Mechanisms of phage detection by bacterial innate immune proteins

Abstract

Bacteria are under constant threat from their viral predators, known as bacteriophages (or phages). As a result, bacteria have evolved diverse immune mechanisms to protect themselves from phage infection, such as restriction modification, CRISPR-Cas, and abortive infection (Abi) systems. Because Abi systems function through killing infected cells to protect the bacterial population, they must stay inactive prior to infection, but rapidly detect phages and promptly trigger an immune response. Although many novel Abi systems have been discovered in recent years, how they detect phage infection remains poorly understood. Here, I demonstrated that CapRel_SJ46, an anti-phage protein from E. coli, senses phage infection by directly binding to the newly synthesized major capsid proteins (MCPs) of certain phages. Binding to the MCPs releases autoinhibition of the CapRel_SJ46 toxin domain, enabling it to pyrophosphorylate tRNAs, which blocks translation to restrict viral infection. Detection of the MCPs is analogous to how eukaryotic innate immune systems detect foreign invaders through conserved pathogen-associated molecular patterns (PAMPs). In addition to the MCPs, I found that CapRel_SJ46 can directly bind to another unrelated and structurally different phage protein, called Gp54. Bas11 phages harbor two trigger proteins, and both are sensed by CapRel_SJ46 during infection, indicating that a bacterial immunity protein can sense more than one phage-encoded trigger. Additionally, I demonstrated that another CapRel homolog, CapRel_Ebc, senses the inhibition of a host cell division protein by the phage-encoded trigger, which is analogous to effector-triggered immunity in eukaryotes, where innate immune proteins sense virulence-associated activities of pathogens rather than directly sensing PAMPs. Lastly, I characterized another Abi system, named RAZR (ring-activated zinc-finger RNase), and showed that RAZR forms a ring-shaped supramolecular complex of over 1 MDa upon sensing a phage-encoded PAMP, leading to activation of its RNase activity to restrict phage infection. This finding highlights the importance of higher-order molecular assembly in bacterial innate immunity. Collectively, my thesis work has provided new insights into the molecular mechanisms by which bacterial innate immune systems detect phage infection.