Discovery of phage defense systems through component modularity networks

Phage defense systems in bacteria exhibit high degrees of modularity, with sensing, signal transmission, and effector enzymes frequently exchanged among phage defense gene clusters. In this study, we capitalize on this modularity to discover phage defense systems by searching for defense-associated modules in new gene contexts across Pseudomonas aeruginosa genomes. This approach reveals a large and interconnected network of modular components distributed across diverse gene clusters. From over 500 candidate defense systems, we select five for experimental testing and validate three: Dionysus, a TerB-encoding system that interferes with early phage infection (EPI) vesicle formation by jumbo phages; Ophion, a radical S-adenosyl-L-methionine (SAM)-containing system that blocks progression from the EPI vesicle to the nucleus stage during jumbo phage infection; and Hercules, a tightly regulated restriction-modification (RM)-like system. Collectively, we demonstrate that leveraging the modular architecture of phage defense systems is an effective approach to their discovery.