The diverse arsenal of type III CRISPR–Cas-associated CARF and SAVED effectors

Steens, Jurre A.; Salazar, Carl Raymund P.; Staals, Raymond H.J.

doi:10.1042/bst20220289

Cited by 28 publications

(29 citation statements)

References 98 publications

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“…The auxiliary effector proteins encoded in the vicinity of the CRISPR-Cas locus are typically composed of the cA n -sensing CRISPR-associated Rossmann fold (CARF) or SMODS-associated and fused to various effector domains (SAVED) (SMODS-associated) domains fused to a wide variety of effector domains including higher eukaryotes and prokaryotes nucleotide-binding (HEPN)-ribonucleases (Csm6 and Csx1), DNA nucleases (Can1, Can2, and Card1), protease or transcriptional regulator Csa3 ( 4 ). The CARF7 clade of the CARF domain family shares a conserved CARF-wHTH core that is often fused to the RelE ribonuclease-like domain identified as a toxin in the type II toxin-antitoxin RelE-RelB family ( 5 ).…”

mentioning

confidence: 99%

Ribosomal stalk-captured CARF-RelE ribonuclease inhibits translation following CRISPR signaling

Mogila,

Tamulaitiene,

Keda

et al. 2023

Science

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Prokaryotic type III CRISPR-Cas antiviral systems employ cyclic oligoadenylate (cA n ) signaling to activate a diverse range of auxiliary proteins that reinforce the CRISPR-Cas defense. Here we characterize a class of cA n -dependent effector proteins named CRISPR-Cas-associated messenger RNA (mRNA) interferase 1 (Cami1) consisting of a CRISPR-associated Rossmann fold sensor domain fused to winged helix-turn-helix and a RelE-family mRNA interferase domain. Upon activation by cyclic tetra-adenylate (cA 4 ), Cami1 cleaves mRNA exposed at the ribosomal A-site thereby depleting mRNA and leading to cell growth arrest. The structures of apo-Cami1 and the ribosome-bound Cami1-cA 4 complex delineate the conformational changes that lead to Cami1 activation and the mechanism of Cami1 binding to a bacterial ribosome, revealing unexpected parallels with eukaryotic ribosome-inactivating proteins.

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mentioning

confidence: 99%

Ribosomal stalk-captured CARF-RelE ribonuclease inhibits translation following CRISPR signaling

Mogila,

Tamulaitiene,

Keda

et al. 2023

Science

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“…As a result, LKMFCA had an increased potential to scavenge its environment for nutrients and to efflux toxins out of its cell to survive (85). Interestingly, at the time of environmental transition, LKMFCA gained many genes participating in resistance to viruses, particularly those of type I and III cas , and CARF systems (86, 87). Given that both environments are hyperthermophilic, it is unclear exactly why these genes became enriched in hot spring Korarchaeota.…”

Section: Resultsmentioning

confidence: 99%

Phylogenomics and ancestral reconstruction of Korarchaeota reveals genomic adaptation to habitat switching

Tahon,

Köstlbacher,

Pelve

et al. 2023

Preprint

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Our knowledge of archaeal diversity and evolution has expanded rapidly in the past decade. However, hardly any genomes of the phylum Korarchaeota have been obtained due to the difficulty in accessing their natural habitats and – possibly – their limited abundance. As a result, many aspects of Korarchaeota biology, physiology and evolution remain enigmatic. Here, we expand this phylum with five high-quality metagenome-assembled genomes. This improved taxon sampling combined with sophisticated phylogenomic analyses robustly places Korarchaeota at the base of TACK and Asgard clades, revisiting the phylum’s long-assumed position. Furthermore, we observe a clear split between terrestrial and marine thermal clades. Gene tree-aware ancestral reconstructions suggest that the last Korarchaeota common ancestor was a thermophilic autotroph. In contrast, Korarchaeaceae, the lineage where environmental transitions occurred, shifted towards a heterotrophic lifestyle. Terrestrial Korarchaeota gained manycasand CARF genes indicating they may need to manage viral infections. Together, our study provides new insights into these early diverging Archaea and suggests that gradual gene gain and loss shaped their adaptation to different thermal environments.ImportanceKorarchaeota are an ancient group of archaea, but their biology, physiology and evolution have remained obscure. Analysis of five novel Korarchaeota MAGs, and publicly available reference data provides robust phylogenomic evidence that Korarchaeota are placed at the base of Asgard archaea and TACK, revisiting the phylum’s long-assumed position. Gene content reconstruction suggests a versatile thermophilic and autotrophic last Korarchaeota common ancestor. Environmental distribution surveying of public databases places all Korarchaeota in thermophilic environments and indicates that their habitat is limited to hydrothermal vents and hot springs. Our modeling indicates at least two transitions linked to habitat switching between these environments in the evolutionary history of Korarchaeota. Both are linked to a significant alteration of the inferred ancestral gene content, including a shift towards a heterotrophic and potential scavenging lifestyle. Furthermore, hot spring Korarchaeota acquired various genes participating in resistance to viruses, suggesting they may need to manage frequent viral threats.

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“…The cyclic oligoadenylate signaling mechanism of type III CRISPR-Cas systems is used to activate a broad spectrum of ancillary proteins, including CARF domain-containing effector nucleases (13,(43)(44)(45). As the nuclease activity of these enzymes leads to growth arrest and cell death upon persistent activation of the type III CRISPR-Cas system (28,46), this necessitates stringent control of the cOAdependent activation mechanism.…”

Section: Discussionmentioning

confidence: 99%

Substrate selectivity and catalytic activation of the type III CRISPR ancillary nuclease Can2

Jungfer,

Sigg,

Jinek

2023

Preprint

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Type III CRISPR-Cas systems provide adaptive immunity against foreign mobile genetic elements through RNA-guided interference. Sequence-specific recognition of RNA targets by the type III effector complex triggers the generation of cyclic oligoadenylate (cOA) second messengers that activate ancillary effector proteins, thus reinforcing the host immune response. The ancillary nuclease Can2 is activated by cyclic tetra- AMP (cA4); however, the mechanisms underlying cA4-mediated activation and substrate selectivity remain elusive. Here we report crystal structures of Thermoanaerobacter brockii Can2 (TbrCan2) in substrate- and product-bound complexes. We show that TbrCan2 is a single strand-selective DNase and RNase that binds substrates via a conserved SxTTS active site motif, and reveal molecular interactions underpinning its sequence preference for CA dinucleotides. Furthermore, we identify a molecular interaction relay linking the cA4 binding and nuclease catalytic sites to enable divalent metal cation coordination and catalytic activation. These findings provide key insights into the molecular mechanisms of Can2 nucleases in type III CRISPR-Cas immunity and may guide their technological development for nucleic acid detection applications.

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The diverse arsenal of type III CRISPR–Cas-associated CARF and SAVED effectors

Cited by 28 publications

References 98 publications

Ribosomal stalk-captured CARF-RelE ribonuclease inhibits translation following CRISPR signaling

Ribosomal stalk-captured CARF-RelE ribonuclease inhibits translation following CRISPR signaling

Phylogenomics and ancestral reconstruction of Korarchaeota reveals genomic adaptation to habitat switching

Substrate selectivity and catalytic activation of the type III CRISPR ancillary nuclease Can2

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