A Single Amino Acid Residue Defines the Difference in NLRP3 Inflammasome Activation between NEK7 and NEK6.

The NOD-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome is a critical component of the innate immune system that is activated by microbial infections and cellular stress signals. The molecular mechanism of NLRP3 inflammasome activation remains not fully understood. As an NLRP3-interacting partner, NEK7 has emerged as a critical mediator for NLRP3 inflammasome activation. In contrast to NEK7, NEK6, the closely related member of the NEK family, does not support NLRP3 inflammasome activation. In this study, we show that the mouse NEK7 catalytic domain, which shares high sequence identity with the counterpart of NEK6, mediates its interaction with NLRP3 and inflammasome activation in mouse macrophages. Within their catalytic domains, a single amino acid residue at a corresponding position (R121NEK7, Q132NEK6) differentiates their function in NLRP3 inflammasome activation. Surprisingly, substitution of the glutamine residue to an arginine residue at position 132 confers NEK6 the ability of NLRP3 binding and inflammasome activation in mouse macrophages. Furthermore, our results suggest a structural pocket surrounding the residue R121 of NEK7 that is essential for NLRP3 binding and inflammasome activation.

The NLRP3 Inflammasome: An Overview of Mechanisms of Activation and Regulation.

The NLRP3 inflammasome is a critical component of the innate immune system that mediates caspase-1 activation and the secretion of proinflammatory cytokines IL-1ß/IL-18 in response to microbial infection and cellular damage. However, the aberrant activation of the NLRP3 inflammasome has been linked with several inflammatory disorders, which include cryopyrin-associated periodic syndromes, Alzheimer's disease, diabetes, and atherosclerosis. The NLRP3 inflammasome is activated by diverse stimuli, and multiple molecular and cellular events, including ionic flux, mitochondrial dysfunction, and the production of reactive oxygen species, and lysosomal damage have been shown to trigger its activation. How NLRP3 responds to those signaling events and initiates the assembly of the NLRP3 inflammasome is not fully understood. In this review, we summarize our current understanding of the mechanisms of NLRP3 inflammasome activation by multiple signaling events, and its regulation by post-translational modifications and interacting partners of NLRP3.

Innate immune sensing of macromolecule homeostasis.

The innate immune system uses a distinct set of germline-encoded pattern recognition receptors to recognize molecular patterns initially thought to be unique to microbial invaders, named pathogen-associated molecular patterns. The concept was later further developed to include similar molecular patterns originating from host cells during tissue damage, known as damage-associated molecular patterns. However, recent advances in the mechanism of monogenic inflammatory diseases have highlighted a much more expansive repertoire of cellular functions that are monitored by innate immunity. Here, we summarize several examples in which an innate immune response is triggered when homeostasis of macromolecule in the cell is disrupted in non-infectious or sterile settings. These ever-growing sensing mechanisms expand the repertoire of innate immune recognition, positioning it not only as a key player in host defense but also as a gatekeeper of cellular homeostasis. Therapeutics inspired by these advances to restore cellular homeostasis and correct the immune system could have far-reaching implications.

STING trafficking as a new dimension of immune signaling.

The cGAS-STING pathway is an evolutionarily conserved immune signaling pathway critical for microbial defense. Unlike other innate immune pathways that largely rely on stationary cascades of signaling events, STING is highly mobile in the cell. STING is activated on the ER, but only signals after it arrives on the Golgi, and then it is quickly degraded by the lysosome. Each step of STING trafficking through the secretory pathway is regulated by host factors. Homeostatic STING trafficking via COPI-, COPII-, and clathrin-coated vesicles is important for maintaining baseline tissue and cellular immunity. Aberrant vesicular trafficking or lysosomal dysfunction produces an immune signal through STING, which often leads to tissue pathology in mice and humans. Many trafficking-mediated diseases of STING signaling appear to impact the central nervous system, leading to neurodegeneration. Therefore, STING trafficking introduces a new dimension of immune signaling that likely has broad implications in human disease.

Interruption of post-Golgi STING trafficking activates tonic interferon signaling.

Activation of the cGAS-STING pathway is traditionally considered a "trigger-release" mechanism where detection of microbial DNA or cyclic di-nucleotides sets off the type I interferon response. Whether this pathway can be activated without pathogenic ligand exposure is less well understood. Here we show that loss of Golgi-to-lysosome STING cofactors, but not ER-to-Golgi cofactors, selectively activates tonic interferon signalling. Impairment of post-Golgi trafficking extends STING Golgi-dwell time, resulting in elevated immune signalling and protection against infection. Mechanistically, trans-Golgi coiled coil protein GCC2 and several RAB GTPases act as key regulators of STING post-Golgi trafficking. Genomic deletion of these factors potently activates cGAS-STING signalling without instigating any pathogenic trigger for cGAS. Gcc2-/- mice develop STING-dependent serologic autoimmunity. Gcc2-deleted or Rab14-deleted cancer cells induce T-cell and IFN-dependent anti-tumour immunity and inhibit tumour growth in mice. In summary, we present a "basal flux" mechanism for tonic cGAS-STING signalling, regulated at the level of post-Golgi STING trafficking, which could be exploited for cancer immunotherapy.