Discovery of the mechanism of the antiviral immune system PARIS reveals new function of viral tRNA

Bacteriophages, viruses that infect bacterial cells, are natural predators and enemies of bacteria. When a phage infects a cell, it is lysed and dozens or even hundreds of new virus particles – phage progeny – are released. The ongoing arms race between bacteria and phages has led to the evolution of a variety of bacterial innate and adaptive immune systems and anti-immune systems in phages.

At the same time, phages can sometimes act on the host side and encode immune systems that help them compete with other phages. One of these immune systems is PARIS (Phage Anti-Restriction-Induced System).

PARIS consists of two components – the AriA sensor and the AriB toxin/effector. The system is activated by the Ocr (Overcoming classical restriction) protein, which mimics the structure of DNA and, by binding to DNA-recognizing immune proteins, helps the T7 phage avoid the action of restriction modification systems and the BREX (BacteRiophage EXclusion) defense system.

A team of scientists from Skoltech, the USA and France have deciphered the mechanism of the PARIS system and presented the results in a new article in the journal Nature.

“The PARIS system is so special from our point of view because it directly demonstrates the coevolution of bacteria and phages. It is a link between anti-immune proteins and bacterial immunity. Bacteria have more than 150 immune systems, which means that phages also have to adapt to them and encode different anti-immune proteins. PARIS is a kind of countermeasure of the bacteria that allows them to protect themselves against phages that have already evolved to inhibit other immune systems.

“In addition, PARIS can detect a viral infection by the presence of viral proteins in the cell rather than viral DNA, which is similar to the antigen-antibody principle characteristic of eukaryotic immunity. Many similar systems have been discovered in recent years (~AVAST, CapRel, Thoeris, etc.). At the same time, the appearance of viral proteins in the cell is often a sign that the infection has already progressed far enough and systems specifically designed to degrade phage DNA have not done their job.

“Under these conditions, programmed cell death is an effective defense strategy that stops the development of a viral infection at the population level. We managed to decipher the mechanism of such a 'suicide program' for the PARIS system,” comments Artem Isaev, corresponding author of the study, assistant professor and head of the Laboratory of Metagenomic Analysis at the Skoltech Bio Center.

The researchers showed that AriA and AriB are assembled into a supramolecular immune complex and deciphered its structure using cryo-electron microscopy. The AriA sensor is arranged in a propeller-shaped frame that coordinates the three AriB subunits and keeps the toxin inactive.

When a trigger occurs in the cell – that is, an activator protein such as Ocr – the AriA sensor binds directly to it, resulting in the release of the active dimer of the toxin effector AriB. The authors showed that AriB is an RNase that cleaves lysine-tRNA in a cell, leading to translational inhibition and cell death.

“The 'arms race' at PARIS can be observed in its purest form: the immune system of bacteria causes the formation of anti-immune proteins in viruses. In response, the PARIS system is formed in bacteria. Some phages have even gone further: the bacteriophage T5 has learned to inhibit the PARIS defense system.

“Like a crime thriller, it suddenly turned out that the T5 phages in Moscow and in Paris were slightly different phages: the Moscow phage lost a fragment of the genome and several tRNA genes, which made it sensitive to the protection of PARIS.

“We showed that the phage requires viral lysine tRNA for successful infection of PARIS cells, as it is not cleaved by the AriB protein. Thus, the T5 phage protects the cell from the toxic effects of the PARIS system, only to produce its own progeny,” added one of the first co-authors of the paper, Svetlana Belukhina, a doctoral student in the Skoltech Life Sciences program.

“Such a consistent evolutionary struggle, the emergence of new pathways of protection and anti-protection, immunity and its inhibition, still allows microbial populations, bacteria and phages to coexist and evolve together. This is a great example that evolution cannot be stopped. In addition, our work has provided a new answer to a long-known puzzle: why do viruses, in principle, encode their own tRNAs?

“It turns out that in some cases, tRNA molecules help the virus cope with the bacterial immune system. Presumably, tRNAs play a similar role in some viruses that infect humans,” added Mikhail Skutel, study co-author and doctoral student in the Skoltech Life Sciences program.

The authors emphasized that their work was made possible by open data sharing and equal contribution from multiple laboratories.

“Modern biology is impossible without cooperation and interdisciplinary expertise. In our study, we had to use methods of structural biology, bioinformatics and microbiology and work with a radioactive label, which often cannot be combined in the same laboratory. Without international cooperation and strong competition, which greatly stimulated our work, the preparation of this publication would hardly have been possible,” Isaev added.