The interferon-inducible protein Z-DNA binding protein 1 (ZBP1, also known as DNA-dependent activator of IFN-regulatory factors (DAI) and DLM-1) was identified as a dsDNA sensor, which instigates innate immune responses. However, this classification has been disputed and whether ZBP1 functions as a pathogen sensor during an infection has remained unknown. Herein, we demonstrated ZBP1-mediated sensing of the influenza A virus (IAV) proteins NP and PB1, triggering cell death and inflammatory responses via the RIPK1–RIPK3–Caspase-8 axis. ZBP1 regulates NLRP3 inflammasome activation as well as induction of apoptosis, necroptosis and pyroptosis in IAV-infected cells. Importantly, ZBP1 deficiency protected mice from mortality during IAV infection owing to reduced inflammatory responses and epithelial damage. Overall, these findings indicate that ZBP1 is an innate immune sensor of IAV and highlight its importance in the pathogenesis of IAV infection.

SUMMARY The inflammasome is an intracellular signaling complex, which on recognition of pathogens and physiological aberration, drives activation of caspase-1, pyroptosis, and the release of the pro-inflammatory cytokines IL-1β and IL-18. Bacterial ligands must secure entry into the cytoplasm to activate inflammasomes, however, the mechanism by which concealed ligands are liberated in the cytoplasm have remained unclear. Here, we showed that the interferon-inducible protein IRGB10 is essential for activation of the DNA-sensing AIM2 inflammasome by Francisella novicida, and contributed to the activation of the LPS-sensing caspase-11 and NLRP3 inflammasome by Gram-negative bacteria. IRGB10 directly targeted cytoplasmic bacteria through a mechanism requiring guanylate-binding proteins. Localization of IRGB10 to the bacterial cell membrane compromised bacterial structural integrity and mediated cytosolic release of ligands for recognition by inflammasome sensors. Overall, our results reveal IRGB10 as part of a conserved signaling hub at the interface between cell-autonomous immunity and innate immune sensing pathways.

The inflammasome is a complex of proteins that through the activity of caspase-1 and the downstream substrates gasdermin D, IL-1β, and IL-18 execute an inflammatory form of cell death termed pyroptosis. Activation of this complex often involves the adaptor protein ASC and upstream sensors including NLRP1, NLRP3, NLRC4, AIM2, and pyrin, which are activated by different stimuli including infectious agents and changes in cell homeostasis. Here we discuss new regulatory mechanisms that have been identified for the canonical inflammasomes, the most recently identified NLRP9b inflammasome, and the new gasdermin family of proteins that mediate pyroptosis and other forms of regulated cell death.

The cellular stress response has a vital role in regulating homeostasis by modulating cell survival and death. Stress granules are cytoplasmic compartments that enable cells to survive various stressors. Defects in the assembly and disassembly of stress granules are linked to neurodegenerative diseases, aberrant antiviral responses and cancer 1-5. Inflammasomes are multiprotein heteromeric complexes that sense molecular patterns that are associated with damage or intracellular pathogens, and assemble into cytosolic compartments known as ASC specks to

Programmed cell death plays crucial roles in organismal development and host defense. Recent studies have highlighted mechanistic overlaps and extensive, multifaceted crosstalk between pyroptosis, apoptosis, and necroptosis, three programmed cell death pathways traditionally considered autonomous. The growing body of evidence, in conjunction with the identification of molecules controlling the concomitant activation of all three pathways by pathological triggers, has led to the development of the concept of PANoptosis. During PANoptosis, inflammatory cell death occurs through the collective activation of pyroptosis, apoptosis, and necroptosis, which can circumvent pathogen-mediated inhibition of individual death pathways. Many of the molecular details of this emerging pathway are unclear. Here, we describe the activation of PANoptosis by bacterial and viral triggers and report protein interactions that reveal the formation of a PANoptosome complex. Infection of macrophages with influenza A virus, vesicular stomatitis virus, Listeria monocytogenes, or Salmonella enterica serovar Typhimurium resulted in robust cell death and the hallmarks of PANoptosis activation. Combined deletion of the PANoptotic components caspase-1 (CASP1), CASP11, receptor-interacting serine/threonine-protein kinase 3 (RIPK3), and CASP8 largely protected macrophages from cell death induced by these pathogens, while deletion of individual components provided reduced or no protection. Further, molecules from the pyroptotic, apoptotic, and necroptotic cell death pathways interacted to form a single molecular complex that we have termed the PANoptosome. Overall, our study identifies pathogens capable of activating PANoptosis and the formation of a PANoptosome complex.

Inflammasomes are multi-component signaling complexes critical to the initiation of pyroptotic cell death in response to invading pathogens and cellular damage. A number of innate immune receptors have been reported to serve as inflammasome sensors. Activation of these sensors leads to the proteolytic activation of caspase-1, a proinflammatory caspase responsible for the cleavage of proinflammatory cytokines interleukin-1β and interleukin-18 and the effector of pyroptotic cell death, gasdermin D. Though crucial to the innate immune response to infection, dysregulation of inflammasome activation can lead to the development of inflammatory diseases, neurodegeneration, and cancer. Therefore, clinical interest in the modulation of inflammasome activation is swiftly growing. As such, it is imperative to develop a mechanistic understanding of the regulation of these complexes. In this review, we divide the regulation of inflammasome activation into three parts. We discuss the transcriptional regulation of inflammasome components and related proteins, the post-translational mechanisms of inflammasome activation, and advances in the understanding of the structural basis of inflammasome activation.

Inflammasome activation is critical for host defenses against various microbial infections. Activation of the NLRC4 inflammasome requires detection of flagellin or type III secretion system (T3SS) components by NLR family apoptosis inhibitory proteins (NAIPs); yet how this pathway is regulated is unknown. Here, we found that interferon regulatory factor 8 (IRF8) is required for optimal activation of the NLRC4 inflammasome in bone-marrow-derived macrophages infected with Salmonella Typhimurium, Burkholderia thailandensis, or Pseudomonas aeruginosa but is dispensable for activation of the canonical and non-canonical NLRP3, AIM2, and Pyrin inflammasomes. IRF8 governs the transcription of Naips to allow detection of flagellin or T3SS proteins to mediate NLRC4 inflammasome activation. Furthermore, we found that IRF8 confers protection against bacterial infection in vivo, owing to its role in inflammasome-dependent cytokine production and pyroptosis. Altogether, our findings suggest that IRF8 is a critical regulator of NAIPs and NLRC4 inflammasome activation for defense against bacterial infection.

Guanylate-binding proteins (GBPs) are essential components of cell-autonomous immunity. In response to IFN signaling, GBPs are expressed in the cytoplasm of immune and nonimmune cells, where they unleash their antimicrobial activity toward intracellular bacteria, viruses, and parasites. Recent studies have revealed that GBPs are essential for mediating activation of the caspase-1 inflammasome in response to the gram-negative bacteria Salmonella enterica serovar Typhimurium, Francisella novicida, Chlamydia muridarum, Chlamydia trachomatis, Legionella pneumophila, Vibrio cholerae, Enterobacter cloacae, and Citrobacter koseri During infection with vacuolar-restricted gram-negative bacteria, GBPs disrupt the vacuolar membrane to ensure liberation of LPS for cytoplasmic detection by caspase-11 and the noncanonical NLRP3 inflammasome. In response to certain cytosolic bacteria, GBPs liberate microbial DNA for activation of the DNA-sensing AIM2 inflammasome. GBPs also promote the recruitment of antimicrobial proteins, including NADPH oxidase subunits and autophagy-associated proteins to the Mycobacterium-containing vacuole to mediate intracellular bacterial killing. Here, we provide an overview on the emerging relationship between GBPs and activation of the inflammasome in innate immunity to microbial pathogens.