Investigating the mechanism of infection and therapeutic potential of Pseudomonas nucleus-forming phages

Project Summary Nucleus-forming bacteriophages (phages) are a recently discovered class of lytic phages that protect their genome by encapsulating it in a nucleus-like protein shell (phage nucleus). The goal of the proposed research is to determine whether the unique properties of nucleus-forming phages make them advantageous therapies for human Pseudomonas aeruginosa infections. ΦKZ and ΦPA3 are nucleus-forming phages that infect P. aeruginosa and demonstrate the ability to selectively localize proteins to their phage nucleus and exclude others, including DNA-targeting bacterial phage defense proteins. Preliminary data show that ΦKZ gp69 is a conserved protein in P. aeruginosa nucleus-forming phages and plays a key role in mediating selective protein trafficking into the phage nucleus. This research will determine the structure of gp69 in complex with other nuclear shell-associated proteins, the mechanism by which gp69 mediates protein transport into the phage nucleus, and other factors that facilitate transport into the phage nucleus. Furthermore, this research will determine the motifs that signal a protein to be imported into the phage nucleus, how those motifs differ between P. aeruginosa infecting phage, and the competitive advantages selective protein import imparts on nucleus-forming phages. Lastly, resistance to phages is known to select for costly evolutionary trade-offs in the host bacterium. This research will determine how pathogenicity and antibiotic sensitivity are altered in P. aeruginosa resistant to nucleus-forming phages. Together, the results of this study will provide crucial information for the development of nucleus-forming phage therapeutics.