The pyroptosome: a supramolecular assembly of ASC dimers mediating inflammatory cell death via caspase-1 activation

Abstract

Pyroptosis is a caspase-1-dependent inflammatory form of cell death. The adapter protein ASC binds directly to caspase-1 and is critical for caspase-1 activation in response to a broad range of stimuli. To elucidate the mechanism of activation of caspase-1 by ASC and its exact role in macrophage pyroptosis, we performed time-lapse confocal bioimaging analysis on human THP-1 macrophages stably expressing an ASC-GFP fusion protein. We show that stimulation of these cells with several proinflammatory stimuli trigger the formation of a large supramolecular assembly of ASC, termed here pyroptosome. Only one distinct pyroptosome in each stimulated cell is formed, which rapidly recruits and activates caspase-1 resulting in pyroptosis and the release of the intracellular proinflammatory cytokines. The pyroptosome is largely composed of oligomerized ASC dimers. Dimerization of ASC is driven by subphysiological concentrations of potassium as in vitro incubation of purified recombinant ASC in the presence of subphysiological concentrations of potassium induces the assembly of a functional pyroptosome. Furthermore, stimulation of potassium efflux in THP-1 cells with potassium-depleting agents induces formation of the pyroptosome, while increasing potassium concentrations in the culture medium or pharmacological inhibition of this efflux inhibits its assembly. Our results establish that macrophage pyroptosis is mediated by a unique pyroptosome, distinct from the inflammasome.

Pyroptotic Neuronal Cell Death Mediated by the AIM2 Inflammasome

“…17,33 ASC staining is diffuse in unstimulated macrophages, but upon stimulation with poly(dA:dT), ASC coalesces into a single focus previously called the pyroptosome. 18 Although we observed oligomerization of ASC upon stimulation with poly(dA:dT) in neurons, ASC staining remained diffuse. Rather than facilitate pyroptosis, the ASC focus in macrophages may sequester caspase-1 into a single location, restricting its access to substrates that are effectors of pyroptosis.…”

“…These findings indicate assembly of the pyroptosome, a supramolecular complex of ASC oligomers that serves as a platform for caspase-1 activation in pyroptosis. 18 Pyroptosis is also characterized by the formation of discrete pores in the plasma membrane. 19 To investigate if AIM2-mediated neuronal cell death also shares this feature, we applied two membrane impermeable dyes of different sizes to neurons stimulated with poly(dA:dT).…”

dsDNA ASCs for caspase 8-mediated apoptosis

“…In particular, in macrophages it was though also noted that excessive caspase 1 activation results in a most rapid form of inflammatory cell death closely resembling necrosis and thus termed pyroptosis. [3][4][5] While the death-inducing signaling complex (DISC) and the apoptosome are the major activation platforms for apoptosis-related caspases, the inflammasome is generally accepted as the major activation mechanism of caspase 1. 4,5 The inflammasome is a multiprotein complex with often variable composition, generally consisting of a danger-, stressor pathogen-sensing component, an adaptor protein and the proform of caspase 1.…”

“…Activation of various Nod-like receptors (NLRs) or AIM2 (absent in melanoma-2) leads to the recruitment of ASC, which in turn binds and recruits caspase 1, eventually leading to IL-1b and IL-18 processing and associated inflammation. Excessive ASC-mediated caspase 1 activation appears to enhance inflammation by the rapid induction of pyroptosis, 3 in particular in pathogen-infected macrophages. As NLRs and AIM2 are often triggered by bacterial and viral products, for example, cytoplasmic dsDNA and bacterial toxins, pyroptosis may help to limit pathogen spreading and further alerts uninfected immune cells by the induction of inflammatory processes.…”

Another twist in the on and off affair between cell suicide and inflammation

“…This form of pro-inflammatory cell suicide has been dubbed 'pyroptosis'. 15,16 A paper in this edition of Cell Death and Differentiation advances the story even further by showing that the AIM2 and NLRP3 inflammasomes can trigger ASC to activate not only pyroptotic (IL-1b releasing) death pathways, but also an apoptotic pathway, and more surprisingly, caspase 8 is involved in both. 17 Sagulenko et al 17 looked at bone marrow-derived macrophages from normal and gene-deleted mice, and studied their responses to calf thymus or synthetic polydA:polydT DNA that had been electroporated across the plasma membrane.…”