scholarly journals The ubiquitylation of IL-1β limits its cleavage by caspase-1 and targets it for proteasomal degradation

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Swarna L. Vijayaraj ◽  
Rebecca Feltham ◽  
Maryam Rashidi ◽  
Daniel Frank ◽  
Zhengyang Liu ◽  
...  
Keyword(s):  
Electrostatic Interaction ◽  
Inflammatory Diseases ◽  
Salt Bridge ◽  
Proteasomal Degradation ◽  
Interleukin 1Β ◽  
Collateral Damage ◽  
Caspase 1 ◽  
Microbial Responses

AbstractInterleukin-1β (IL-1β) is activated by inflammasome-associated caspase-1 in rare autoinflammatory conditions and in a variety of other inflammatory diseases. Therefore, IL-1β activity must be fine-tuned to enable anti-microbial responses whilst limiting collateral damage. Here, we show that precursor IL-1β is rapidly turned over by the proteasome and this correlates with its decoration by K11-linked, K63-linked and K48-linked ubiquitin chains. The ubiquitylation of IL-1β is not just a degradation signal triggered by inflammasome priming and activating stimuli, but also limits IL-1β cleavage by caspase-1. IL-1β K133 is modified by ubiquitin and forms a salt bridge with IL-1β D129. Loss of IL-1β K133 ubiquitylation, or disruption of the K133:D129 electrostatic interaction, stabilizes IL-1β. Accordingly, Il1bK133R/K133R mice have increased levels of precursor IL-1β upon inflammasome priming and increased production of bioactive IL-1β, both in vitro and in response to LPS injection. These findings identify mechanisms that can limit IL-1β activity and safeguard against damaging inflammation.

scholarly journals The inflammasome-activated cytokine IL-1[beta] is targeted for ubiquitylation and proteasomal degradation to limit its inflammatory potential

2021 ◽  
Author(s):  
Swarna Vijayaraj ◽  
Rebecca Feltham ◽  
Maryam Rashidi ◽  
Daniel Frank ◽  
Zhengyang Liu ◽  
...  
Keyword(s):  
Interleukin 1 ◽  
Salt Bridge ◽  
Beta Activity ◽  
Collateral Damage ◽  
Interleukin 1 Beta ◽  
Wide Spread ◽  
Caspase 1 ◽  
Microbial Responses

Interleukin-1[beta] (IL-1[beta]) is activated by inflammasome-associated caspase-1 in rare autoinflammatory conditions and in wide-spread diseases. Therefore, IL-1[beta] activity must be fine-tuned to enable anti-microbial responses whilst limiting collateral damage. Here we report that precursor IL-1[beta] is rapidly turned over by the proteasome and this correlates with its decoration by K11-, K63- and K48-linked ubiquitin chains. The ubiquitylation of IL-1[beta] is not just a degradation signal triggered by inflammasome priming and activating stimuli, but also limits IL-1[beta] cleavage by caspase-1. We further demonstrate that IL-1[beta] K133 is modified by ubiquitin and forms a salt bridge with IL-1[beta] D129. Loss of IL-1[beta] K133 ubiquitylation, or disruption of the K133:D129 electrostatic interaction, stabilizes IL-1[beta]. Accordingly, IL-1[beta] K133R/K133R mice display increased precursor IL-1[beta] upon inflammasome priming and increased bioactive IL-1[beta], both in vitro and following LPS injection in vivo. These findings reveal new mechanisms for limiting IL-1[beta] activity and safeguarding against damaging inflammation.

Novel irreversible caspase-1 inhibitor attenuates the maturation of intracellular interleukin-1β

2007 ◽  
Vol 85 (1) ◽  
pp. 56-65 ◽  
Author(s):  
Xue-Qin Ma ◽  
Hua-Jie Zhang ◽  
Ya-Hui Zhang ◽  
Yi-Hua Chen ◽  
Fang Wu ◽  
...  
Keyword(s):  
Rheumatoid Arthritis ◽  
Proinflammatory Cytokines ◽  
Caspase 3 ◽  
Inflammatory Diseases ◽  
Interleukin 1Β ◽  
Interleukin 18 ◽  
Caspase 1

. Caspase-1, the most efficient enzyme in processing the proinflammatory cytokines interleukin 1β and interleukin 18 in humans, is associated with inflammatory diseases such as rheumatoid arthritis, osteoarthritis, and some neuronal diseases. We previously reported that isoquinoline-1,3,4-trione and its derivatives are novel caspase-3 inhibitors that could attenuate apoptosis in vitro and in vivo. Here we report a novel derivative of isoquinoline-1,3,4-trione that is highly potent in inhibiting caspase-1 activity in an irreversible and slow-binding manner, thus inhibiting cellular caspase-1 activity and the maturation of interleukin 1β in U-937 cells.

scholarly journals OLT1177, a β-sulfonyl nitrile compound, safe in humans, inhibits the NLRP3 inflammasome and reverses the metabolic cost of inflammation

2018 ◽  
Vol 115 (7) ◽  
pp. E1530-E1539 ◽  
Author(s):  
Carlo Marchetti ◽  
Benjamin Swartzwelter ◽  
Fabia Gamboni ◽  
Charles P. Neff ◽  
Katrin Richter ◽  
...  
Keyword(s):  
Human Blood ◽  
Nlrp3 Inflammasome ◽  
Inflammatory Diseases ◽  
Plasma Concentrations ◽  
Metabolic Cost ◽  
Inflammasome Activation ◽  
Potassium Efflux ◽  
Caspase 1 ◽  
Healthy Humans

Activation of the NLRP3 inflammasome induces maturation of IL-1β and IL-18, both validated targets for treating acute and chronic inflammatory diseases. Here, we demonstrate that OLT1177, an orally active β-sulfonyl nitrile molecule, inhibits activation of the NLRP3 inflammasome. In vitro, nanomolar concentrations of OLT1177 reduced IL-1β and IL-18 release following canonical and noncanonical NLRP3 inflammasome activation. The molecule showed no effect on the NLRC4 and AIM2 inflammasomes, suggesting specificity for NLRP3. In LPS-stimulated human blood-derived macrophages, OLT1177 decreased IL-1β levels by 60% and IL-18 by 70% at concentrations 100-fold lower in vitro than plasma concentrations safely reached in humans. OLT1177 also reduced IL-1β release and caspase-1 activity in freshly obtained human blood neutrophils. In monocytes isolated from patients with cryopyrin-associated periodic syndrome (CAPS), OLT1177 inhibited LPS-induced IL-1β release by 84% and 36%. Immunoprecipitation and FRET analysis demonstrated that OLT1177 prevented NLRP3-ASC, as well as NLRP3-caspase-1 interaction, thus inhibiting NLRP3 inflammasome oligomerization. In a cell-free assay, OLT1177 reduced ATPase activity of recombinant NLRP3, suggesting direct targeting of NLRP3. Mechanistically, OLT1177 did not affect potassium efflux, gene expression, or synthesis of the IL-1β precursor. Steady-state levels of phosphorylated NF-κB and IkB kinase were significantly lowered in spleen cells from OLT1177-treated mice. We observed reduced IL-1β content in tissue homogenates, limited oxidative stress, and increased muscle oxidative metabolism in OLT1177-treated mice challenged with LPS. Healthy humans receiving 1,000 mg of OLT1177 daily for 8 d exhibited neither adverse effects nor biochemical or hematological changes.

scholarly journals The E3 Ubiquitin Ligase TRIM65 Negatively Regulates Inflammasome Activation Through Promoting Ubiquitination of NLRP3

2021 ◽  
Vol 12 ◽  
Author(s):  
Tiantian Tang ◽  
Ping Li ◽  
Xinhui Zhou ◽  
Rui Wang ◽  
Xiuqin Fan ◽  
...  
Keyword(s):  
Nlrp3 Inflammasome ◽  
Inflammatory Diseases ◽  
Critical Role ◽  
Neutrophil Migration ◽  
Inflammasome Activation ◽  
Regulation Mechanism ◽  
Caspase 1 ◽  
Nlrp3 Inflammasome Activation

The dysregulation of NLRP3 inflammasome plays a critical role in pathogenesis of various human inflammatory diseases, thus NLRP3 inflammasome activation must be tightly controlled at multiple levels. However, the underlying mechanism regulating NLRP3 inflammasome activation remains unclear. Herein, the effects of Tripartite motif-containing protein 65 (TRIM65) on NLRP3 inflammasome activation and the underlying molecular mechanism were investigated in vitro and in vivo. Inhibition or deletion of Trim65 could significantly strengthen agonist induced NLRP3 inflammasome activation in THP-1 cells and BMDMs, indicated by increased caspase-1 activation and interleukin-1β secretion. However, TRIM65 had no effect on poly (dA: dT)-induced AIM2 inflammasome activation or flagellin-induced IPAF inflammasome activation. Mechanistically, immunoprecipitation assays demonstrated that TRIM65 binds to NACHT domain of NLRP3, promotes lys48- and lys63- linked ubiquitination of NLRP3 and restrains the NEK7-NLRP3 interaction, thereby inhibiting NLRP3 inflammasome assembly, caspase-1 activation, and IL-1β secretion. In vivo, three models of inflammatory diseases were used to confirm the suppression role of TRIM65 in NLRP3 inflammasome activation. TRIM65-deficient mice had a higher production of IL-1β induced by lipopolysaccharide in sera, and more IL-1β secretion and neutrophil migration in the ascites, and more severity of joint swelling and associated IL-1β production induced by monosodium urate, suggesting that TRIM65 deficiency was susceptible to inflammation. Therefore, the data elucidate a TRIM65-dependent negative regulation mechanism of NLRP3 inflammasome activation and provide potential therapeutic strategies for the treatment of NLRP3 inflammasome-related diseases.

scholarly journals Neutrophils Facilitate Prolonged Inflammasome Response in the DAMP-Rich Inflammatory Milieu

2021 ◽  
Vol 12 ◽  
Author(s):  
Seunghwan Son ◽  
Sung-Hyun Yoon ◽  
Byeong Jun Chae ◽  
Inhwa Hwang ◽  
Do-Wan Shim ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Inflammatory Diseases ◽  
Macrophage Polarization ◽  
Interleukin 1Β ◽  
Inflammasome Activation ◽  
M2 Macrophage ◽  
Mitochondrial Depolarization ◽  
Activated Neutrophils

Aberrant inflammasome activation contributes to various chronic inflammatory diseases; however, pyroptosis of inflammasome-active cells promptly terminates local inflammasome response. Molecular mechanisms underlying prolonged inflammasome signaling thus require further elucidation. Here, we report that neutrophil-specific resistance to pyroptosis and NLRP3 desensitization can facilitate sustained inflammasome response and interleukin-1β secretion. Unlike macrophages, inflammasome-activated neutrophils did not undergo pyroptosis, indicated by using in vitro cell-based assay and in vivo mouse model. Intriguingly, danger-associated molecular patterns (DAMP)-rich milieu in the inflammatory region significantly abrogated NLRP3-activating potential of macrophages, but not of neutrophils. This macrophage-specific NLRP3 desensitization was associated with DAMP-induced mitochondrial depolarization that was not observed in neutrophils due to a lack of SARM1 expression. Indeed, valinomycin-induced compulsory mitochondrial depolarization in neutrophils restored inflammasome-dependent cell death and ATP-induced NLRP3 desensitization in neutrophils. Alongside prolonged inflammasome-activating potential, neutrophils predominantly secreted interleukin-1β rather than other proinflammatory cytokines upon NLRP3 stimulation. Furthermore, inflammasome-activated neutrophils did not trigger efferocytosis-mediated M2 macrophage polarization essential for the initiation of inflammation resolution. Taken together, our results indicate that neutrophils can prolong inflammasome response via mitochondria-dependent resistance to NLRP3 desensitization and function as major interleukin-1β-secreting cells in DAMP-rich inflammatory region.

scholarly journals Synthetic 4-Hydroxy Auxarconjugatin B, a Novel Autophagy Inducer, Attenuates Gouty Inflammation by Inhibiting the NLRP3 Inflammasome

Cells ◽  
2020 ◽  
Vol 9 (2) ◽  
pp. 279 ◽  
Author(s):  
Chih-Yu Hsieh ◽  
Lan-Hui Li ◽  
Yulin Lam ◽  
Zhanxiong Fang ◽  
Chin Heng Gan ◽  
...  
Keyword(s):  
Uric Acid ◽  
Nlrp3 Inflammasome ◽  
Inflammatory Diseases ◽  
Gouty Arthritis ◽  
Mechanistic Study ◽  
Caspase 1 ◽  
Autophagy Induction ◽  
Active Caspase

Gouty arthritis results from the generation of uric acid crystals within the joints. These uric acid crystals activate the NACHT, LRR and PYD domains-containing protein 3 (NLRP3) inflammasome, which is involved in chronic inflammatory diseases, including gouty arthritis. This study identified the polyenylpyrrole derivative 4-hydroxy auxarconjugatin B (4-HAB), a novel autophagy inducer, which attenuated uric acid crystals-mediated activation of the NLRP3 inflammasome in vitro and in vivo. 4-HAB dose-dependently reduced the release of interleukin (IL)-1β, IL-18, active caspase-1 and apoptosis-associated speck-like protein (ASC) in uric acid crystals-activated macrophages. In a mechanistic study, 4-HAB was shown to inhibit uric acid crystals-induced mitochondrial damage, lysosomal rupture and ASC oligomerization. Additionally, 4-HAB inhibited the NLRP3 inflammasome through Sirt1-dependent autophagy induction. Furthermore, the anti-inflammatory properties of 4-HAB were confirmed in a mouse model of uric acid crystals-mediated peritonitis by the reduced levels of neutrophil influx, IL-1β, active caspase-1, IL-6 and MCP-1 in lavage fluids. In conclusion, 4-HAB attenuates gouty inflammation, in part by attenuating activation of the NLRP3 inflammasome through the Sirt1/autophagy induction pathway.

scholarly journals Structural mechanism of CARD8 regulation by DPP9

2021 ◽  
Author(s):  
Humayun Sharif ◽  
L. Robert Hollingsworth ◽  
Andrew R. Griswold ◽  
Jeffrey C. Hsiao ◽  
Qinghui Wang ◽  
...  
Keyword(s):  
Pattern Recognition ◽  
Ternary Complex ◽  
Proteasomal Degradation ◽  
Full Length ◽  
Inflammasome Activation ◽  
Structural Mechanism ◽  
Danger Signals ◽  
Caspase 1 ◽  
Recognition Receptor

SUMMARYCARD8 is a germline-encoded pattern recognition receptor that detects intracellular danger signals. Like the related inflammasome sensor NLRP1, CARD8 undergoes constitutive autoprocessing within its function-to-find domain (FIIND), generating two polypeptides that stay associated and autoinhibited. Certain pathogen- and danger-associated activities, including the inhibition of the serine dipeptidases DPP8 and DPP9 (DPP8/9), induce the proteasome-mediated degradation of the N-terminal (NT) fragment, releasing the C-terminal (CT) fragment to form a caspase-1 activating inflammasome. DPP8/9 also bind directly to the CARD8 FIIND, but the role that this interaction plays in CARD8 inflammasome regulation is not yet understood. Here, we solved several cryo-EM structures of CARD8 bound to DPP9, with or without the DPP inhibitor Val-boroPro (VbP), which revealed a ternary complex composed of one DPP9, the full-length CARD8, and one CARD8-CT. Through structure-guided biochemical and cellular experiments, we demonstrated that DPP9’s structure restrains CARD8-CT after proteasomal degradation. Moreover, although DPP inhibitors do not directly displace CARD8 from DPP9 in vitro, we show that they can nevertheless destabilize this complex in cells. Overall, these results demonstrate that DPP8/9 inhibitors cause CARD8 inflammasome activation via at least two distinct mechanisms, one upstream and one downstream of the proteasome.

scholarly journals Role of ASM/Cer/TXNIP Signaling Module in the NLRP3 Inflammasome Activation

2020 ◽  
Author(s):  
Jianjun Jiang ◽  
Jin Yang ◽  
Yining Shi ◽  
Jiyu Cao ◽  
Youjin Lu ◽  
...  
Keyword(s):  
Nlrp3 Inflammasome ◽  
Drug Targets ◽  
Inflammatory Diseases ◽  
Acid Sphingomyelinase ◽  
Inflammasome Activation ◽  
Caspase 1 ◽  
Nlrp3 Inflammasome Activation ◽  
Ceramide Production

Abstract Background: The NLRP3 inflammasome serves as a crucial component in an array of inflammatory conditions by boosting the secretion of pro-inflammatory cytokines: IL-1β and IL-18. Hence, a thorough investigation of the underlying mechanism of NLRP3 activation could ascertain the requisite directionality to the ongoing studies, along with the identification of the novel drug targets for the management of inflammatory diseases. Previous studies have established the vital role of the Acid sphingomyelinase (ASM)/Ceramide (Cer) pathway in the functional outcome of cells, with a particular emphasis on the inflammatory processes. ASM mediates the ceramide production by sphingomyelin hydrolysis. Furthermore, the participation of the ASM/Cer in NLRP3 activation remains ambiguous. Methods: We employed lipopoysaccharide (LPS)/Adenosine Triphosphate (ATP)-induced activation of NLRP3 inflammasome in J774A.1 cells as an in vitro inflammatory model. Results: We observed that imipramine, a well-known inhibitor of ASM, significantly inhibited ASM activity & increased ceramide accumulation, which indicates ASM activation. Besides, it also suppressed the LPS/ATP-induced expression of proteins and mRNA: Thioredoxin interacting protein (TXNIP), NLRP3, Caspase-1, IL-1β and IL-18. Interestingly verapamil, a TXNIP inhibitor, suppressed LPS/ATP-induced TXNIP/NLRP3 inflammasome activation; however, it did not affect LPS/ATP-induced ASM activity and ceramide production. Further examination showed that the exogenous C2-ceramide-treated J774A.1 cells induce the overexpression of TXNIP, NLRP3, Caspase-1, IL-1β, and IL-18. Furthermore, verapamil inhibited C2-Ceramide mediated TXNIP overexpression and NLRP3 inflammasome activation. These findings infer that TXNIP overexpression leads to Cer mediated NLRP3 inflammasome activation. Conclusion: Our study validated the crucial role of the ASM/Cer/TXNIP signaling pathway in NLRP3 inflammasome activation.

scholarly journals Caspase-1 inhibition prevents glial inflammasome activation and pyroptosis in models of multiple sclerosis

2018 ◽  
Vol 115 (26) ◽  
pp. E6065-E6074 ◽  
Author(s):  
Brienne A. McKenzie ◽  
Manmeet K. Mamik ◽  
Leina B. Saito ◽  
Roobina Boghozian ◽  
Maria Chiara Monaco ◽  
...  
Keyword(s):  
Multiple Sclerosis ◽  
Demyelinating Disease ◽  
Inflammatory Diseases ◽  
Disease Process ◽  
Molecular Evidence ◽  
Inflammasome Activation ◽  
Neurobehavioral Performance ◽  
Human Microglia ◽  
Caspase 1

Multiple sclerosis (MS) is a progressive inflammatory demyelinating disease of the CNS of unknown cause that remains incurable. Inflammasome-associated caspases mediate the maturation and release of the proinflammatory cytokines IL-1β and IL-18 and activate the pore-forming protein gasdermin D (GSDMD). Inflammatory programmed cell death, pyroptosis, was recently shown to be mediated by GSDMD. Here, we report molecular evidence for GSDMD-mediated inflammasome activation and pyroptosis in both myeloid cells (macrophages/microglia) and, unexpectedly, in myelin-forming oligodendrocytes (ODCs) in the CNS of patients with MS and in the MS animal model, experimental autoimmune encephalomyelitis (EAE). We observed inflammasome activation and pyroptosis in human microglia and ODCs in vitro after exposure to inflammatory stimuli and demonstrate caspase-1 inhibition by the small-molecule inhibitor VX-765 in both cell types. GSDMD inhibition by siRNA transduction suppressed pyroptosis in human microglia. VX-765 treatment of EAE animals reduced the expression of inflammasome- and pyroptosis-associated proteins in the CNS, prevented axonal injury, and improved neurobehavioral performance. Thus, GSDMD-mediated pyroptosis in select glia cells is a previously unrecognized mechanism of inflammatory demyelination and represents a unique therapeutic opportunity for mitigating the disease process in MS and other CNS inflammatory diseases.

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