Costs and benefits of diversity-generating immune mechanisms

Abstract

Organisms across the tree of life have evolved diversity-generating immune mechanisms (DGMs) to counteract selective pressures imposed by their parasites. Increased host diversity has a major impact on parasite epidemics as well as host evolution. Being virtually ubiquitous, bacteria and their predators, bacteriophage (phage), are essential to every ecological niche on earth and key players in industrial and healthcare applications. Bacterial DGMs include CRISPRCas and Restriction-Modification (RM) shufflons. Type I RM methylates self-DNA and cleaves unmethylated invasive DNA, however phage can escape from this response by becoming methylated themselves. Shufflons recombine genes coding for the RM specificity subunit, creating population-level diversity in recognition sequences; this is thought to limit phage escape. We investigate the Mycoplasma pulmonis Mpu shufflon, which has the capacity to generate 30 different specificity subunits, of which we predict at least 12 to be functional. We create a model system by adapting the Mpu shufflon for expression in Pseudomonas aeruginosa PA14. Transforming a CRISPR-deficient PA14 host with RM, we uncover large autoimmune costs when inducing a novel RM system with only limited benefits of low-level phage resistance. When expressed together, CRISPR-Cas and RM provide PA14 with complete resistance against most Pseudomonas phages tested and partial resistance against lipopolysaccharide-specific phage LMA2. Surprisingly, the RM restriction subunit is not an essential component for this effect; the mechanistic basis of this synergistic interaction between DNA methylation and CRISPR-Cas systems requires further investigation. The lack of detectable spacer acquisition, required for CRISPR-Cas to effectively target the infecting phage, suggests these effects are likely due to altered host gene expression that in turn impacts the ability of phage to infect. Future studies need to address questions about the molecular basis of resistance in this model system.