Leveraging defense system modularity to discover anti-phage systems

Abstract Bacterial exposure to constant phage attack drives rapid diversification of anti-phage defense systems, often through the exchange of modular defensive domains. Here, we leverage this modularity signature to identify new defense systems by systematically searching for operons encoding defensive domains in non-canonical configurations. We identified 214,164 candidate defense operons in E. coli genomes, representing a ∼2.2-fold expansion over defense systems detected by DefenseFinder. Experimental testing of 9 candidates validated 6 with anti-phage activity. These include DarTG and ietAS system variants that have acquired helicase modules, and a Gabija system in which a MazF-like protein replaces GajA, implying novel anti-phage mechanisms. We also identified a new clade of Pycsar and show that it synergizes with type IV Thoeris to broaden phage protection. Our findings demonstrate that mining modular defensive domains provides a powerful strategy to predict and characterize new anti-phage systems, expanding the known repertoire of bacterial immunity. Teaser Modular domains of anti-phage defense systems are harnessed to predict novel defense systems Competing Interest Statement The authors have declared no competing interest.