Proteome Reallocation Reveals Dynamics and Mechanisms of Phage Infection and Defense in Staphylococcus aureus

The coexistence of bacteria and phages is marked by dynamic interactions that determine infection outcomes. However, the mechanisms by which the host allocates its resources to cope with phage infection remain largely unknown. In this study, using longitudinal proteomics, we elucidated these interactions for the temperate staphylococcal phage ϕ NM1 and strains of Staphylococcus aureus either harboring its cognate prophage or lacking it. We demonstrated that infection of non-lysogenic S. aureus with ϕ NM1 induces a dramatic shutdown of host translation, reducing proteome allocation by over 20%. Quantitative analysis of the economics of ϕ NM1 infection revealed that the diversion of these cellular resources toward phage replication imposes a significant metabolic burden, thereby impairing cell growth. In contrast, lysogenic cells cope with phage infection and prevent culture collapse through a coordinated response of prophage-encoded defenses, host-encoded stress effectors, and reprogrammed cellular metabolism, thereby avoiding translation shutdown. Through coinfection with the wildtype phage and an engineered phage-like particle carrying a CRISPR-Cas phagemid, we revealed that synthetic DNA cargos evade host defenses and hijack the transcriptional machinery, altering infection outcomes. Without immunity, coinfection could collapse the non-lysogens more quickly than the native phage by overexpressing the cargo proteins, suppressing carbohydrate metabolism, and accelerating structural phage protein production through increased phage genome replication. Together, these findings provide a systems-level understanding of phage infection in S. aureus , uncovering the mechanisms for host takeover and prophage-mediated defense, with implications for next-generation phage therapy.