Volume 33, No 3, Mar 2023

ISSN: 1001-0602 
EISSN: 1748-7838 2018 
impact factor 17.848* 
(Clarivate Analytics, 2019)

Volume 33 Issue 3, March 2023: 229-244

ORIGINAL ARTICLES

The compact Casπ (Cas12l) ‘bracelet’ provides a unique structural platform for DNA manipulation

Ao Sun^1,2,† , Cheng-Ping Li^1,2,† , Zhihang Chen^1,2,† , Shouyue Zhang^1,2,† , Dan-Yuan Li^1,2,† , Yun Yang^1,2 , Long-Qi Li^1,2 , Yuqian Zhao^1,2 , Kaichen Wang² , Zhaofu Li¹ , Jinxia Liu^3,4 , Sitong Liu^3,4 , Jia Wang^1,2,* , Jun-Jie Gogo Liu^1,2,*

¹Beijing Advanced Innovation Center for Structural Biology & Frontier Research Center for Biological Structure, School of Life Sciences, Tsinghua University, Beijing, China
²Tsinghua-Peking Center for Life Sciences, School of Life Sciences, Tsinghua University, Beijing, China
³Department of Environmental Engineering, Peking University, Beijing, China
⁴Key Laboratory of Water and Sediment Sciences, Ministry of Education of China, Beijing, China
^†These authors contributed equally: Ao Sun, Cheng-Ping Li, Zhihang Chen, Shouyue Zhang
Correspondence: Jia Wang(wangjia2016@tsinghua.edu.cn)Jun-Jie Gogo Liu(junjiegogoliu@tsinghua.edu.cn)

CRISPR-Cas modules serve as the adaptive nucleic acid immune systems for prokaryotes, and provide versatile tools for nucleic acid manipulation in various organisms. Here, we discovered a new miniature type V system, CRISPR-Casπ (Cas12l) (~860 aa), from the environmental metagenome. Complexed with a large guide RNA (~170 nt) comprising the tracrRNA and crRNA, Casπ (Cas12l) recognizes a unique 5′ C-rich PAM for DNA cleavage under a broad range of biochemical conditions, and generates gene editing in mammalian cells. Cryo-EM study reveals a ‘bracelet’ architecture of Casπ effector encircling the DNA target at 3.4 Å resolution, substantially different from the canonical ‘two-lobe’ architectures of Cas12 and Cas9 nucleases. The large guide RNA serves as a ‘two-arm’ scaffold for effector assembly. Our study expands the knowledge of DNA targeting mechanisms by CRISPR effectors, and offers an efficient but compact platform for DNA manipulation.

https://doi.org/10.1038/s41422-022-00771-2

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