Phage T7 DNA mimic protein Ocr is a potent inhibitor of BREX defence

Isaev, Artem; Drobiazko, Alena; Sierro, Nicolas; Gordeeva, Julia; Yosef, Ido; Qimron, Udi; Ivanov, Nikolai V.; Severinov, Konstantin

doi:10.1093/nar/gkaa290

Cited by 66 publications

(63 citation statements)

References 43 publications

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“…3B). Whole genome sequencing of 4 mutants revealed that all of them carried a mutation (T54V) in gene 0.3 which encodes Ocr, an anti-restriction protein that inhibits RM and BREX systems by mimicking the structure of DNA (46,47). In the presence of the system, the transformation efficiency of an Ocr-expressing plasmid was ~100-fold lower than in the absence of the system, while there was no difference when transforming a GFP-encoding plasmid as a control (Fig.…”

Section: Paris Triggers Growth Arrest Upon Sensing An Anti-restrictiomentioning

confidence: 99%

Prophage-encoded hotspots of bacterial immune systems

Rousset

Dowding

Bernheim

et al. 2021

Preprint

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The arms race between bacteria and phages led to the emergence of a variety of genetic systems used by bacteria to defend against viral infection, some of which were repurposed as powerful biotechnological tools. While numerous defense systems have been identified in genomic regions termed defense islands, it is believed that many more remain to be discovered. Here, we show that P2- like prophages and their P4-like satellites have genomic hotspots that represent a significant source of novel anti-phage systems. We validate the defense activity of 14 systems spanning various protein domains and describe PARIS, an abortive infection system triggered by a phage-encoded anti-restriction protein. Immunity hotspots are present across prophages of distant bacterial species, highlighting their biological importance in the competition between bacteria and phages.

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Section: Paris Triggers Growth Arrest Upon Sensing An Anti-restrictiomentioning

confidence: 99%

Prophage-encoded hotspots of bacterial immune systems

Rousset

Dowding

Bernheim

et al. 2021

Preprint

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“…For example, the T7 phage encoded Ocr protein is highly acidic and forms a dimer with a bend similar to B-DNA ( 48 , 49 ). Ocr was initially discovered as a Type I restriction enzyme system inhibitor and was more recently shown to inhibit the anti-phage system, BREX ( 50 ). Importantly, systematic mutation of Ocr's acidic residues revealed it to be highly recalcitrant to breakage, similar to AcrIF2*, maintaining inhibitory activity against Type I R-M even with up to 33% of acidic residues mutated ( 48 ).…”

Section: Discussionmentioning

confidence: 99%

Mobile element warfare via CRISPR and anti-CRISPR in Pseudomonas aeruginosa

León

Park

Borges

et al. 2021

Nucleic Acids Research

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Bacteria deploy multiple defenses to prevent mobile genetic element (MGEs) invasion. CRISPR–Cas immune systems use RNA-guided nucleases to target MGEs, which counter with anti-CRISPR (Acr) proteins. Our understanding of the biology and co-evolutionary dynamics of the common Type I-C CRISPR–Cas subtype has lagged because it lacks an in vivo phage-host model system. Here, we show the anti-phage function of a Pseudomonas aeruginosa Type I-C CRISPR–Cas system encoded on a conjugative pKLC102 island, and its Acr-mediated inhibition by distinct MGEs. Seven genes with anti-Type I-C function (acrIC genes) were identified, many with highly acidic amino acid content, including previously described DNA mimic AcrIF2. Four of the acr genes were broad spectrum, also inhibiting I-E or I-F P. aeruginosa CRISPR–Cas subtypes. Dual inhibition comes at a cost, however, as simultaneous expression of Type I-C and I-F systems renders phages expressing the dual inhibitor AcrIF2 more sensitive to targeting. Mutagenesis of numerous acidic residues in AcrIF2 did not impair anti-I-C or anti-I-F function per se but did exacerbate inhibition defects during competition, suggesting that excess negative charge may buffer DNA mimics against competition. Like AcrIF2, five of the Acr proteins block Cascade from binding DNA, while two function downstream, likely preventing Cas3 recruitment or activity. One such inhibitor, AcrIC3, is found in an ‘anti-Cas3’ cluster within conjugative elements, encoded alongside bona fide Cas3 inhibitors AcrIF3 and AcrIE1. Our findings demonstrate an active battle between an MGE-encoded CRISPR–Cas system and its diverse MGE targets.

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“…Recently another microbial self-defense mechanism, called Bacteriophage Exclusion (BREX) system has been identified [ 71 ]. Although the exact mechanism of action is not well understood, BREX is thought to methylate host DNA asymmetrically to distinguish host DNA from newly acquired phage integration [ 72 ]. The BREX system is usually composed of a core system consisting of proteins containing an ATPase domain, a DNA methyltransferase domain and phosphatase domain [ 71 ].…”

Section: Discussionmentioning

confidence: 99%

Hybrid genome de novo assembly with methylome analysis of the anaerobic thermophilic subsurface bacterium Thermanaerosceptrum fracticalcis strain DRI-13T

2021

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Background There is a dearth of sequenced and closed microbial genomes from environments that exceed > 500 m below level terrestrial surface. Coupled with even fewer cultured isolates, study and understanding of how life endures in the extreme oligotrophic subsurface environments is greatly hindered. Using a de novo hybrid assembly of Illumina and Oxford Nanopore sequences we produced a circular genome with corresponding methylome profile of the recently characterized thermophilic, anaerobic, and fumarate-respiring subsurface bacterium, Thermanaerosceptrum fracticalcis, strain DRI-13T to understand how this microorganism survives the deep subsurface. Results The hybrid assembly produced a single circular genome of 3.8 Mb in length with an overall GC content of 45%. Out of the total 4022 annotated genes, 3884 are protein coding, 87 are RNA encoding genes, and the remaining 51 genes were associated with regulatory features of the genome including riboswitches and T-box leader sequences. Approximately 24% of the protein coding genes were hypothetical. Analysis of strain DRI-13T genome revealed: 1) energy conservation by bifurcation hydrogenase when growing on fumarate, 2) four novel bacterial prophages, 3) methylation profile including 76.4% N6-methyladenine and 3.81% 5-methylcytosine corresponding to novel DNA methyltransferase motifs. As well a cluster of 45 genes of unknown protein families that have enriched DNA mCpG proximal to the transcription start sites, and 4) discovery of a putative core of bacteriophage exclusion (BREX) genes surrounded by hypothetical proteins, with predicted functions as helicases, nucleases, and exonucleases. Conclusions The de novo hybrid assembly of strain DRI-13T genome has provided a more contiguous and accurate view of the subsurface bacterium T. fracticalcis, strain DRI-13T. This genome analysis reveals a physiological focus supporting syntrophy, non-homologous double stranded DNA repair, mobility/adherence/chemotaxis, unique methylome profile/recognized motifs, and a BREX defense system. The key to microbial subsurface survival may not rest on genetic diversity, but rather through specific syntrophy niches and novel methylation strategies.

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Phage T7 DNA mimic protein Ocr is a potent inhibitor of BREX defence

Cited by 66 publications

References 43 publications

Prophage-encoded hotspots of bacterial immune systems

Prophage-encoded hotspots of bacterial immune systems

Mobile element warfare via CRISPR and anti-CRISPR in Pseudomonas aeruginosa

Hybrid genome de novo assembly with methylome analysis of the anaerobic thermophilic subsurface bacterium Thermanaerosceptrum fracticalcis strain DRI-13T

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