Bacteriophage anti-defense genes that neutralize TIR and STING immune responses
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ABSTRACT:
PROVIDER: PRJEB58053 | ENA |
REPOSITORIES: ENA
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Project description:Programmed cell suicide of infected bacteria, known as abortive infection (Abi), serves as a central immune defense strategy to prevent the spread of bacteriophage viruses and other invasive genetic elements across a population. Many Abi systems utilize bespoke cyclic nucleotide immune messengers generated upon infection to rapidly mobilize cognate death effectors. Here, we identify a large family of bacteriophage nucleotidyltransferases (NTases) which synthesize competitor cyclic dinucleotide (CDN) ligands and inhibit NAD-depleting TIR effectors activated through a linked STING CDN sensor domain (TIR-STING). Through a functional screen of NTase-adjacent phage genes, we uncover candidate inhibitors of host TIR-STING suicide signaling. Among these, we demonstrate that a virus MazG-like nucleotide pyrophosphatase, Atd1, depletes the starvation alarmone (p)ppGpp, revealing a role for the alarmone-activated host toxin MazF as a key executioner of TIR-driven abortive infection. Phage NTases and counter-defenses like Atd1 preserve host viability to ensure virus propagation, and may be exploited as tools to modulate TIR and STING immune responses.
2023-03-27 | GSE222071 | GEO
Project description:Bacteriophage antidefense genes that neutralize TIR and STING immune responses
| PRJNA917585 | ENA
Project description:Bacteria and bacteriophages have been engaged in a relentless evolutionary arms race, driving a rapid evolution of bacterial defense mechanisms and leading to their scattered distribution across genomes. We hypothesized that the variability in defense systems presence in bacterial genomes leads to equally variable counter-defense repertoires in phage genomes. To test this, we analyzed the variable regions in Pseudomonas model phages of the Pbunavirus genus, uncovering five anti-defense genes inhibiting Zorya type I, RADAR, Hypnos, Druantia type I and III, and Thoeris type III. Remarkably, a typical Pbunavirus encodes up to five known anti-defense genes, some inhibiting four unrelated defense systems with distinct nucleic acid-targeting mechanisms. Structural searches revealed that these broad-acting inhibitors are encoded across diverse phage taxa infecting multiple bacterial hosts. The presence of both broad and specific inhibitors suggests that defense systems exert a strong selective pressure, and their utility presents opportunities to improve phage-based therapeutics.
2026-02-23 | PXD063439 | Pride
Project description:Capsule is a critical virulence factor that significantly contributes to phage resistance in Acinetobacter baumannii. To investigate the interplay between capsule-based defense and phage predation, we applied phage selection pressure to A. baumannii to generate isogenic phage-resistant mutants. Utilizing transcriptomic analysis, we subsequently characterized the global alterations in the biological regulatory network of a capsule-deficient, phage-resistant mutant in comparison to its parental strain. This approach allowed us to identify key transcriptional reprogramming events associated with the acquisition of phage resistance in the absence of a functional capsule.
2025-10-01 | GSE308794 | GEO
Project description:Retrons are prokaryotic genetic elements involved in anti-phage defense and consist of a non-coding RNA, a reverse transcriptase (RT), and various effector proteins. Retron-Eco7 (previously known as Retron-Ec78) from Escherichia coli encodes two effector proteins (a PtuA ATPase and a PtuB nuclease) and degrades host tRNATyr upon phage infection, thereby protecting host cells against invading phages. However, its defense mechanism remains elusive. Here, we report the cryo-electron microscopy structures of the Retron-Eco7 complex, comprising the RT, multicopy single-stranded DNA (msDNA), PtuA, and PtuB. The Retron-Eco7 structure reveals that the RT–msDNA complex associates with two PtuA–PtuB complexes, potentially inhibiting their nuclease activity and suppressing bacterial growth arrest prior to phage infection. Furthermore, we found that a phage-encoded D15 nuclease acts as a trigger for the Retron-Eco7 system, cleaving the msDNA bound to the complex and facilitating the dissociation of PtuA–PtuB from RT–msDNA. Our data indicate that msDNA cleavage by D15 is the initial step required for the specific cleavage of host tRNATyr by the PtuA–PtuB nuclease, which leads to abortive infection. Overall, this study provides mechanistic insights into the Retron-Eco7 system and highlights the diversity of prokaryotic anti-phage defense mechanisms.
2025-10-23 | GSE293394 | GEO
Project description:Bacteria encode diverse defense systems including restriction-modification and CRISPR-Cas that cleave nucleic acid to protect against phage infection. Bioinformatic analyses demonstrate many recently identified anti-phage defense operons are comprised of a nuclease and NTPase protein, suggesting additional nucleic acid targeting systems remain to be understood. Here we develop large-scale comparative cell biology and biochemical approaches to analyze 16 nuclease-NTPase systems and define molecular features that control anti-phage defense. Purification, biochemical characterization, and in vitro reconstitution of nucleic acid degradation demonstrates protein–protein complex formation is a shared feature of multi-gene nuclease-NTPase systems. We show that AbpAB, Hachiman, and PD-T4-8 system nucleases use highly degenerate recognition site preferences to enable broad nucleic acid degradation, and the Azaca system exhibits specific phage targeting through the recognition of modified phage genomic DNA. Our results uncover principles of anti-phage defense system function and highlight the mechanistic diversity of nuclease-NTPase systems in bacterial immunity.
2026-02-18 | GSE303276 | GEO
Project description:Antiviral STANDs (Avs) are bacterial anti-phage proteins that are considered as the evolutionary ancestors of immune pattern-recognition receptors of the NLR family. Following the recognition of a conserved phage protein, Avs proteins exhibit cellular toxicity and abort phage propagation by killing the infected cell. Type 2 Avs proteins (Avs2) were suggested to recognize the large terminase subunit of the phage by direct binding as a signature of phage infection based on co-expression assays. Here, we analyzed the binding partners of a type 2 Avs protein from Klebsiella pneumoniae (KpAvs2) expressed in Escherichia coli during SECphi18 phage infection and showed that rather than the large terminase subunit, KpAvs2 binds a small phage protein of unknown function during infection.
2024-12-04 | PXD048766 | Pride
Project description:Phage therapy has garnered significant attention due to the rise of life-threatening multidrug-resistant pathogenic bacteria and the growing awareness of the transfer of resistance genes between pathogens. In light of this, phage therapy applications are now being extended to target plant pathogenic bacteria, like Erwinia amylovora that causes fire blight in apple and pear orchards. Understanding the mechanisms of phage resistance development is crucial for enhancing the effectiveness of phage therapy. Despite the challenges of naturally developing a bacteriophage resistant mutant (BIM) of E. amylovora (without traditional mutagenesis methods), this study successfully created a BIM mutant against the podovirus Ea46-1-A1. The parent strain, E. amylovora D7, and the BIM mutant, B6-2 were compared at the transcriptomic level.
2025-12-09 | GSE291879 | GEO
Project description:Zorya is a recently identified and widely distributed bacterial immune system that protects bacteria from viral (phage) infections. Three Zorya subtypes have been discovered, each containing predicted membrane-embedded ZorAB complexes paired with soluble subunits that differ among Zorya subtypes, notably ZorC and ZorD in type I Zorya systems1,2. Here, we investigate the molecular basis of Zorya defense using cryo-electron microscopy, mutagenesis, fluorescence microscopy, proteomics, and functional studies. We present cryo-EM structures of ZorAB and show that it shares stoichiometry and features of other 5:2 inner membrane ion-driven rotary motors. The ZorA5B2 complex contains a dimeric ZorB peptidoglycan binding domain and a pentameric α-helical coiled-coil tail made of ZorA that projects approximately 70 nm into the cytoplasm. We also characterize the structure and function of the soluble Zorya components, ZorC and ZorD, finding that they harbour DNA binding and nuclease activity, respectively. Comprehensive functional and mutational analyses demonstrate that all Zorya components work in concert to protect bacterial cells against invading phages. We provide evidence that ZorAB operates as a proton-driven motor that becomes activated upon sensing of phage invasion. Subsequently, ZorAB transfers the phage invasion signal through the ZorA cytoplasmic tail to recruit and activate the soluble ZorC and ZorD effectors, which facilitate degradation of the phage DNA. In summary, our study elucidates the foundational mechanisms of Zorya function as an anti-phage defense system.
2025-01-24 | PXD047450 | Pride
Project description:Bacteriophages must overcome diverse bacterial immune systems, yet the molecular principles enabling such escape remain poorly understood. Here, we show that the phage homing endonuclease SegB facilitates immune evasion by promoting the segmental amplification of anti-defense loci. The antiphage defense Septu inhibits phage T6 replication by cleaving the variable loop of tRNATyr. We show that SegB enables immune evasion by amplifying a genomic segment that contains the full-length tRNATyr gene. This repeat expansion increases tRNATyr expression, allowing the phage to overcome Septu immunity. Remarkably, SegB also mediates in trans amplification of distinct anti-defense genes that counteract OLD and toxin-antitoxin ToxIN defense systems. Collectively, our findings demonstrate that SegB-mediated segmental amplification represents a versatile mechanism by which phages rapidly adapt to and circumvent diverse bacterial antiphage defenses.
2026-02-05 | GSE303005 | GEO