Finally, thank you to @mfwhite2.bsky.social for the wonderful summary of these stories! www.nature.com/articles/d41...

Congrats as well to @erinedoherty.bsky.social and @benadler.bsky.social et al. in the @doudna-lab.bsky.social lab for their excellent complementary work on Panoptes! www.nature.com/articles/s41...

Huge shoutout and thanks to my lab mates in the @aaronwhiteley.bsky.social lab and our collaborators in the @benmorehouse.bsky.social lab. This work would not have been possible without you all! 🫶🏻

Very excited to see our work appear alongside findings from @aaronwhiteley.bsky.social & @benmorehouse.bsky.social !

www.nature.com/articles/s41...

Thank you to everyone who helped make this story happen!

Obligatory meme to summarize:
“How I imagine phage facing CBASS and mCpol”

Our sister-story from our friends @aesully98.bsky.social , @benmorehouse.bsky.social and @aaronwhiteley.bsky.social is out today as well! Congratulations!!
www.nature.com/articles/s41...

Also—check out the complementary work led by @erinedoherty.bsky.social and @benadler.bsky.social in @doudna-lab.bsky.social lab that just went up on bioRxiv!

Huge shout-out to all of the authors from the @aaronwhiteley.bsky.social and @benmorehouse.bsky.social labs that were so instrumental to this project! A special thank you to our collaborators L. Aravind and Max Burroughs for their incredible bioinformatic analyses!

In total, we uncovered a novel mechanism of immune evasion sensing in bacteria. The Panoptes system is part of a “layered” immune system that flips the liability of a nucleotide-derived second messenger in immune signaling against the phage, adding an interesting dimension to phage-bacteria warfare.

Finally, we wanted to explore the relationship between CBASS and Panoptes, given the importance of Acb2 in both nucleotide-based signaling systems. We found that 53% of all Panoptes systems co-occur with CBASS, suggesting that Panoptes systems are predominantly guardians of CBASS in bacteria.

Consistent with this, we found that OptE robustly inhibited bacterial growth unless an OptS-derived nucleotide was present and that OptS constitutively synthesizes 2′,3′-c-di-AMP in vivo in the absence of phage infection.

These data led us to hypothesize that OptS-synthesized nucleotides were actually inhibiting, rather than activating, the OptE effector. Upon phage infection, however, Acb2 sequesters the OptS product away from OptE, unleashing its growth inhibition effect.

These findings left us puzzled: How does the Acb2 nucleotide sponge protein activate the Panoptes system? Based on its ability to bind diverse nucleotide products, we proposed that Acb2 also binds the product of OptS (2′,3′-c-di-AMP), which is exactly what we found.

Acb2 was recently discovered by the Bondy-Denomy and Chen Labs to be an anti-defense protein that acts as a nucleotide “sponge,” sequestering CBASS-derived cyclic oligonucleotide signaling molecules.

Intriguingly, the escaper phages we identified encoded loss-of-function mutations exclusively in the anti-CBASS 2 (acb2) gene. Consistent with this, we found that acb2 was necessary and sufficient to activate Panoptes defense.

We now knew that nucleotide signaling was a key component of the Panoptes system, but we were left wondering: how does the phage activate defense? To answer this question, we turned to phages that were able to escape Panoptes.

We tested the activity of OptS by incubating it with a range of NTPs and found that it used ATP as a substrate to specifically produce 2′,3′-c-di-AMP, an isomer of c-di-AMP. We used a series of phosphodiesterases to show that the linkage included a 3′–5′ and 2′–5′ bond.