Search

Cytosolic caspase-8 is a mediator of death receptor signaling. While caspase-8 expression is lost in some tumors, it is increased in others, indicating a conditional pro-survival function of caspase-8 in cancer. Here, we show that tumor cells employ DNA-damage-induced nuclear caspase-8 to override the p53-dependent G2/M cell-cycle checkpoint. Caspase-8 is upregulated and localized to the nucleus in multiple human cancers, correlating with treatment resistance and poor clinical outcome. Depletion of caspase-8 causes G2/M arrest, stabilization of p53, and induction of p53-dependent intrinsic apoptosis in tumor cells. In the nucleus, caspase-8 cleaves and inactivates the ubiquitin-specific peptidase 28 (USP28), preventing USP28 from de-ubiquitinating and stabilizing wild-type p53. This results in de facto p53 protein loss, switching cell fate from apoptosis toward mitosis. In summary, our work identifies a non-canonical role of caspase-8 exploited by cancer cells to override the p53-dependent G2/M cell-cycle checkpoint., Competing Interests: Declaration of Interests The authors declare no competing interests., (Copyright © 2020 Elsevier Inc. All rights reserved.)

TAK1 is a key mediator of proinflammatory signals. In this issue of Developmental Cell, Naito et al. (2019) report that TAK1 loss from endothelial cells in adult mice results in intestinal and hepatic vascular destruction due to TNF-induced death of endothelial cells. Additionally, endothelial TAK1 deletion reduces tumor burden., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Constitutively activated NF-κB signaling has long been known to be oncogenic. In this issue of Immunity, O'Reilly et al. (2018) unveil a link between loss of NF-κB1, aberrant STAT1 signaling, sterile inflammation, and the increased expression of immune checkpoint molecules as cancer drivers., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Concomitant hepatocyte apoptosis and regeneration is a hallmark of chronic liver diseases (CLDs) predisposing to hepatocellular carcinoma (HCC). Here, we mechanistically link caspase-8-dependent apoptosis to HCC development via proliferation- and replication-associated DNA damage. Proliferation-associated replication stress, DNA damage, and genetic instability are detectable in CLDs before any neoplastic changes occur. Accumulated levels of hepatocyte apoptosis determine and predict subsequent hepatocarcinogenesis. Proliferation-associated DNA damage is sensed by a complex comprising caspase-8, FADD, c-FLIP, and a kinase-dependent function of RIPK1. This platform requires a non-apoptotic function of caspase-8, but no caspase-3 or caspase-8 cleavage. It may represent a DNA damage-sensing mechanism in hepatocytes that can act via JNK and subsequent phosphorylation of the histone variant H2AX., (Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.)

Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is known for specifically killing cancer cells, whereas in resistant cancers, TRAIL/TRAIL-R can promote metastasis via Rac1 and PI3K. It remains unknown, however, whether and to what extent TRAIL/TRAIL-R signaling in cancer cells can affect the immune microenvironment. Here we show that TRAIL-triggered cytokine secretion from TRAIL-resistant cancer cells is FADD dependent and identify the TRAIL-induced secretome to drive monocyte polarization to myeloid-derived suppressor cells (MDSCs) and M2-like macrophages. TRAIL-R suppression in tumor cells impaired CCL2 production and diminished both lung MDSC presence and tumor growth. In accordance, the receptor of CCL2, CCR2, is required to facilitate increased MDSC presence and tumor growth. Finally, TRAIL and CCL2 are co-regulated with MDSC/M2 markers in lung adenocarcinoma patients. Collectively, endogenous TRAIL/TRAIL-R-mediated CCL2 secretion promotes accumulation of tumor-supportive immune cells in the cancer microenvironment, thereby revealing a tumor-supportive immune-modulatory role of the TRAIL/TRAIL-R system in cancer biology., (Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.)

Recruitment of the deubiquitinase CYLD to signaling complexes is mediated by its interaction with HOIP, the catalytically active component of the linear ubiquitin chain assembly complex (LUBAC). Here, we identify SPATA2 as a constitutive direct binding partner of HOIP that bridges the interaction between CYLD and HOIP. SPATA2 recruitment to TNFR1- and NOD2-signaling complexes is dependent on HOIP, and loss of SPATA2 abolishes CYLD recruitment. Deficiency in SPATA2 exerts limited effects on gene activation pathways but diminishes necroptosis induced by tumor necrosis factor (TNF), resembling loss of CYLD. In summary, we describe SPATA2 as a previously unrecognized factor in LUBAC-dependent signaling pathways that serves as an adaptor between HOIP and CYLD, thereby enabling recruitment of CYLD to signaling complexes., (Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.)

Intestinal epithelial cells (IECs) regulate gut immune homeostasis, and impaired epithelial responses are implicated in the pathogenesis of inflammatory bowel diseases (IBD). IEC-specific ablation of nuclear factor κB (NF-κB) essential modulator (NEMO) caused Paneth cell apoptosis and impaired antimicrobial factor expression in the ileum, as well as colonocyte apoptosis and microbiota-driven chronic inflammation in the colon. Combined RelA, c-Rel, and RelB deficiency in IECs caused Paneth cell apoptosis but not colitis, suggesting that NEMO prevents colon inflammation by NF-κB-independent functions. Inhibition of receptor-interacting protein kinase 1 (RIPK1) kinase activity or combined deficiency of Fas-associated via death domain protein (FADD) and RIPK3 prevented epithelial cell death, Paneth cell loss, and colitis development in mice with epithelial NEMO deficiency. Therefore, NEMO prevents intestinal inflammation by inhibiting RIPK1 kinase activity-mediated IEC death, suggesting that RIPK1 inhibitors could be effective in the treatment of colitis in patients with NEMO mutations and possibly in IBD., (Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.)

Ubiquitination and deubiquitination are crucial for assembly and disassembly of signaling complexes. LUBAC-generated linear (M1) ubiquitin is important for signaling via various immune receptors. We show here that the deubiquitinases CYLD and A20, but not OTULIN, are recruited to the TNFR1- and NOD2-associated signaling complexes (TNF-RSC and NOD2-SC), at which they cooperate to limit gene activation. Whereas CYLD recruitment depends on its interaction with LUBAC, but not on LUBAC's M1-chain-forming capacity, A20 recruitment requires this activity. Intriguingly, CYLD and A20 exert opposing effects on M1 chain stability in the TNF-RSC and NOD2-SC. While CYLD cleaves M1 chains, and thereby sensitizes cells to TNF-induced death, A20 binding to them prevents their removal and, consequently, inhibits cell death. Thus, CYLD and A20 cooperatively restrict gene activation and regulate cell death via their respective activities on M1 chains. Hence, the interplay between LUBAC, M1-ubiquitin, CYLD, and A20 is central for physiological signaling through innate immune receptors., (Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.)

Many cancers harbor oncogenic mutations of KRAS. Effectors mediating cancer progression, invasion, and metastasis in KRAS-mutated cancers are only incompletely understood. Here we identify cancer cell-expressed murine TRAIL-R, whose main function ascribed so far has been the induction of apoptosis as a crucial mediator of KRAS-driven cancer progression, invasion, and metastasis and in vivo Rac-1 activation. Cancer cell-restricted genetic ablation of murine TRAIL-R in autochthonous KRAS-driven models of non-small-cell lung cancer (NSCLC) and pancreatic ductal adenocarcinoma (PDAC) reduces tumor growth, blunts metastasis, and prolongs survival by inhibiting cancer cell-autonomous migration, proliferation, and invasion. Consistent with this, high TRAIL-R2 expression correlates with invasion of human PDAC into lymph vessels and with shortened metastasis-free survival of KRAS-mutated colorectal cancer patients., (Copyright © 2015 Elsevier Inc. All rights reserved.)

UBE2L3 is associated with increased susceptibility to numerous autoimmune diseases, but the underlying mechanism is unexplained. By using data from a genome-wide association study of systemic lupus erythematosus (SLE), we observed a single risk haplotype spanning UBE2L3, consistently aligned across multiple autoimmune diseases, associated with increased UBE2L3 expression in B cells and monocytes. rs140490 in the UBE2L3 promoter region showed the strongest association. UBE2L3 is an E2 ubiquitin-conjugating enzyme, specially adapted to function with HECT and RING-in-between-RING (RBR) E3 ligases, including HOIL-1 and HOIP, components of the linear ubiquitin chain assembly complex (LUBAC). Our data demonstrate that UBE2L3 is the preferred E2 conjugating enzyme for LUBAC in vivo, and UBE2L3 is essential for LUBAC-mediated activation of NF-κB. By accurately quantifying NF-κB translocation in primary human cells from healthy individuals stratified by rs140490 genotype, we observed that the autoimmune disease risk UBE2L3 genotype was correlated with basal NF-κB activation in unstimulated B cells and monocytes and regulated the sensitivity of NF-κB to CD40 stimulation in B cells and TNF stimulation in monocytes. The UBE2L3 risk allele correlated with increased circulating plasmablast and plasma cell numbers in SLE individuals, consistent with substantially elevated UBE2L3 protein levels in plasmablasts and plasma cells. These results identify key immunological consequences of the UBE2L3 autoimmune risk haplotype and highlight an important role for UBE2L3 in plasmablast and plasma cell development., (Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.)

Linear ubiquitination is crucial for innate and adaptive immunity. The linear ubiquitin chain assembly complex (LUBAC), consisting of HOIL-1, HOIP, and SHARPIN, is the only known ubiquitin ligase that generates linear ubiquitin linkages. HOIP is the catalytically active LUBAC component. Here, we show that both constitutive and Tie2-Cre-driven HOIP deletion lead to aberrant endothelial cell death, resulting in defective vascularization and embryonic lethality at midgestation. Ablation of tumor necrosis factor receptor 1 (TNFR1) prevents cell death, vascularization defects, and death at midgestation. HOIP-deficient cells are more sensitive to death induction by both tumor necrosis factor (TNF) and lymphotoxin-α (LT-α), and aberrant complex-II formation is responsible for sensitization to TNFR1-mediated cell death in the absence of HOIP. Finally, we show that HOIP's catalytic activity is necessary for preventing TNF-induced cell death. Hence, LUBAC and its linear-ubiquitin-forming activity are required for maintaining vascular integrity during embryogenesis by preventing TNFR1-mediated endothelial cell death., (Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.)

Nucleotide-binding and oligomerization domain (NOD)-like receptors constitute a first line of defense against invading bacteria. X-linked Inhibitor of Apoptosis (XIAP) is implicated in the control of bacterial infections, and mutations in XIAP are causally linked to immunodeficiency in X-linked lymphoproliferative syndrome type-2 (XLP-2). Here, we demonstrate that the RING domain of XIAP is essential for NOD2 signaling and that XIAP contributes to exacerbation of inflammation-induced hepatitis in experimental mice. We find that XIAP ubiquitylates RIPK2 and recruits the linear ubiquitin chain assembly complex (LUBAC) to NOD2. We further show that LUBAC activity is required for efficient NF-κB activation and secretion of proinflammatory cytokines after NOD2 stimulation. Remarkably, XLP-2-derived XIAP variants have impaired ubiquitin ligase activity, fail to ubiquitylate RIPK2, and cannot facilitate NOD2 signaling. We conclude that XIAP and LUBAC constitute essential ubiquitin ligases in NOD2-mediated inflammatory signaling and propose that deregulation of NOD2 signaling contributes to XLP-2 pathogenesis., (Copyright © 2012 Elsevier Inc. All rights reserved.)

TNF is a key inflammatory cytokine. Using a modified tandem affinity purification approach, we identified HOIL-1 and HOIP as functional components of the native TNF-R1 signaling complex (TNF-RSC). Together, they were shown to form a linear ubiquitin chain assembly complex (LUBAC) and to ubiquitylate NEMO. We show that LUBAC binds to ubiquitin chains of different linkage types and that its recruitment to the TNF-RSC is impaired in TRADD-, TRAF2-, and cIAP1/2- but not in RIP1- or NEMO-deficient MEFs. Furthermore, the E3 ligase activity of cIAPs, but not TRAF2, is required for HOIL-1 recruitment to the TNF-RSC. LUBAC enhances NEMO interaction with the TNF-RSC, stabilizes this protein complex, and is required for efficient TNF-induced activation of NF-kappaB and JNK, resulting in apoptosis inhibition. Finally, we demonstrate that sustained stability of the TNF-RSC requires LUBAC's enzymatic activity, thereby adding a third form of ubiquitin linkage to the triggering of TNF signaling by the TNF-RSC.

Human cytomegalovirus (HCMV) can suppress and evade the immune system. We have identified as a mechanism the ability of HCMV to infect dendritic cells (DC), which initiate the antiviral immune response. HCMV-infected DC show enhanced expression of costimulatory molecules. In contrast, MHC molecules are partially downregulated, leading to a reduced antigen-presenting capacity. Moreover, the apoptosis-inducing ligands CD95L (FasL) and tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) are upregulated, thereby enabling HCMV-infected DC to delete activated T lymphocytes. This additional layer of viral defense is complemented by nondeletional mechanisms, which suppress surviving T cells. Thus, infection of DC allows the virus to blunt the antiviral T cell response by a multilayered defense strategy and could play a pivotal role in HCMV-triggered immunosuppression.

Apoptosis induced by tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL/APO-2L) has been shown to exert important functions during various immunological processes. The involvement of the death adaptor proteins FADD/MORT1, TRADD, and RIP and the apoptosis-initiating caspases-8 and -10 in death signaling by the two death-inducing TRAIL receptors 1 and 2 (TRAIL-R1 and TRAIL-R2) are controversial. Analysis of the native TRAIL death-inducing signaling complex (DISC) revealed ligand-dependent recruitment of FADD/MORT1 and caspase-8. Differential precipitation of ligand-stimulated TRAIL receptors demonstrated that FADD/MORT1 and caspase-8 were recruited to TRAIL-R1 and TRAIL-R2 independently of each other. FADD/MORT1- and caspase-8-deficient Jurkat cells expressing only TRAIL-R2 were resistant to TRAIL-induced apoptosis. Thus, FADD/MORT1 and caspase-8 are essential for apoptosis induction via TRAIL-R2.