Two HEPN domains dictate CRISPR RNA maturation and target cleavage in Cas13d

Bo Zhang, Yangmiao Ye, Weiwei Ye, Vanja Perčulija, Han Jiang, Yiyang Chen, Yu Li, Jing Chen, Jinying Lin, Siqi Wang, Qi Chen, Yu-San Han and Songying Ouyang ()
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Bo Zhang: Fujian Normal University
Yangmiao Ye: Fujian Normal University
Weiwei Ye: Fujian Normal University
Vanja Perčulija: Fujian Normal University
Han Jiang: Fujian Normal University
Yiyang Chen: Fujian Normal University
Yu Li: Fujian Normal University
Jing Chen: Fujian Normal University
Jinying Lin: Fujian Normal University
Siqi Wang: Fujian Normal University
Qi Chen: Fujian Normal University
Yu-San Han: National Taiwan University
Songying Ouyang: Fujian Normal University

Nature Communications, 2019, vol. 10, issue 1, 1-11

Abstract: Abstract Cas13d, the type VI-D CRISPR-Cas effector, is an RNA-guided ribonuclease that has been repurposed to edit RNA in a programmable manner. Here we report the detailed structural and functional analysis of the uncultured Ruminococcus sp. Cas13d (UrCas13d)-crRNA complex. Two hydrated Mg2+ ions aid in stabilizing the conformation of the crRNA repeat region. Sequestration of divalent metal ions does not alter pre-crRNA processing, but abolishes target cleavage by UrCas13d. Notably, the pre-crRNA processing is executed by the HEPN-2 domain. Furthermore, both the structure and sequence of the nucleotides U(-8)-C(-1) within the repeat region are indispensable for target cleavage, and are specifically recognized by UrCas13d. Moreover, correct base pairings within two separate spacer regions (an internal and a 3′-end region) are essential for target cleavage. These findings provide a framework for the development of Cas13d into a tool for a wide range of applications.

Date: 2019
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Persistent link: https://EconPapers.repec.org/RePEc:nat:natcom:v:10:y:2019:i:1:d:10.1038_s41467-019-10507-3

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DOI: 10.1038/s41467-019-10507-3

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