小柯机器人

美国加州大学伯克利分校Jennifer A. Doudna团队通过CRISPR修剪器-整合酶进行基因组扩展。2023年6月14日，《自然》杂志在线发表了这项成果。

研究人员展示了I-E型系统中一个优雅的替代途径，其使用内部的DnaQ样外切酶（DEDDh）来选择和处理DNA，以便使用原间隔基序（PAM）进行整合。天然的Cas1-Cas2/外切酶融合（修剪器-整合酶）催化协调的DNA捕获、修剪和整合。CRISPR修剪器-整合酶的五个冷冻电镜结构，在DNA整合之前和期间都是可视化的，显示了不对称处理如何产生尺寸确定的、含有PAM的底物。在基因组整合之前，PAM序列被Cas1释放并被外切酶切割，将插入的DNA标记为自身，并防止CRISPR对宿主的反常定位。这些数据共同支持一个模型，即缺乏Cas4的CRISPR系统使用融合或招募的种外切酶来忠实地获得新的CRISPR免疫序列。

据介绍，CRISPR-Cas适应性免疫系统从入侵的移动遗传元件中捕获DNA片段，并将其整合到宿主基因组中，为RNA引导的免疫提供模板。CRISPR系统通过区分自我和非自我来维持基因组的完整性并避免自身免疫，在这个过程中，CRISPR/Cas1-Cas2整合酶是必要的，但不是充分的。在一些微生物中，Cas4内切酶协助CRISPR适应，但许多CRISPR-Cas系统缺乏Cas4。

附：英文原文

Title: Genome expansion by a CRISPR trimmer-integrase

Author: Wang, Joy Y., Tuck, Owen T., Skopintsev, Petr, Soczek, Katarzyna M., Li, Gary, Al-Shayeb, Basem, Zhou, Julia, Doudna, Jennifer A.

Issue&Volume: 2023-06-14

Abstract: CRISPR–Cas adaptive immune systems capture DNA fragments from invading mobile genetic elements and integrate them into the host genome to provide a template for RNA-guided immunity1. CRISPR systems maintain genome integrity and avoid autoimmunity by distinguishing between self and non-self, a process for which the CRISPR/Cas1–Cas2 integrase is necessary but not sufficient2,3,4,5. In some microorganisms, the Cas4 endonuclease assists CRISPR adaptation6,7, but many CRISPR–Cas systems lack Cas48. Here we show here that an elegant alternative pathway in a type I-E system uses an internal DnaQ-like exonuclease (DEDDh) to select and process DNA for integration using the protospacer adjacent motif (PAM). The natural Cas1–Cas2/exonuclease fusion (trimmer-integrase) catalyses coordinated DNA capture, trimming and integration. Five cryo-electron microscopy structures of the CRISPR trimmer-integrase, visualized both before and during DNA integration, show how asymmetric processing generates size-defined, PAM-containing substrates. Before genome integration, the PAM sequence is released by Cas1 and cleaved by the exonuclease, marking inserted DNA as self and preventing aberrant CRISPR targeting of the host. Together, these data support a model in which CRISPR systems lacking Cas4 use fused or recruited9,10 exonucleases for faithful acquisition of new CRISPR immune sequences.

DOI: 10.1038/s41586-023-06178-2

Source: https://www.nature.com/articles/s41586-023-06178-2

Nature：《自然》，创刊于1869年。隶属于施普林格·自然出版集团，最新IF：69.504
官方网址：http://www.nature.com/
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