Inflammasomes induce cell death in response to infection, chemical entities and cell damage. Human genetics and animal models have identified the NLRP3 inflammasome as a gatekeeper of caspase-1-dependent pyroptosis. NLRP3 activation induces polymerization of ASC into a single, micron-scale perinuclear punctum, where caspase-1 is activated. These puncta have yet to be imaged at sufficient resolution to resolve their ultrastructure. Here, we apply correlative cryo-light microscopy and cryo-electron tomography to visualize ASC/caspase-1 puncta and mitochondrial membrane remodelling within NLRP3-activated cells. The puncta are composed primarily of branched ASC filaments. The N-terminal pyrin domain forms an 8-10 nm-diameter tubular core, which is decorated by the flexibly linked C-terminal caspase recruitment domain. The variable filament density allows ribosomes and trans-Golgi-like vesicles to permeate the network. We propose this provides structural integrity while allowing macromolecules and vesicles to diffuse in or bind with the necessary density, timing and localization.
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