Prophage-encoded RexAB-type phage defense system in Pseudomonas putida

Temperate bacteriophages in their prophage form are abundant residents in bacterial genomes. As the fitness of a prophage depends on the welfare of its host, prophages often encode genes that enhance their host's fitness, thereby also contributing to their own survival. One such strategy for mutual survival is to protect the host from superinfection by other phages. We recently showed that four cryptic prophages contribute to the phage resistance of Pseudomonas putida, a metabolically versatile and industrially promising model bacterium. Here, we show that prophages 1, 2, and 3 exhibit anti-phage activity against specific phages in our CEPEST collection. Focusing on P1-mediated phage defense, we uncover that P1 genes PP_5643 and PP_5644 encode a two-component RexAB-type phage immunity system. The RexA_pp is a DNA-binding protein that shares structural similarity with phage lambda RexA. RexB_pp is a membrane protein that acts as a cell growth-suppressing effector when stimulated by RexA_pp overexpression, and probably by phage infection. The activation of RexB_pp results in membrane damage, as evidenced by increased membrane permeability to SYTOX Green and propidium iodide, as well as leakage of cytoplasmic enzymes. Our data suggest that RexB_pp causes growth cessation of infected bacteria and thus aborts the infection.