Caspase-4 binds to LPS membranes with positive curvature for non-canonical inflammasome activation

The non-canonical inflammasome, comprising caspase-4 in humans, initiates pyroptosis upon sensing cytosolic lipopolysaccharide (LPS) from Gram-negative bacteria. Caspase-4 activation also depends on several guanylate-binding proteins (GBPs), which associate with the surface of cytosolic bacteria. Here, we investigated how caspase-4 accesses its cognate ligand, the hydrophobic lipid A moiety of LPS, and the role of GBPs in this process. GBP1 was essential for caspase-4 activation during infection. Mechanistically, GBP1 deformed the LPS-containing outer membrane of cytosolic bacteria, acting as a GTP-dependent mechanoenzyme. In vitro, GBP1 fragmented LPS micelles and promoted caspase-4/LPS complex formation, thereby enhancing LPS-induced caspase-4 activation. Fragmented LPS micelles presented additional micelle tips that served as binding and activation sites for caspase-4, indicating that caspase-4 engages LPS membranes with defined geometry rather than individual LPS molecules. Thus, GBP-mediated deformation of the LPS-rich outer bacterial membrane generates regions of positive curvature that expose lipid A, enabling caspase-4 binding, oligomerization, and activation.