或自然阵列收缩的细胞——并动态上调获取以迅速扩大小的免疫记忆库。随着CRISPR阵列的扩展，crRNA丰度的增加反过来降低了apoCas9的可用性，从而抑制了获取，以减轻与获取升高相关的自身免疫风险。

虽然apoCas9的核酸酶叶本身就足以刺激获取，但在低免疫深度的细胞中，只有全长Cas9响应crRNA水平才能促进获取。最后，课题组研究人员发现这种活性在多个II-C型Cas9同源基因中是进化保守的。总之，该研究团队建立了一个自动补充反馈机制，其中apoCas9通过作为crRNA传感器和间隔获取调节器来保护CRISPR免疫深度。

据悉，原核生物通过获取外源DNA片段（称为间隔片段）进入CRISPR阵列来创造适应性免疫记忆。在II型CRISPR-Cas系统中，RNA引导的效应体Cas9也通过从原间隔器相邻基序（PAM）侧翼的DNA中选择间隔器来辅助获取机制。

附：英文原文

Title: Cas9 senses CRISPR RNA abundance to regulate CRISPR spacer acquisition

Author: Zhou, Xufei, Diao, Rucheng, Li, Xin, Ziegler, Christine A., Gramelspacher, Max J., Freddolino, Lydia, Hou, Zhonggang, Zhang, Yan

Issue&Volume: 2025-09-03

Abstract: Prokaryotes create adaptive immune memories by acquiring foreign DNA snippets, known as spacers, into the CRISPR array1. In type II CRISPR-Cas systems, the RNA-guided effector Cas9 also assists the acquisition machinery by selecting spacers from protospacer adjacent motif (PAM)-flanked DNA2,3. Here, we uncover the first biological role for Cas9 that is independent of its dual RNA partners. Following depletion of crRNA and/or tracrRNA, Neisseria apoCas9 stimulates spacer acquisition efficiency. Physiologically, Cas9 senses low levels of crRNA in cells with short CRISPR arrays – such as those undergoing array neogenesis or natural array contractions – and dynamically upregulates acquisition to quickly expand the small immune memory banks. As the CRISPR array expands, rising crRNA abundance in turn reduces apoCas9 availability, thereby dampening acquisition to mitigate autoimmunity risks associate with elevated acquisition. While apoCas9’s nuclease lobe alone suffices for stimulating acquisition, only full-length Cas9 responses to crRNA levels to boost acquisition in cells with low immunity depth. Finally, we show that this activity is evolutionarily conserved across multiple type II-C Cas9 orthologs. Altogether, we establish an auto-replenishing feedback mechanism in which apoCas9 safeguards CRISPR immunity depth by acting as both a crRNA sensor and a regulator of spacer acquisition.

DOI: 10.1038/s41586-025-09577-9

Source: https://www.nature.com/articles/s41586-025-09577-9