Project description:In the ongoing arms race with phages, bacteria have evolved diverse defense systems, such as CRISPR-Cas and restriction-modification systems. The DNA double-strand break (DSB) repair system represents a core mechanism for maintaining genomic integrity and is vital for cell survival. However, it remains unknown whether and how these repair systems contribute to phage resistance. This study systematically investigates the role of the non-homologous end joining (NHEJ) during phage infection in Mycobacterium smegmatis. We found that NHEJ deficiency compromises host resistance to phage SWU1, as evidenced by increased plaque counts and reduced bacterial survival. Mechanistically, phages exploit host NHEJ for genomic repair; however, the error-prone nature of NHEJ leads to imperfect repair at phage cos sites, thereby blocking replication. The host modulates the balance between NHEJ and homologous recombination (HR) to control repair fidelity: NHEJ loss shifts the balance towards high-fidelity HR, which in turn promotes phage survival. Furthermore, NHEJ deficiency exacerbates infection-induced oxidative stress, leading to a compromise in bacterial viability. Our findings reveal the multifaceted functions of NHEJ in mycobacterium-phage interactions and provide new insights into how DNA repair systems shape antiphage defense and coevolution.
2026-01-01 | GSE315142 | GEO
Project description:Genetic architecture facilitates then constrains adaptation in a host-parasite coevolutionary arms race
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.
Project description:Bacteria are constantly threatened by their viral predators (phages), which has resulted in the development of defense systems for bacterial survival. One family of defense systems found widely across bacteria are OLD (for overcoming lysogeny defect) family nucleases. Despite recent discoveries regarding Class 2 and 4 OLD family nucleases and how phages overcome them, Class 1 OLD family nucleases warrant further study as there has only been one anti-phage Class 1 OLD family nuclease described to date. Here, we identify the Vibrio cholerae-encoded Class 1 OLD family nuclease Vc OLD and describe its disruption of genome replication of the lytic vibriophage ICP1. Furthermore, we examine its in vitro activity, identifying Vc OLD as a DNA nickase. Finally, we identify the first direct inhibitor of a Class 1 OLD family nuclease, the ICP1-encoded Oad1. Our research further illuminates Class 1 OLD family nucleases’ role in phage defense and explores the dynamic arms race between V. cholerae and its predatory phage ICP1.
Project description:Bacteria and their viruses (bacteriophages or phages) are in a dynamic arms race that balances predation and resistance, each deploying various strategies, including protein post-translational modifications (PTMs), to achieve dominance. To better understand the role PTMs play in phage infection, we infected Cellulophaga baltica bacteria with three previously characterized phages that represent diverse genomes and infection efficiencies (phi18:1, phi18:4, and phi38:1) to identify proteome-wide and protein-specific trends of PTMs. Approximately double the number of methylated residues on proteins were detected in phage-infected cells (virocells) compared to uninfected cells, and significantly increased frequencies of protein methylation were observed during the early stages of infection. This notable result led to a focus on protein methylation. Phage proteins were detectably methylated in both virocells and free virions, neither of which has been previously reported. Host proteins with known importance to phage infection--including GTPase EF-Tu, chaperone DnaK, and gliding motility proteins--were frequently methylated and/or exhibited methylation patterns in virocells that contrasted those in uninfected cells. Collectively, our results expand on the growing interest of the important role PTMs play in phage infection by demonstrating the dynamic methylation of phage proteins as well as host proteins important to phage infection.
Project description:Six isolates of PT21/28 and six of PT32 were analysed by CGH using UBECarray3 microarrays (containing probes for E. coli K-12 str. MG1655 and O157:H7 str. EDL933 and Sakai) to define genotypic differences between phage types. gDNA from E.coli O157 str. Sakai was hybridised to all arrays to provide a universal control channel on all arrays.
Project description:Bacteria and bacteriophages are in a constant arms race to develop bacterial defense and phage counter-defense systems. The prevailing view so far has been that phage counter-defense systems target specific bacterial defense systems. Here, we uncover a mechanism by which the T7 bacteriophage broadly manipulates host anti-phage defenses using protein phosphorylation. We show that the T7 protein kinase (gene 0.7, or T7K), which was believed to be specific to a defined set host factors, is in fact a hyperpromiscuous dual-specifity kinase enacting a massive wave of phosphorylation on virtually all host and phage proteins during infection. The scale of phosphorylation vastly exceeds the number of previously known phosphorylation events in E. coli, has no sequence motif specificity, and results in a higher proteome-wide phosphorylation density than even in mammalian cells that have ~500 kinases. Stoichiometry analysis of phosphorylation sites revealed a striking bias of T7K activity towards nucleic acid-binding substrates, which we show is mediated by a DNA-binding domain within T7K. This specificity for high stoichiometry phosphorylation of DNA binding proteins enables deactivation of diverse DNA targeting anti-defense systems. We provide mechanistic insight into how T7K weakens the Eco9 bacterial defense system through specific phosphorylation events, with phosphomimetic mutations in key sites of the toxin protein RcaT deactivating its defense. Finally, screening a genetically diverse collection of E. coli strains showed that half of strains with evidence of defense against T7 are also sensitive to T7K demonstrating a high prevalence and significance of T7K-mediated host interactions in nature.