Dual phage sensing activates an RNA-targeting reverse transcriptase for anti-phage defense

Defense-associated reverse transcriptases (DRTs) play a crucial role in antiphage responses, but the mechanisms behind their activation and activation of unconventional DNA synthesis for antiviral effector activation are not entirely clear. Researchers have now identified DRT4 as a dual-sensor immune system that detects two different phage-derived signals.

Through cryo-EM analysis, they observed that DRT4 forms a complex structure called a dimer-of-trimers hexamer, with each protomer being responsible for protein-primed, template-independent synthesis of a unique ssDNA product. The DRT4 polymerase shows a strong preference for purine nucleotides, which is due to specific base interactions within its active site.

Genes involved in phage escape demonstrate that DRT4 activation is triggered by two independent factors: phage ribonucleotide reductase NrdAB, which raises intracellular levels of dNTPs, and the SSB-like protein 0087, which binds directly to the ssDNA produced by DRT4. Once activated, DRT4 is capable of degrading RNA using its dNTP-dependent RNase activity.

This process links phage sensing to RNA cleavage and abortive infection. In summary, the findings reveal that DRT4-generated ssDNA serves as a signaling intermediate that connects phage-induced metabolic changes with direct recognition of viral replication proteins, demonstrating how protein-primed DNA synthesis can be adapted for infection sensing and antiviral effector activation.