2'3'-cGAMP-induced membrane shearing promotes broad antiphage immunity.

Area of Science:

• Microbiology
• Molecular Biology
• Structural Biology

Background:

• The cyclic-oligonucleotide-based anti-phage signaling system (CBASS) is a key prokaryotic antiviral defense.
• Transmembrane (TM) proteins are common CBASS effectors, but their immune activation mechanisms are poorly understood.

Purpose of the Study:

• To elucidate the mechanism by which three transmembrane (3TM)-SMODS-associated fused to various effector domains (SAVED) effectors mediate membrane disruption in CBASS signaling.
• To understand how ligand binding triggers effector assembly and membrane permeabilization.

Main Methods:

• Cryo-electron microscopy (Cryo-EM) to determine the structure of the 3TM-SAVED effector.
• Biochemical assays to study effector assembly and function in vitro.
• Liposome-based membrane disruption assays.

Main Results:

• 3TM-SAVED effectors assemble stepwise from monomers to filaments upon binding 2'3'-cyclic GMP-AMP (cGAMP).
• Filament formation involves reorientation of TM helices and amphipathic hairpins, creating vertically offset arrays.
• These arrays drive vertical lipid shearing, forming pores that permeabilize membranes and induce cell death.

Conclusions:

• This study reveals the molecular mechanism of CBASS TM effectors, demonstrating how they sense cyclic nucleotides and disrupt membranes.
• Vertical membrane shearing is identified as a novel principle of membrane disruption applicable across life domains.
• Findings provide insights into the evolution of innate immunity pathways, including the mammalian cGAS-STING pathway.