Filament formation activates protease and ring nuclease activities of CRISPR SAVED-Lon

Smalakyte, Dalia; Ruksenaite, Audrone; Sasnauskas, Giedrius; Tamulaitiene, Giedre; Tamulaitis, Gintautas

doi:10.1101/2024.05.08.593097

Cited by 2 publications

References 77 publications

(127 reference statements)

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The SAVED domain of the type III CRISPR protease CalpL is a ring nuclease

Binder,

Schneberger,

Schmitz

et al. 2024

Nucleic Acids Research

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Prokaryotic CRISPR-Cas immune systems detect and cleave foreign nucleic acids. In type III CRISPR-Cas systems, the Cas10 subunit of the activated recognition complex synthesizes cyclic oligoadenylates (cOAs), second messengers that activate downstream ancillary effector proteins. Once the viral attack has been weathered, elimination of extant cOA is essential to limit the antiviral response and to allow cellular recovery. Various families of ring nucleases have been identified, specializing in the degradation of cOAs either as standalone enzymes or as domains of effector proteins. Here we describe the ring nuclease activity inherent in the SAVED domain of the cA4-activated CRISPR Lon protease CalpL. We characterize the kinetics of cA4 cleavage and identify key catalytic residues. We demonstrate that cA4-induced oligomerization of CalpL is essential not only for activation of the protease, but is also required for nuclease activity. Further, the nuclease activity of CalpL poses a limitation to the protease reaction, indicating a mechanism for regulation of the CalpL/T/S signaling cascade. This work is the first demonstration of a catalytic SAVED domain and gives new insights into the dynamics of transcriptional adaption in CRISPR defense systems.

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The SAVED domain of the type III CRISPR protease CalpL is a ring nuclease

Binder,

Schneberger,

Schmitz

et al. 2024

Nucleic Acids Research

View full text Add to dashboard Cite

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Diversity and abundance of ring nucleases in type III CRISPR-Cas loci

Hoikkala,

Chi,

Grüschow

et al. 2024

Preprint

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Most type III CRISPR-Cas systems facilitate immune responses against invading mobile genetic elements such as phages by generating cyclic oligoadenylates (cOAs). Downstream effectors activated by cOAs are typically non-specific proteins that induce damage to essential cellular components, thereby preventing phage epidemics. Due to these toxic effects, it is crucial that the production and concentration of cOAs remain under tight regulatory control during infection-free periods or when deactivating the immune response after clearing an infection. Type III CRISPR loci often encode enzymes known as ring nucleases (RNs) that bind and degrade specific cOAs, while some effectors are auto-deactivating. Despite the discovery of several classes of RNs, a comprehensive bioinformatic analysis of type III CRISPR-Cas loci in this context is lacking. Here, we examined 38,742 prokaryotic genomes to provide a global overview of type III CRISPR loci, focussing on the known and predicted RNs. The candidate RNs Csx16 and Csx20 are confirmed as active enzymes, joining Crn1-3. Distributions and patterns of co-occurrence of RNs and associated effectors are explored, allowing the conclusion that a sizeable majority of type III CRISPR systems regulate cOA levels by degrading the signalling molecules, which has implications for cell fate following viral infection.

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Filament formation activates protease and ring nuclease activities of CRISPR SAVED-Lon

Cited by 2 publications

References 77 publications

The SAVED domain of the type III CRISPR protease CalpL is a ring nuclease

The SAVED domain of the type III CRISPR protease CalpL is a ring nuclease

Diversity and abundance of ring nucleases in type III CRISPR-Cas loci

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