The Molecular Architecture for RNA-Guided RNA Cleavage by Cas13a

Liang Liu; Xueyan Li; Jun Ma; Zongqiang Li; Lilan You; Jiuyu Wang; Min Wang; Xinzheng Zhang; Yanli Wang

doi:10.1016/j.cell.2017.06.050

The Molecular Architecture for RNA-Guided RNA Cleavage by Cas13a

Cell. 2017 Aug 10;170(4):714-726.e10. doi: 10.1016/j.cell.2017.06.050. Epub 2017 Jul 27.

Authors

Liang Liu¹, Xueyan Li², Jun Ma³, Zongqiang Li¹, Lilan You², Jiuyu Wang¹, Min Wang¹, Xinzheng Zhang⁴, Yanli Wang⁵

Affiliations

¹ Key Laboratory of RNA Biology, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China.
² Key Laboratory of RNA Biology, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China; University of Chinese Academy of Sciences, Beijing 100049, China.
³ National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China.
⁴ University of Chinese Academy of Sciences, Beijing 100049, China; National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China; Center for Biological Imaging, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China. Electronic address: xzzhang@ibp.ac.cn.
⁵ Key Laboratory of RNA Biology, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China; University of Chinese Academy of Sciences, Beijing 100049, China; Collaborative Innovation Center of Genetics and Development, Shanghai 200438, China. Electronic address: ylwang@ibp.ac.cn.

PMID: 28757251
DOI: 10.1016/j.cell.2017.06.050

Abstract

Cas13a, a type VI-A CRISPR-Cas RNA-guided RNA ribonuclease, degrades invasive RNAs targeted by CRISPR RNA (crRNA) and has potential applications in RNA technology. To understand how Cas13a is activated to cleave RNA, we have determined the crystal structure of Leptotrichia buccalis (Lbu) Cas13a bound to crRNA and its target RNA, as well as the cryo-EM structure of the LbuCas13a-crRNA complex. The crRNA-target RNA duplex binds in a positively charged central channel of the nuclease (NUC) lobe, and Cas13a protein and crRNA undergo a significant conformational change upon target RNA binding. The guide-target RNA duplex formation triggers HEPN1 domain to move toward HEPN2 domain, activating the HEPN catalytic site of Cas13a protein, which subsequently cleaves both single-stranded target and collateral RNAs in a non-specific manner. These findings reveal how Cas13a of type VI CRISPR-Cas systems defend against RNA phages and set the stage for its development as a tool for RNA manipulation.

Keywords: C2c2; CRISPR-Cas; Cas13a; RNA-guided RNA cleavage; RNase.

MeSH terms

Bacterial Proteins / chemistry*
Bacterial Proteins / ultrastructure
Base Sequence
CRISPR-Associated Proteins / chemistry*
CRISPR-Associated Proteins / ultrastructure
CRISPR-Cas Systems*
Leptotrichia / chemistry
Leptotrichia / immunology*
Leptotrichia / metabolism
Leptotrichia / virology
Models, Molecular
RNA Processing, Post-Transcriptional
RNA, Bacterial / chemistry
RNA, Bacterial / genetics
RNA, Bacterial / ultrastructure
RNA, Guide, CRISPR-Cas Systems / chemistry
RNA, Guide, CRISPR-Cas Systems / genetics
RNA, Guide, CRISPR-Cas Systems / ultrastructure
RNA, Viral / chemistry
X-Ray Diffraction

Substances

Bacterial Proteins
CRISPR-Associated Proteins
RNA, Bacterial
RNA, Guide, CRISPR-Cas Systems
RNA, Viral