Biology subjects

Isaev, A.

Publications and source records attributed to Isaev, A..

11 recordsLinked to original sources

bioRxiv2026

Water-mass origin and transformation shapes seawater microbiomes in the northeastern Barents Sea and the St. Anna Trough

The St. Anna Trough region is a key Eurasian Arctic gateway connecting the northeastern Barents Sea, northern Kara Sea, and central Arctic Ocean. The region is characterized by complex hydrodynamics, with multiple water masses of local and Atlantic origin interacting and undergoing transformation. In this study, we investigated whether seawater microbiome composition correlates with water-mass origin and can be used to help delineate water masses. We collected 115 seawater samples representing major regional water masses - Arctic surface water, meltwater, cold dense water, Barents Sea branch water, the recently discovered Keldysh Current water, and mixed waters - and analyzed them using 16S rRNA gene amplicon sequencing. We found that microbiome composition closely followed water mass structure, which, together with thermohaline properties and depth, explained the majority (61.2%) of microbiome composition variation. Distinct microbial patterns were associated with Atlantic and Arctic water masses, except for the Arctic surface water, which lacked a distinct microbiome and resembled those of meltwater or the Keldysh Current microbiomes. Differential abundance and network analyses revealed characteristic bacterial markers: SAR11 (Clades Ia and IV), Roseibacillus, SV1-3 (Microbacteriaceae), SAR86 clade, and AEGEAN-169 marine group in the Keldysh Current; Halieaceae, Planktomarina, NS5 marine group, and Sulfitobacter in meltwater; SUP05 cluster, SAR86, SAR92, and SAR324 clade in cold dense water; Nitrincolaceae and Amylibacter in Barents Sea branch water; Flavobacteriaceae and Chitinophagales in mixed waters. These differences were driven primarily by shifts in relative abundance of widespread Arctic bacterial taxa rather than by differential prevalence of bacterial groups. Our results demonstrate that seawater microbiomes can serve as sensitive biological tracers of water mass origin and transformation in the Eurasian Arctic. Furthermore, the distinct Keldysh Current microbiome supports its recognition as a novel hydrographic feature and indicates its role as a biological corridor transporting Atlantic-associated biota eastward toward the Kara Sea. These findings suggest that "Atlantification" of Eurasian Arctic includes a microbial component, highlighting marine microbiomes as indicators of ongoing ecological changes in the Arctic Ocean.

microbiology↗

bioRxiv2026

H-NS silences antiviral immunity through 3D chromatin compaction

Bacterial antiviral immunity systems can be toxic to their hosts and must be tightly regulated. Immunity genes are often encoded by AT-rich mobile genetic elements subjected to silencing by nucleoid binding protein H-NS. Using RNA-seq, ChIP-seq, and chromosome 3D reconstruction with Micro-C, we find that H-NS binds and compacts bacterial immunity loci. H-NS deletion reveals anti-phage activity in model Escherichia coli strains generally considered phage-sensitive. Extending this approach to environmental isolates, we show that removal of H-NS silencing enhances defense, allowing bacteria to restrict phages with anti-defense proteins. We also discover Madara, a novel immunity system that complements the co-regulated BREX defense. Our results establish H-NS as a master regulator of bacterial immunity and highlight the importance of expression levels for anti-phage activity in native hosts.

molecular biology↗

bioRxiv2026

OLD sentinel: an abortive tRNase surveys phage replication and DNA defects in RecBCD-compromised cells

OLD, an abortive immunity protein from prophage P2, consists of an ABC ATPase sensor and a TOPRIM nuclease effector -- a core architecture shared by a large protein family, including components of antiphage systems Gabija, PARIS, Septu, and Lamassu. OLD was originally identified for its lethality in recB-deficient cells and inhibition of bacteriophage {lambda} infection, but the mechanisms governing its activation have remained elusive. Here, we present the cryo-EM structure of an inactive OLD tetramer and show that destabilization into dimeric form opens the TOPRIM catalytic site, stimulating tRNA cleavage. This activity arrests translation, a phenotype rescued by phage-encoded tRNAs. We demonstrate that OLD activation is not strictly RecBCD-dependent: OLD binds aberrant DNA structures in recB-deficient cells, but activation during infection requires recognition of single-stranded DNA hairpins at the phage replication origin. Collectively, our findings reveal how host and phage DNA processing factors create a complex landscape controlling OLD-mediated immunity.

molecular biology↗

bioRxiv2026

Phages infecting the common gut commensal Escherichia coli HS reveal tropism for its Klebsiella-like capsule

Tailed dsDNA phages are ubiquitous and thought to infect almost any bacterial species. Escherichia coli HS - a commensal strain used as a model in gut colonization studies - has not been reported as a host for dsDNA phages and is resistant to more than 100 coliphages from the BASEL collection. Here, we report the first phages infecting E. coli HS, characterize their interaction with an endogenous BREX system, and provide a detailed genomic and phylogenetic description. Phages {phi}HS1 (Queuovirinae) and {phi}HS2 (Ackermannviridae) possess dPreQ and 5-NeOmdU modifications that confer resistance to restriction digestion. {phi}HS3 is a temperate phage, related to the native HS prophage {phi}HS4, and represents a founding member of a novel genus (Hueyvirus) of P22-like phages capable of lateral transduction. We reveal exchange of tailspike genes between {phi}HS2, {phi}HS3, and Klebsiella-specific phages. Furthermore, we demonstrate that E. coli HS encodes a Klebsiella-like K47 capsule required for phage infection. These results suggest that the broad phage resistance of E. coli HS may be linked to its capsular type.

microbiology↗

bioRxiv2025

A census of anti-CRISPR proteins reveals AcrIE9 as an inhibitor of Escherichia coli K12 Type IE CRISPR-Cas system

CRISPR-Cas adaptive immunity systems provide defense against mobile genetic elements and are often countered by diverse anti-CRISPR (Acr) proteins. The Type IE CRISPR-Cas of Escherichia coli K12 has been a model for structural and functional studies and is a part of the species core genome. However, this system is transcriptionally silent, which has fueled questions about its true biological function. To clarify the role of this system in defense, we carried out a census of Acr proteins found in Enterobacterales and identified AcrIE9 as a potent inhibitor of the E. coli K12 Type IE CRISPR-Cas system. While sharing little sequence identity, AcrIE9 proteins from Pseudomonas and Escherichia both interact with the Cas7 subunit of the Cascade complex, thus preventing its binding to DNA. We further show that AcrIE9 is genetically linked to AcrIE10, forming the most widespread anti-CRISPR cluster in Enterobacterales, and this module often co-occurs with a novel HTH-like protein with unusual architecture.

microbiology↗

bioRxiv2025

Taxonomically different symbiotic communities of sympatric Arctic sponge species show functional similarity with specialization at species level

Marine sponges harbor diverse communities of associated organisms, including eukaryotes, viruses, and bacteria. Sponge associated microbiomes contribute to the health of the host organisms by defending them against invading bacteria and providing them with essential metabolites. Here we describe microbiomes of three sympatric species of cold-water marine sponges - Halichondria panicea, Halichondria sitiens, and Isodictya palmata - sampled over a period of six years at the White Sea. We identified the sponges as low microbial abundance species and detected stably associated bacteria that represent new taxa of sponge symbionts within Alpha- and Gammaproteobacteria. The sponges carried unique sets of unrelated species of symbiotic bacteria illustrating varying complexity of microbiomes. On a community level, sponge associated microbiomes shared common symbiotic features; they encoded multiple eukaryotic-like proteins, biosynthetic pathways, and transporters of amino acids and vitamins essential for sponges. On a species level, however, different classes of eukaryotic-like proteins and pathways were distributed between dominant and minor symbionts indicating specialization within microbiomes. Particularly, taurine and sulfoacetate metabolism pathways were associated exclusively with dominant symbionts in all three sponge species. Our study demonstrates strong functional convergence and co-evolution of microbiomes of sympatric cold-water sponge species with a distribution of functions between community members. Additionally, we observed dramatic shifts in compositions of sponge microbiomes coinciding with abnormally high water temperatures during the 2018 season, highlighting the vulnerability of cold-water ecosystems to global warming.

microbiology↗

bioRxiv2025

Specificity and Mechanism of tRNA cleavage by the AriB Toprim nuclease of the PARIS bacterial immune system

Transfer RNA molecules have been recently recognized as widespread targets of bacterial immune systems. Translation inhibition through tRNA cleavage or modification inhibits phage propagation, thereby protecting the bacterial population. To counteract this, some viruses encode their own tRNA molecules, allowing infection to take place. The AriB effector of the PARIS defence system is a Toprim nuclease previously shown to target the E. coli tRNALys(UUU), but not a tRNALys(UUU) variant encoded by bacteriophage T5. We demonstrate here that the T5-tRNALys(UUU) is required but not sufficient to bypass PARIS immunity. Combining tRNA-sequencing, genetics, phage infection and in vitro biochemical data, we reveal that the E. coli tRNAThr(UGU), is another prime target of AriB and tRNAAsn(GUU) represents a secondary, yet biologically relevant, target of the PARIS effector. Activated AriB protein cleaves these targets in vitro, and the cleavage reaction is not dependent on the presence of specific tRNA modifications. We show that the overexpression of phage T5 tRNALys(UUU), tRNAThr(UGU) and tRNAAsn(GUU) variants is sufficient to inhibit PARIS anti-viral defence. Finally, we propose a model for tRNA recognition by the AriB dimer and provide molecular details of its nuclease activity and specificity.

microbiology↗

bioRxiv2024

Molecular basis of foreign DNA recognition by BREX anti-phage immunity system

Anti-phage systems of the BREX (BacteRiophage EXclusion) superfamily rely on epigenetic DNA methylation to discriminate between the host and invading DNA, but their mechanism of protection remains enigmatic. We demonstrate that in Type I BREX systems, both defense and methylation are based on site-specific DNA recognition by the BrxX (PglX) methyltransferase and require the S-adenosyl methionine cofactor. We present a 2.2-[A] cryoEM structure of Escherichia coli BrxX bound to target dsDNA, which reveals the molecular details of DNA recognition by BREX and paves the way for rational engineering of BREX specificity. We show that BrxX alone does not support methylation, and BREX activity requires an assembly of a supramolecular BrxBCXZ immune complex. Finally, we present a cryoEM structure of BrxX bound to a phage-encoded inhibitor Ocr that sequesters an inactive dimeric form of BrxX. Together, these results allow us to propose a model of BREX-mediated DNA sensing and anti-phage defense.

molecular biology↗

bioRxiv2024

Viral proteins activate PARIS-mediated tRNA degradation and viral tRNAs rescue infection

Viruses compete with each other for limited cellular resources, and some viruses deliver defense mechanisms that protect the host from competing genetic parasites. PARIS is a defense system, often encoded in viral genomes, that is composed of a 53 kDa ABC ATPase (AriA) and a 35 kDa TOPRIM nuclease (AriB). Here we show that AriA and AriB assemble into a 425 kDa supramolecular immune complex. We use cryo-EM to determine the structure of this complex which explains how six molecules of AriA assemble into a propeller-shaped scaffold that coordinates three subunits of AriB. ATP-dependent detection of foreign proteins triggers the release of AriB, which assembles into a homodimeric nuclease that blocks infection by cleaving the host tRNALys. Phage T5 subverts PARIS immunity through expression of a tRNALys variant that prevents PARIS-mediated cleavage, and thereby restores viral infection. Collectively, these data explain how AriA functions as an ATP-dependent sensor that detects viral proteins and activates the AriB toxin. PARIS is one of an emerging set of immune systems that form macromolecular complexes for the recognition of foreign proteins, rather than foreign nucleic acids.

biochemistry↗

bioRxiv2023

Benchmarking DNA Isolation Methods for Marine Metagenomics Studies

Metagenomics is a powerful tool to study marine microbial communities. However, obtaining high-quality environmental DNA suitable for downstream sequencing applications is a challenging task. The quality and quantity of isolated DNA heavily depend on the choice of purification procedure and the type of sample. Selection of an appropriate DNA isolation method for a new type of material often entails a lengthy trial and error process. Further, each DNA purification approach introduces biases and thus affects the composition of the studied community. To account for these problems and biases, we systematically investigated efficiency of DNA purification from three types of samples (water, sea sediment, and digestive tract of a model invertebrate Magallana gigas) with eight commercially available microbial DNA isolation kits. For each kit-sample combination we measured the quantity of purified DNA, extent of DNA fragmentation, the presence of PCR-inhibiting contaminants, admixture of eukaryotic DNA, alpha-diversity, and reproducibility of the resulting community composition based on 16S rRNA amplicons sequencing. Additionally, we determined a "kitome", e.g., a set of contaminating taxa inherent for each type of purification kit used. The resulting matrix of evaluated parameters allows one to select the best DNA purification procedure for a given type of sample.

bioinformatics↗

bioRxiv2023

Phage T3 overcomes the BREX defence through SAM cleavage and inhibition of SAM synthesis

Bacteriophage T3 encodes a SAMase that through cleavage of S-adenosyl-methionine (SAM) circumvents the SAM-dependent Type I Restriction-Modification defence of the host bacterium Escherichia coli. Here, we show that the SAMase also allows T3 to evade BREX defence. SAM degradation weakly affects BREX methylation of host DNA, but completely inhibits the defensive function of BREX, suggesting that SAM is required as a co-factor for BREX-mediated exclusion of phage DNA. The anti-BREX activity of the T3 SAMase is mediated by two independent mechanisms: enzymatic degradation of SAM and downregulation of SAM synthesis through direct inhibition of the host SAM synthase MetK. We determined a 2.8 [A] cryo-EM structure of the eight-subunit T3 SAMase-MetK complex. Structure guided mutagenesis of the SAMase-MetK interface revealed that the interaction with MetK stabilizes the T3 SAMase in vivo, thus further stimulating its anti-BREX activity. This work provides insights in the versatility and intricacy of bacteriophage counter-defence mechanisms and highlights the role of SAM as an important co-factor of diverse phage-defence systems.

microbiology↗