National Natural Science Foundation of China (NSFC)
32270761
中国
National Natural Science Foundation of China (NSFC)
31741027
中国
National Natural Science Foundation of China (NSFC)
81672722
中国
Ministry of Science and Technology (MoST, China)
2022YFC2303700
中国
Ministry of Science and Technology (MoST, China)
2021YFA301900
中国
National Natural Science Foundation of China (NSFC)
32222040
中国
National Natural Science Foundation of China (NSFC)
32070049
中国
Other government
2022M712272
中国
引用
ジャーナル: Innovation (Camb) / 年: 2023 タイトル: DnaQ mediates directional spacer acquisition in the CRISPR-Cas system by a time-dependent mechanism. 著者: Dongmei Tang / Tingting Jia / Yongbo Luo / Biqin Mou / Jie Cheng / Shiqian Qi / Shaohua Yao / Zhaoming Su / Yamei Yu / Qiang Chen / 要旨: In the spacer acquisition stage of CRISPR-Cas immunity, spacer orientation and protospacer adjacent motif (PAM) removal are two prerequisites for functional spacer integration. Cas4 has been ...In the spacer acquisition stage of CRISPR-Cas immunity, spacer orientation and protospacer adjacent motif (PAM) removal are two prerequisites for functional spacer integration. Cas4 has been implicated in both processing the prespacer and determining the spacer orientation. In Cas4-lacking systems, host 3'-5' DnaQ family exonucleases were recently reported to play a Cas4-like role. However, the molecular details of DnaQ functions remain elusive. Here, we characterized the spacer acquisition of the adaptation module of the type I-E system, in which a DnaQ domain naturally fuses with Cas2. We presented X-ray crystal structures and cryo-electron microscopy structures of this adaptation module. Our biochemical data showed that DnaQ trimmed PAM-containing and PAM-deficient overhangs with different efficiencies. Based on these results, we proposed a time-dependent model for DnaQ-mediated spacer acquisition to elucidate PAM removal and spacer orientation determination in Cas4-lacking CRISPR-Cas systems.