Phage N4 uses a SAR endolysin-holin system for host cell lysis

Abstract Bacteriophages (phages) cause active host cell lysis to terminate their infection and release progeny into the environment. However, some phages have systems to delay their lysis, and thereby increase progeny yield, in specific conditions: a phenomenon called lysis inhibition (LIN). Two dissimilar phages of Escherichia coli are known to exhibit LIN: T4 and N4. In T4, a multi-protein mechanism stalls lysis and maintains the LIN state in response to superinfection in a high phage population density. However, the lysis proteins responsible for T4 lysis and LIN are not conserved with phage N4. In this study, we characterize the phage N4 proteins involved in lysis by molecular and genetic means. We define the functions of the minimal gene set required for lysis through heterologous expression and complementation. Furthermore, by sequence comparison with a selected mutant library that does not induce LIN, we have identified genomic regions both within and outside the lysis cassette involved in N4 LIN. We propose a model where N4 lysis proteins that execute rapid N4 lysis can be regulated to induce LIN. Despite the lack of conservation with T4 components, our study suggests that direct modulation of lysis initiation may be common and provides a springboard for identifying similar mechanisms in other phages. Importance Phages are viruses that kill only bacteria. To do so, they use lysis proteins to disrupt the structural components of the bacterial envelope. There are commonalities between the phages that infect bacteria with similar structures, but there are also interesting differences we can exploit to learn more about specific phages or their bacterial hosts. In this study, we characterize the lysis proteins of the E. coli phage N4. Our results suggest that lysis regulation controls phage yield in N4 differently than in well-studied phage T4. Future studies on the relationship between lysis proteins and phage yield could provide handles to use this information for optimal large-scale phage production for clinical or industrial applications.