Proteolytic Processing of Interleukin-1 Family Cytokines: Variations on a Common Theme

Members of the extended interleukin-1 (IL-1) cytokine family, such as IL-1, IL-18, IL-33, and IL-36, play a pivotal role in the initiation and amplification of immune responses. However, deregulated production and/or activation of these cytokines can lead to the development of multiple inflammatory disorders. IL-1 family members share a broadly similar domain organization and receptor signaling pathways. Another striking similarity between IL-1 family members is the requirement for proteolytic processing in order to unlock their full biological potential. Although much emphasis has been put on the role of caspase-1, another emerging theme is the involvement of neutrophil- and mast cell-derived proteases in IL-1 family cytokine processing. Elucidating the regulation of IL-1 family members by proteolytic processing is of great interest for understanding inflammation and immunity. Here, we review the identity of the proteases involved in the proteolytic processing of IL-1 family cytokines and the therapeutic implications in inflammatory disease

Deletion of Nlrp3 protects from inflammation-induced skeletal muscle atrophy

BACKGROUND: Critically ill patients develop atrophic muscle failure, which increases morbidity and mortality. Interleukin-1β (IL-1β) is activated early in sepsis. Whether IL-1β acts directly on muscle cells and whether its inhibition prevents atrophy is unknown. We aimed to investigate if IL-1β activation via the Nlrp3 inflammasome is involved in inflammation-induced atrophy. METHODS: We performed an experimental study and prospective animal trial. The effect of IL-1β on differentiated C2C12 muscle cells was investigated by analyzing gene-and-protein expression, and atrophy response. Polymicrobial sepsis was induced by cecum ligation and puncture surgery in Nlrp3 knockout and wild type mice. Skeletal muscle morphology, gene and protein expression, and atrophy markers were used to analyze the atrophy response. Immunostaining and reporter-gene assays showed that IL-1β signaling is contained and active in myocytes. RESULTS: Immunostaining and reporter gene assays showed that IL-1β signaling is contained and active in myocytes. IL-1β increased Il6 and atrogene gene expression resulting in myocyte atrophy. Nlrp3 knockout mice showed reduced IL-1β serum levels in sepsis. As determined by muscle morphology, organ weights, gene expression, and protein content, muscle atrophy was attenuated in septic Nlrp3 knockout mice, compared to septic wild-type mice 96 h after surgery. CONCLUSIONS: IL-1β directly acts on myocytes to cause atrophy in sepsis. Inhibition of IL-1β activation by targeting Nlrp3 could be useful to prevent inflammation-induced muscle failure in critically ill patients

Neutrophil-Derived Proteases Escalate Inflammation through Activation of IL-36 Family Cytokines

Recent evidence has strongly implicated the IL-1 family cytokines IL-36α, IL-36β, and IL-36γ as key initiators of skin inflammation. Similar to the other members of the IL-1 family, IL-36 cytokines are expressed as inactive precursors and require proteolytic processing for activation; however, the responsible proteases are unknown. Here, we show that IL-36α, IL-36β, and IL-36γ are activated differentially by the neutrophil granule-derived proteases cathepsin G, elastase, and proteinase-3, increasing their biological activity ∼500-fold. Active IL-36 promoted a strong pro-inflammatory signature in primary keratinocytes and was sufficient to perturb skin differentiation in a reconstituted 3D human skin model, producing features resembling psoriasis. Furthermore, skin eluates from psoriasis patients displayed significantly elevated cathepsin G-like activity that was sufficient to activate IL-36β. These data identify neutrophil granule proteases as potent IL-36-activating enzymes, adding to our understanding of how neutrophils escalate inflammatory reactions. Inhibition of neutrophil-derived proteases may therefore have therapeutic benefits in psoriasis

Trabid epigenetically drives expression of IL-12 and IL-23

The production of interleukin 12 (IL-12) and IL-23 in dendritic cells is strictly regulated via epigenetic silencing. This transcriptional repression is overcome with the help of the deubiquitinase Trabid and has functional implications in a mouse model of multiple sclerosis

Defining the combinatorial space of PKC::CARD-CC signal transduction nodes.

peer reviewedSignal transduction typically displays a so-called bow-tie topology: Multiple receptors lead to multiple cellular responses but the signals all pass through a narrow waist of central signaling nodes. One such signaling node for several inflammatory and oncogenic signaling pathways is the CARD-CC/BCL10/MALT1 (CBM) complexes, which get activated by protein kinase C (PKC)-mediated phosphorylation of the caspase activation and recruitment domain (CARD)-coiled-coil domain (CC) component. In humans, there are four CARD-CC family proteins (CARD9, CARD10, CARD11, and CARD14) and 9 true PKC isozymes (α to ι). At this moment, less than a handful of PKC::CARD-CC relationships are known. In order to explore the biologically relevant combinatorial space out of all 36 potential permutations in this two-component signaling event, we made use of CARD10-deficient human embryonic kidney 293T cells for subsequent pairwise cotransfections of all CARD-CC family members and all activated PKCs. Upon analysis of NF-κB-dependent reporter gene expression, we could define specific PKC::CARD-CC relationships. Surprisingly, as many as 21 PKC::CARD-CC functional combinations were identified. CARD10 was responsive to most PKCs, while CARD14 was mainly activated by PKCδ. The CARD11 activation profile was most similar to that of CARD9. We also discovered the existence of mixed protein complexes between different CARD-CC proteins, which was shown to influence their PKC response profile. Finally, multiple PKCs were found to use a common phosphorylation site to activate CARD9, while additional phosphorylation sites contribute to CARD14 activation. Together, these data reveal the combinatorial space of PKC::CARD-CC signal transduction nodes, which will be valuable for future studies on the regulation of CBM signaling

CARD14 Signalling Ensures Cell Survival and Cancer Associated Gene Expression in Prostate Cancer Cells.

peer reviewedProstate cancer (PCa) is one of the most common cancer types in men and represents an increasing global problem due to the modern Western lifestyle. The signalling adapter protein CARD14 is specifically expressed in epithelial cells, where it has been shown to mediate NF-κB signalling, but a role for CARD14 in carcinoma has not yet been described. By analysing existing cancer databases, we found that CARD14 overexpression strongly correlates with aggressive PCa in human patients. Moreover, we showed that CARD14 is overexpressed in the LNCaP PCa cell line and that knockdown of CARD14 severely reduces LNCaP cell survival. Similarly, knockdown of BCL10 and MALT1, which are known to form a signalling complex with CARD14, also induced LNCaP cell death. MALT1 is a paracaspase that mediates downstream signalling by acting as a scaffold, as well as a protease. Recent studies have already indicated a role for the scaffold function of MALT1 in PCa cell growth. Here, we also demonstrated constitutive MALT1 proteolytic activity in several PCa cell lines, leading to cleavage of A20 and CYLD. Inhibition of MALT1 protease activity did not affect PCa cell survival nor activation of NF-κB and JNK signalling, but reduced expression of cancer-associated genes, including the cytokine IL-6. Taken together, our results revealed a novel role for CARD14-induced signalling in regulating PCa cell survival and gene expression. The epithelial cell type-specific expression of CARD14 may offer novel opportunities for more specific therapeutic targeting approaches in PCa

Granzyme B-dependent proteolysis acts as a switch to enhance the proinflammatory activity of IL-1α

Granzyme B is a cytotoxic lymphocyte-derived protease that plays a central role in promoting apoptosis of virus-infected target cells, through direct proteolysis and activation of constituents of the cell death machinery. However, previous studies have also implicated granzymes A and B in the production of proinflammatory cytokines, via a mechanism that remains undefined. Here we show that IL-1α is a substrate for granzyme B and that proteolysis potently enhanced the biological activity of this cytokine in vitro as well as in vivo. Consistent with this, compared with full-length IL-1α, granzyme B-processed IL-1α exhibited more potent activity as an immunoadjuvant in vivo. Furthermore, proteolysis of IL-1α within the same region, by proteases such as calpain and elastase, was also found to enhance its biological potency. Thus, IL-1α processing by multiple immune-related proteases, including granzyme B, acts as a switch to enhance the proinflammatory properties of this cytokin