Mechanistic insights into activation of bacterial Retron-Eco8 immunity by phage protein SSB

The Retron-Eco8 system, comprising a reverse transcriptase (RT), a non-coding RNA (ncRNA), and an OLD-family nuclease effector, protects bacteria from phage infection via abortive infection upon sensing a phage single-stranded DNA-binding protein (SSB). However, the molecular basis of this immunity remained unclear. Here, we report cryo-electron microscopy (cryo-EM) structures of Retron-Eco8 in inactive and activated states, revealing mechanisms of phage-triggered activation and effector function. Retron-Eco8 assembles into a tetrameric complex in which each protomer contains an RT, msrRNA-msdDNA duplex, and effector in an autoinhibited conformation. Upon phage infection, phage SSB binds msdDNA, relieving autoinhibition and activating the nuclease effector to degrade both phage and host DNA, triggering cell death to block phage propagation. Host SSB fails to activate the system, while DNA binding and oligomerization of phage SSB are essential for this activation, highlighting its specificity. These findings elucidate the molecular mechanism of Retron-Eco8-mediated immunity, facilitating retron-based biotechnological applications.