Zorya - a novel abortive infection system widespread in bacteria, with a broad anti-phage activity

The perpetual arms race between phages and bacteria is driving the innovative evolution of multiple, sophisticated defense strategies in prokaryotes that can be adapted to invaluable molecular biology tools such as restriction enzymes and CRISPR-Cas9. Zorya, a newly discovered defense system, is of particular interest as its mechanism of immunity contains homologs of components of a proton channel of bacterial flagella – domains that had never been identified before as being involved in defense.

In our studies, Zorya showed strong protection against a broad range of phages, including several morphological families of dsDNA and ssDNA phages. We first measured the culture dynamics of multiple phage infections using a robot-based system and showed that Zorya is an abortive infection system, i.e., infected bacterial cells commit suicide, preventing lysis of the entire cell population. Next, using several experimental setups with fluorescently labeled proteins, we microscopically examined the successful completion of each step of phage infection. We showed that although the phage successfully injects its DNA, no protein synthesis of the phage occurs. We then calculated the ratio of phage DNA to bacterial DNA during infection and showed that Zorya system abolishes phage DNA replication. To gain a deeper understanding of the mechanism of action of Zorya system, we used super-resolution microscopy to localize components of the system during phage infection. Also, we successfully co-purified and characterized several proteins that are bound to Zorya protein complex. The details of the defense mechanism of Zorya system will be further discussed.