Single phage proteins sequester TIR- and cGAS-generated signaling molecules

Area of Science:

• Microbiology
• Molecular Biology
• Immunology

Background:

• Prokaryotic anti-phage systems utilize TIR and cGAS enzymes to generate cyclic nucleotide signals that restrict phage replication.
• The mechanisms by which phages counteract these defense systems remain largely uncharacterized.

Approach:

• Investigated the anti-defense mechanisms of Thoeris anti-phage proteins Tad1 and Tad2.
• Utilized in vivo and in vitro assays to assess the binding affinities and inhibitory activities of Tad1 and Tad2 against CBASS cyclic oligonucleotides.
• Characterized the stoichiometry and binding site specificities of Tad1 and Tad2 for cyclic di-/trinucleotides (CDNs/CTNs) and glycocyclic ADPR (gcADPR).

Key Points:

• Tad1 and Tad2 exhibit anti-CBASS activity by sequestering cyclic oligonucleotides, including CDNs, CTNs, and gcADPR.
• Tad1 possesses six binding sites for CDNs/gcADPR and two high-affinity sites for CTNs, inhibiting CBASS systems.
• Tad2 sequesters various CDNs and gcADPR through distinct binding pockets, demonstrating dual inhibition.

Conclusions:

• Tad1 and Tad2 represent a novel class of phage-encoded inhibitors with broad specificity for cyclic nucleotides involved in TIR- and cGAS-based immunity.
• These proteins act as two-pronged inhibitors, effectively neutralizing prokaryotic anti-phage defenses.
• The findings establish a new paradigm for phage counter-defense strategies involving flexible sequestration of signaling molecules.