DNA targeting and interference by a bacterial Argonaute nuclease

Kuzmenko, Anton; Oguienko, Anastasiya; Esyunina, Daria; Yudin, Denis; Petrova, Mayya; Kudinova, A. G.; Маслова, О. А.; Ninova, Maria; Ryazansky, Sergei; Leach, David R. F.; Aravin, Alexei A.; Kulbachinskiy, Andrey

doi:10.1038/s41586-020-2605-1

Cited by 141 publications

(230 citation statements)

References 46 publications

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“…However, when normalized to the length of corresponding replicons, the ratio of smDNAs mapping to the plasmid and chromosomal DNA (24 and 44 in the two replicas) roughly equaled the known copy number of the plasmid vector (pET28, about 20 copies per cell). Thus, unlike previously studied pAgos (8,17,20), KmAgo-bound guides are not enriched in sequences derived from plasmid DNA.…”

Section: Resultscontrasting

confidence: 76%

“…Previously studied pAgo proteins were shown to bind small guide nucleic acids when expressed in their native hosts or in the heterologous E. coli system (8,13,14,17,20,22). To explore if KmAgo is associated with guide nucleic acids when expressed in E. coli cells, we extracted and labeled nucleic acids from the KmAgo samples purified by metal-affinity chromatography.…”

Section: Resultsmentioning

confidence: 99%

“…Similarly to eukaryotic Agos, pAgos have been implicated in the cell’s defense against mobile genetic elements (1,4,6). Recently, CbAgo was shown to protect bacterial cells from invaders, including plasmids and phages (17). Similarly, thermophilic TtAgo and PfAgo were shown to decrease plasmid DNA content and transformation efficiency (7,8).…”

Section: Introductionmentioning

confidence: 99%

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A programmable pAgo nuclease with universal guide and target specificity from the mesophilic bacteriumKurthia massiliensis

Kropocheva

Kuzmenko

Aravin

et al. 2021

Preprint

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Argonaute proteins are programmable nucleases that are found in both eukaryotes and prokaryotes and provide defense against invading genetic elements. Although some prokaryotic Argonautes (pAgos) were shown to recognize RNA targets in vitro, the majority of studied pAgos have strict specificity toward DNA, which limits their practical use in RNA-centric applications. Here, we describe a unique KmAgo nuclease from the mesophilic bacterium Kurthia massiliensis that can be programmed with either DNA or RNA guides and can precisely cleave both DNA and RNA targets. KmAgo preferentially binds 16-20 nt long 5'-phosphorylated guide molecules with no strict specificity towards their sequence and is active in a wide range of temperatures. In bacterial cells, KmAgo is loaded with small DNAs with no obvious sequence preferences suggesting that it can uniformly target genomic sequences. Target cleavage by KmAgo depends on the formation of secondary structure indicating that KmAgo can be used for structural probing of RNA targets. Mismatches between the guide and target sequences greatly affect the efficiency and precision of target cleavage, depending on the mismatch position and the nature of the reacting nucleic acid. These properties of KmAgo open the way for its use for highly specific nucleic acid detection and cleavage.

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Section: Resultscontrasting

confidence: 76%

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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A programmable pAgo nuclease with universal guide and target specificity from the mesophilic bacteriumKurthia massiliensis

Kropocheva

Kuzmenko

Aravin

et al. 2021

Preprint

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“…Of particular interest is the Argonaute homolog and restriction endonuclease that function as viral and mobile element defense mechanisms [87, 88]. Recently, a bacterial Argonaute nuclease from Clostridium butyricum was shown to target multicopy genetic elements and suppress the propagation of plasmids and infection by phages via DNA interference [89]. In addition to the RM systems, this diverse suite of genes in Nasonia specifically from Nashville suggests enhanced protection against viral infection.…”

Section: Resultsmentioning

confidence: 99%

Genomes of gut bacteria fromNasoniawasps shed light on phylosymbiosis and microbe-assisted hybrid breakdown

Cross

Leigh

Hatmaker

et al. 2021

Preprint

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Phylosymbiosis is a cross-system trend whereby microbial community relationships recapitulate the host phylogeny. In Nasonia parasitoid wasps, phylosymbiosis occurs throughout development, is distinguishable between sexes, and benefits host development and survival. Moreover, the microbiome shifts in hybrids as a rare Proteus bacteria in the microbiome becomes dominant. The larval hybrids then catastrophically succumb to bacterial-assisted lethality and reproductive isolation between the species. Two important questions for understanding phylosymbiosis and bacterial-assisted lethality in hybrids are: (i) Do the Nasonia bacterial genomes differ from other animal isolates and (ii) Are the hybrid bacterial genomes the same as those in the parental species? Here we report the cultivation, whole genome sequencing, and comparative analyses of the most abundant gut bacteria in Nasonia larvae, Providencia rettgeri and Proteus mirabilis. Characterization of new isolates shows Proteus mirabilis forms a more robust biofilm than Providencia rettgeri and when grown in co-culture, Proteus mirabilis significantly outcompetes Providencia rettgeri. Providencia rettgeri genomes from Nasonia are similar to each other and more divergent to pathogenic, human associates. Proteus mirabilis from N. vitripennis, N. giraulti, and their hybrid offspring are nearly identical and relatively distinct from human isolates based. These results indicate that members of the larval gut microbiome within Nasonia are most similar to each other, and the strain of the dominant Proteus mirabilis in hybrids is resident in parental species. Holobiont interactions between shared, resident members of the wasp microbiome and the host underpin phylosymbiosis and hybrid breakdown.

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“…The best understood prokaryotic Ago (pAgo) proteins are the so called full-length pAgos, which are composed of N, PAZ, MID and PIWI domains, and thus closely resemble eAgo proteins. It has been demonstrated that fulllength pAgos function as prokaryotic antiviral systems, with the PIWI domain performing cleavage of invading nucleic acids [7]. Furthermore, a function beyond immunity -decatenation of circular chromosomes during replication -has also been documented [8].…”

Section: Introductionmentioning

confidence: 99%

Prokaryotic Argonaute From Archaeoglobus Fulgidus Interacts With DNA as a Homodimer

Golovinas

Rutkauskas

Manakova

et al. 2020

Preprint

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Argonaute (Ago) proteins are found in all three domains of life. The best characterized group is eukaryotic Argonautes (eAgos), which are the core of RNA interference. The best understood prokaryotic Ago (pAgo) proteins are full-length pAgos. They are composed of four major structural/functional domains (N, PAZ, MID and PIWI) and thereby closely resemble eAgos. It was demonstrated that full-length pAgos function as prokaryotic antiviral systems, with the PIWI domain performing cleavage of invading nucleic acids. However, the majority of identified pAgos are shorter and catalytically inactive (encode just MID and inactive PIWI domains), thus their action mechanism and function remain unknown. In this work we focus on AfAgo, a short pAgo protein encoded by an archaeon Archaeoglobus fulgidus. We find that in all previously solved AfAgo structures, its two monomers form substantial dimerization interfaces involving the C-terminal β-sheets. Led by this finding, we have employed various biochemical and biophysical assays, including SEC-MALS, SAXS, single-molecule FRET and AFM, to show that AfAgo is indeed a homodimer in solution, which is capable of simultaneous interaction with two DNA molecules. This finding underscores the diversity of prokaryotic Agos and broadens the range of currently known Argonaute-nucleic acid interaction mechanisms.

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DNA targeting and interference by a bacterial Argonaute nuclease

Cited by 141 publications

References 46 publications

A programmable pAgo nuclease with universal guide and target specificity from the mesophilic bacteriumKurthia massiliensis

A programmable pAgo nuclease with universal guide and target specificity from the mesophilic bacteriumKurthia massiliensis

Genomes of gut bacteria fromNasoniawasps shed light on phylosymbiosis and microbe-assisted hybrid breakdown

Prokaryotic Argonaute From Archaeoglobus Fulgidus Interacts With DNA as a Homodimer

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DNA targeting and interference by a bacterial Argonaute nuclease

Cited by 141 publications

References 46 publications

A programmable pAgo nuclease with universal guide and target specificity from the mesophilic bacteriumKurthia massiliensis

A programmable pAgo nuclease with universal guide and target specificity from the mesophilic bacteriumKurthia massiliensis

Genomes of gut bacteria fromNasoniawasps shed light on phylosymbiosis and microbe-assisted hybrid breakdown

Prokaryotic Argonaute From Archaeoglobus Fulgidus Interacts With DNA&nbsp;as a Homodimer

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Prokaryotic Argonaute From Archaeoglobus Fulgidus Interacts With DNA as a Homodimer