CRISPR-Cas III-A Csm6 CARF Domain Is a Ring Nuclease Triggering Stepwise cA4 Cleavage with ApA>p Formation Terminating RNase Activity

Jia, Ning; Jones, Roger; Yang, Guangli; Ouerfelli, Ouathek; Patel, Dinshaw J.

doi:10.1016/j.molcel.2019.06.014

Title: CRISPR-Cas III-A Csm6 CARF Domain Is a Ring Nuclease Triggering Stepwise cA4 Cleavage with ApA>p Formation Terminating RNase Activity

Journal Article · Sun Sep 01 00:00:00 EDT 2019 · Molecular Cell

DOI:https://doi.org/10.1016/j.molcel.2019.06.014· OSTI ID:1658391

Jia, Ning; Jones, Roger; Yang, Guangli; Ouerfelli, Ouathek; Patel, Dinshaw J.

Type III-A CRISPR-Cas surveillance complexes containing multi-subunit Csm effector, guide, and target RNAs exhibit multiple activities, including formation of cyclic-oligoadenylates (cA_n) from ATP and subsequent cA_n-mediated cleavage of single-strand RNA (ssRNA) by the trans-acting Csm6 RNase. Our structure-function studies have focused on Thermococcus onnurineus Csm6 to deduce mechanistic insights into how cA₄ binding to the Csm6 CARF domain triggers the RNase activity of the Csm6 HEPN domain and what factors contribute to regulation of RNA cleavage activity. Furthermore, we demonstrate that the Csm6 CARF domain is a ring nuclease, whereby bound cA₄ is stepwise cleaved initially to ApApApA>p and subsequently to ApA>p in its CARF domain-binding pocket, with such cleavage bursts using a timer mechanism to regulate the RNase activity of the Csm6 HEPN domain. In addition, we establish T. onnurineus Csm6 as an adenosine-specific RNase and identify a histidine in the cA4 CARF-binding pocket involved in autoinhibitory regulation of RNase activity.

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Research Organization:: Argonne National Laboratory (ANL), Argonne, IL (United States). Advanced Photon Source (APS)

Sponsoring Organization:: USDOE Office of Science (SC); National Institutes of Health (NIH)

Grant/Contract Number:: AC02-06CH11357; P30CA008748; S10 RR029205; P30 GM124165

OSTI ID:: 1658391

Alternate ID(s):: OSTI ID: 1561311

Journal Information:: Molecular Cell, Journal Name: Molecular Cell Vol. 75 Journal Issue: 5; ISSN 1097-2765

Publisher:: Cell Press - ElsevierCopyright Statement

Country of Publication:: United States

Language:: English

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Cited by: 55 works

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Structure of Csx1-cOA4 complex reveals the basis of RNA decay in Type III-B CRISPR-Cas Molina, Rafael; Stella, Stefano; Feng, Mingxia Nature Communications, Vol. 10, Issue 1 https://doi.org/10.1038/s41467-019-12244-z	journal	September 2019
Structure and mechanism of a Type III CRISPR defence DNA nuclease activated by cyclic oligoadenylate McMahon, Stephen A.; Zhu, Wenlong; Graham, Shirley Nature Communications, Vol. 11, Issue 1 https://doi.org/10.1038/s41467-019-14222-x	journal	January 2020

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59 BASIC BIOLOGICAL SCIENCES
Type III-A CRISPR-Cas Csm
Csm6 CARF
ring nuclease
RNase activity
autoinhibition
Csm6
adenosine-specific RNase

Title: CRISPR-Cas III-A Csm6 CARF Domain Is a Ring Nuclease Triggering Stepwise cA4 Cleavage with ApA>p Formation Terminating RNase Activity

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