CRISPR-Cas III-A Csm6 CARF domain is a ring nuclease triggering stepwise cA(4) cleavage with ApA>p formation terminating RNase activity Journal Article


Authors: Jia, N.; Jones, R.; Yang, G.; Ouerfelli, O.; Patel, D. J.
Article Title: CRISPR-Cas III-A Csm6 CARF domain is a ring nuclease triggering stepwise cA(4) cleavage with ApA>p formation terminating RNase activity
Abstract: Type III-A CRISPR-Cas surveillance complexes containing multi-subunit Csm effector, guide, and target RNAs exhibit multiple activities, including formation of cyclic-oligoadenylates (cAn) from ATP and subsequent cAn-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. 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. © 2019 Elsevier Inc. In structure-function studies on CRISPR-Cas Csm6 RNase, Jia et al. demonstrate that its CARF domain is a ring nuclease, triggering stepwise cleavage of bound cA4 into ApApApA>p and ApA>p, using a timer mechanism to regulate HEPN domain RNase activity. We also establish that CARF domain His132 plays an autoinhibitory role in controlling RNase activity. © 2019 Elsevier Inc.
Keywords: type iii-a crispr-cas csm; a carf histidine contributes to autoinhibition; csm6 carf is a ring nuclease; csm6 is an adenosine-specific rnase; sequential cleavage bursts regulate rnase activity
Journal Title: Molecular Cell
Volume: 75
Issue: 5
ISSN: 1097-2765
Publisher: Cell Press  
Date Published: 2019-09-05
Start Page: 944
End Page: 956.e6
Language: English
DOI: 10.1016/j.molcel.2019.06.014
PUBMED: 31326273
PROVIDER: scopus
PMCID: PMC6731128
DOI/URL:
Notes: Source: Scopus
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MSK Authors
  1. Dinshaw J Patel
    424 Patel
  2. Guangli Yang
    25 Yang
  3. Ning Jia
    5 Jia