The inflammasome is a critical molecular complex that activates interleukin-1 driven inflammation in response to pathogen- and danger-associated signals. Germline mutations in the inflammasome sensor ...NLRP1 cause Mendelian systemic autoimmunity and skin cancer susceptibility, but its endogenous regulation remains less understood. Here we use a proteomics screen to uncover dipeptidyl dipeptidase DPP9 as a novel interacting partner with human NLRP1 and a related inflammasome regulator, CARD8. DPP9 functions as an endogenous inhibitor of NLRP1 inflammasome in diverse primary cell types from human and mice. DPP8/9 inhibition via small molecule drugs and CRISPR/Cas9-mediated genetic deletion specifically activate the human NLRP1 inflammasome, leading to ASC speck formation, pyroptotic cell death, and secretion of cleaved interleukin-1β. Mechanistically, DPP9 interacts with a unique autoproteolytic domain (Function to Find Domain (FIIND)) found in NLRP1 and CARD8. This scaffolding function of DPP9 and its catalytic activity act synergistically to maintain NLRP1 in its inactive state and repress downstream inflammasome activation. We further identified a single patient-derived germline missense mutation in the NLRP1 FIIND domain that abrogates DPP9 binding, leading to inflammasome hyperactivation seen in the Mendelian autoinflammatory disease Autoinflammation with Arthritis and Dyskeratosis. These results unite recent findings on the regulation of murine Nlrp1b by Dpp8/9 and uncover a new regulatory mechanism for the NLRP1 inflammasome in primary human cells. Our results further suggest that DPP9 could be a multifunctional inflammasome regulator involved in human autoinflammatory diseases.
Coatomer complex I (COPI) mediates retrograde vesicular trafficking from Golgi to the endoplasmic reticulum (ER) and within Golgi compartments. Deficiency in subunit alpha causes COPA syndrome and is ...associated with type I IFN signalling, although the upstream innate immune sensor involved was unknown. Using in vitro models we find aberrant activation of the STING pathway due to deficient retrograde but probably not intra-Golgi transport. Further we find the upstream cytosolic DNA sensor cGAS as essentially required to drive type I IFN signalling. Genetic deletion of COPI subunits COPG1 or COPD similarly induces type I IFN activation in vitro, which suggests that inflammatory diseases associated with mutations in other COPI subunit genes may exist. Finally, we demonstrate that inflammation in COPA syndrome patient peripheral blood mononuclear cells and COPI-deficient cell lines is ameliorated by treatment with the small molecule STING inhibitor H-151, suggesting targeted inhibition of the cGAS/STING pathway as a promising therapeutic approach.
Purpose of Review
Type I interferons (IFNαβ) induce the expression of hundreds of genes; thus, it is unsurprising that the initiation, transmission, and resolution of the IFNαβ-mediated immune ...response is tightly controlled. Mutations that alter nucleic acid processing and recognition, ablate IFNαβ-specific negative feedback mechanisms, or result in dysfunction of the proteasome system can all induce pathogenic IFNαβ signalling and are the focus of this review.
Recent Findings
Recent advances have delineated the precise cytoplasmic mechanisms that facilitate self-DNA to be recognised by cGAS and self-RNA to be recognised by RIG-I or MDA-5. This helps clarify interferonopathies associated with mutations in genes which code for DNase-II and ADAR1, among others. Similarly, loss of function mutations in Pol α, which lowers the presence of antagonistic ligands in the cytosol, or gain of function mutations in RIG-I and MDA-5, result in increased propensity for receptor activation and therefore IFNαβ induction.
Summary
As the aetiology of monogenic autoinflammatory diseases are uncovered, novel and sometimes unsuspected molecular interactions and signalling pathways are being defined. This review covers developments that have come to light over the past 3 years, with reference to the study of interferonopathies.
Purpose of Review
Autoinflammatory diseases are driven by abnormal innate immune activation. In the case of inflammasomopathies, these are all attributable to activation of an inflammasome complex, ...nucleated by an innate immune sensor such as NLRP3. This review will focus on recent advances that have helped to elucidate the role of three other sensors (NLRP1, NLRC4 and pyrin) which can also cause inflammasomopathies.
Recent Findings
Mutations in pyrin (S242R or E244K) destroy an inhibitory 14-3-3 binding site and result in the newly characterised disease pyrin-associated autoinflammation with neutrophilic dermatosis (PAAND). Moreover, a separate autoinflammatory disease driven by mevalonate kinase deficiency leads to defective RhoGTPase prenylation and subsequent loss of pyrin S242R phosphorylation, suggesting a shared mechanism of disease. Other inflammasomes such as NLRP1 and NLRC4 have had novel mutations described recently, which inform about the specific domains required for activation and autoinhibition.
Summary
This review covers recent advances in the study of inflammasomopathies, focussing on gene discoveries that elucidate new pathogenic mechanisms.
Human NLRP1 (NACHT, LRR, and PYD domain-containing protein 1) is an innate immune sensor predominantly expressed in the skin and airway epithelium. Here, we report that human NLRP1 senses the ...ultraviolet B (UVB)- and toxin-induced ribotoxic stress response (RSR). Biochemically, RSR leads to the direct hyperphosphorylation of a human-specific disordered linker region of NLRP1 (NLRP1
) by MAP3K20/ZAKα kinase and its downstream effector, p38. Mutating a single ZAKα phosphorylation site in NLRP1
abrogates UVB- and ribotoxin-driven pyroptosis in human keratinocytes. Moreover, fusing NLRP1
to CARD8, which is insensitive to RSR by itself, creates a minimal inflammasome sensor for UVB and ribotoxins. These results provide insight into UVB sensing by human skin keratinocytes, identify several ribotoxins as NLRP1 agonists, and establish inflammasome-driven pyroptosis as an integral component of the RSR.
Purpose of Review
The nuclear factor κB (NF-κB) pathway is tightly regulated through multiple posttranslational mechanisms including ubiquitination. Mutations in these regulatory pathways can cause ...disease and are the focus of this review.
Recent Findings
The linear ubiquitin chain assembly complex (LUBAC) is a trimer made up of HOIL-1L, SHARPIN, and the catalytic subunit HOIP. Loss of function mutations in HOIL-1L and HOIP result in largely overlapping phenotypes, characterized by multi-organ autoinflammation, immunodeficiency, and amylopectinosis. Interestingly, patient fibroblasts exhibited diminished IL-1β- and TNF-induced NF-κB activation, yet monocytes were hyper-responsive to IL-1β, hinting at cell type or target specific roles of LUBAC-mediated ubiquitination. Ubiquitin-driven signaling is counterbalanced by deubiquitinase enzymes (DUBs), such as OTULIN and A20. Hypomorphic mutations in OTULIN result in elevated NF-κB signaling causing an autoinflammatory syndrome. Similarly, patients with high-penetrance heterozygous mutations in the gene encoding A20 (haploinsufficiency of A20 (HA20)) display excessive ubiquitination and increased activity of NF-κB and of NLRP3 inflammasome activation. HA20 patients present with Behçet-like characteristics or an autoimmune lymphoproliferative syndrome (ALPS)-like phenotype, indicating diverse protein functions.
Summary
This review summarizes recent discoveries in the field of NF-kB-related autoinflammatory diseases (relopathies) within the past 3 years and points to several questions that still remain unanswered.
Structural rearrangements of the genome can drive lung tumorigenesis through the generation of fusion genes with oncogenic properties. Advanced genomic approaches have identified the presence of a ...genetic fusion between fibroblast growth factor receptor 3 (FGFR3) and transforming acidic coiled-coil 3 (TACC3) in non-small cell lung cancer (NSCLC), providing a novel target for FGFR inhibition. To interrogate the functional consequences of the FGFR3-TACC3 fusion in the transformation of lung epithelial cells, we generated a novel transgenic mouse model that expresses FGFR3-TACC3 concomitant with loss of the p53 tumor suppressor gene. Intranasal delivery of an Ad5-CMV-Cre virus promoted seromucinous glandular transformation of olfactory cells lining the nasal cavities of FGFR3-TACC3 (LSL-F3T3) mice, which was further accelerated upon loss of p53 (LSL-F3T3/p53). Surprisingly, lung tumors failed to develop in intranasally infected LSL-F3T3 and LSL-F3T3/p53 mice. In line with these observations, we demonstrated that intranasal delivery of Ad5-CMV-Cre induces widespread Cre-mediated recombination in the olfactory epithelium. Intra-tracheal delivery of Ad5-CMV-Cre into the lungs of LSL-F3T3 and LSL-F3T3/p53 mice, however, resulted in the development of lung adenocarcinomas. Taken together, these findings provide in vivo evidence for an oncogenic function of FGFR3-TACC3 in respiratory epithelium.
NLRP1 is an innate immune sensor that can form cytoplasmic inflammasome complexes. Polymorphisms in NLRP1 are linked to asthma; however, there is currently no functional or mechanistic explanation ...for this.
We sought to clarify the role of NLRP1 in asthma pathogenesis.
Results from the GALA II cohort study were used to identify a link between NLRP1 and asthma in Mexican Americans. In vitro and in vivo models for NLRP1 activation were applied to investigate the role of this inflammasome in asthma at the molecular level.
We document the association of an NLRP1 haplotype with asthma for which the single nucleotide polymorphism rs11651270 (M1184V) individually is the most significant. Surprisingly, M1184V increases NLRP1 activation in the context of N-terminal destabilization, but decreases NLRP1 activation on dipeptidyl peptidase 9 inhibition. In vitro studies demonstrate that M1184V increases binding to dipeptidyl peptidase 9, which can account for its inhibitory role in this context. In addition, in vivo data from a mouse model of airway inflammation reveal a protective role for NLRP1 inflammasome activation reducing eosinophilia in this setting.
Linking our in vitro and in vivo results, we found that the NLRP1 variant M1184V reduces inflammasome activation in the context of dipeptidyl peptidase 9 inhibition and could thereby increase asthma severity. Our studies may have implications for the treatment of asthma in patients carrying this variant of NLRP1.
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Proteasome dysfunction can lead to autoinflammatory disease associated with elevated type I interferon (IFN-αβ) and NF-κB signaling; however, the innate immune pathway driving this is currently ...unknown. Here, we identified protein kinase R (PKR) as an innate immune sensor for proteotoxic stress. PKR activation was observed in cellular models of decreased proteasome function and in multiple cell types from patients with proteasome-associated autoinflammatory disease (PRAAS). Furthermore, genetic deletion or small-molecule inhibition of PKR in vitro ameliorated inflammation driven by proteasome deficiency. In vivo, proteasome inhibitor-induced inflammatory gene transcription was blunted in PKR-deficient mice compared with littermate controls. PKR also acted as a rheostat for proteotoxic stress by triggering phosphorylation of eIF2α, which can prevent the translation of new proteins to restore homeostasis. Although traditionally known as a sensor of RNA, under conditions of proteasome dysfunction, PKR sensed the cytoplasmic accumulation of a known interactor, interleukin-24 (IL-24). When misfolded IL-24 egress into the cytosol was blocked by inhibition of the endoplasmic reticulum-associated degradation pathway, PKR activation and subsequent inflammatory signaling were blunted. Cytokines such as IL-24 are normally secreted from cells; therefore, cytoplasmic accumulation of IL-24 represents an internal danger-associated molecular pattern. Thus, we have identified a mechanism by which proteotoxic stress is detected, causing inflammation observed in the disease PRAAS.
Dipeptidyl peptidase 9 (DPP9) is a direct inhibitor of NLRP1, but how it affects inflammasome regulation in vivo is not yet established. Here, we report three families with immune-associated defects, ...poor growth, pancytopenia, and skin pigmentation abnormalities that segregate with biallelic
rare variants. Using patient-derived primary cells and biochemical assays, these variants were shown to behave as hypomorphic or knockout alleles that failed to repress NLRP1. The removal of a single copy of
,
,
, or
, but not
, was sufficient to rescue the lethality of
mutant neonates in mice. Similarly,
deficiency was partially rescued by the inactivation of
, an obligate downstream adapter of the NLRP1 inflammasome, in zebrafish. These experiments suggest that the deleterious consequences of DPP9 deficiency were mostly driven by the aberrant activation of the canonical NLRP1 inflammasome and IL-1β signaling. Collectively, our results delineate a Mendelian disorder of DPP9 deficiency driven by increased NLRP1 activity as demonstrated in patient cells and in two animal models of the disease.
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