RESUMO
Tumor-associated macrophages (TAMs) display pro-tumorigenic phenotypes for supporting tumor progression in response to microenvironmental cues imposed by tumor and stromal cells. However, the underlying mechanisms by which tumor cells instruct TAM behavior remain elusive. Here, we uncover that tumor-cell-derived glucosylceramide stimulated unconventional endoplasmic reticulum (ER) stress responses by inducing reshuffling of lipid composition and saturation on the ER membrane in macrophages, which induced IRE1-mediated spliced XBP1 production and STAT3 activation. The cooperation of spliced XBP1 and STAT3 reinforced the pro-tumorigenic phenotype and expression of immunosuppressive genes. Ablation of XBP1 expression with genetic manipulation or ameliorating ER stress responses by facilitating LPCAT3-mediated incorporation of unsaturated lipids to the phosphatidylcholine hampered pro-tumorigenic phenotype and survival in TAMs. Together, we uncover the unexpected roles of tumor-cell-produced lipids that simultaneously orchestrate macrophage polarization and survival in tumors via induction of ER stress responses and reveal therapeutic targets for sustaining host antitumor immunity.
Assuntos
Estresse do Retículo Endoplasmático , Retículo Endoplasmático/metabolismo , Membranas Intracelulares/metabolismo , Ativação de Macrófagos , Melanoma/metabolismo , Lipídeos de Membrana/metabolismo , Neoplasias Cutâneas/metabolismo , Macrófagos Associados a Tumor/metabolismo , Animais , Linhagem Celular Tumoral , Sobrevivência Celular , Retículo Endoplasmático/ultraestrutura , Glucosilceramidase/metabolismo , Membranas Intracelulares/ultraestrutura , Melanoma/genética , Melanoma/ultraestrutura , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Fenótipo , Fosforilação , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Fator de Transcrição STAT3/genética , Fator de Transcrição STAT3/metabolismo , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/ultraestrutura , Evasão Tumoral , Microambiente Tumoral , Macrófagos Associados a Tumor/ultraestrutura , Proteína 1 de Ligação a X-Box/genética , Proteína 1 de Ligação a X-Box/metabolismoRESUMO
How inflammatory caspases trigger pyroptotic cell death is mostly unexplained. In this issue of Immunity, Núñez and colleagues report that caspase-11 cleaves the transmembrane channel pannexin-1, causing an efflux of cellular ATP that promotes a P2X7 receptor-dependent pyroptosis.
Assuntos
Caspases/metabolismo , Conexinas/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Piroptose/fisiologia , Receptores Purinérgicos P2X7/metabolismo , Choque Séptico/metabolismo , Animais , HumanosRESUMO
BACKGROUND: The cryopyrin-associated periodic syndromes (CAPS) comprise a group of rare autoinflammatory diseases caused by gain-of-function mutations in the NLRP3 gene. NLRP3 contains a leucine-rich repeats (LRR) domain with a highly conserved exonic organization that is subjected to extensive alternative splicing. Aberrant NLRP3 inflammasome assembly in CAPS causes chronic inflammation; however, the mechanisms regulating inflammasome function remain unclear. OBJECTIVE: We aimed to elucidate the mechanisms regulating NLRP3-mediated autoinflammation in human disease, characterizing the role of LRR in inflammasome activation. METHODS: We analyzed sequence read archive data to characterize the pattern of NLRP3 splicing in human monocytes and investigated the role of each LRR-coding exon in inflammasome regulation in genetically modified U937 cells representing CAPS and healthy conditions. RESULTS: We detected a range of NLRP3 splice variants in human primary cells and monocytic cell lines, including 2 yet-undescribed splice variants. We observe that lipopolysaccharides affect the abundance of certain splice variants, suggesting that they may regulate NLRP3 activation by affecting alternative splicing. We showed that exons 4, 5, 7, and 9 are essential for inflammasome function, both in the context of wild-type NLRP3 activation by the agonist molecule nigericin and in a model of CAPS-mediated NLRP3 inflammasome assembly. Moreover, the SGT1-NLRP3 interaction is decreased in nonfunctional variants, suggesting that alternative splicing may regulate the recruitment of proteins that facilitate inflammasome assembly. CONCLUSION: These findings demonstrate the contribution of the LRR domain in inflammasome function and suggest that navigating LRR exon usage within NLRP3 is sufficient to dampen inflammasome assembly in CAPS.
Assuntos
Síndromes Periódicas Associadas à Criopirina , Inflamassomos , Humanos , Inflamassomos/metabolismo , Síndromes Periódicas Associadas à Criopirina/genética , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Leucina/genética , Mutação com Ganho de FunçãoRESUMO
Inflammasomes are intracellular multiprotein signaling platforms that initiate inflammatory responses in response to pathogens and cellular damage. Active inflammasomes induce the enzymatic activity of caspase-1, resulting in the induction of inflammatory cell death, pyroptosis, and the maturation and secretion of inflammatory cytokines IL-1ß and IL-18. Inflammasomes are activated in many inflammatory diseases, including autoinflammatory disorders and arthritis, and inflammasome-specific therapies are under development for the treatment of inflammatory conditions. In this review, we outline the different inflammasome platforms and recent findings contributing to our knowledge about inflammasome biology in health and disease. In particular, we discuss the role of the inflammasome in the pathogenesis of arthritic diseases, including rheumatoid arthritis, gout, ankylosing spondylitis, and juvenile idiopathic arthritis, and the potential of newly developed therapies that specifically target the inflammasome or its products for the treatment of inflammatory diseases.
Assuntos
Artrite/metabolismo , Inflamassomos/metabolismo , Inflamação/metabolismo , Animais , Artrite/imunologia , Proteínas Adaptadoras de Sinalização CARD/metabolismo , Humanos , Imunidade Inata , Inflamação/imunologia , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Proteínas de Neoplasias/metabolismo , Transdução de SinaisRESUMO
Sensors of pathogens, such as Toll-like receptors (TLRs), detect microbes to activate transcriptional programs that orchestrate adaptive responses to specific insults. Here we report that TLR4 and TLR2 specifically activated the endoplasmic reticulum (ER) stress sensor kinase IRE1alpha and its downstream target, the transcription factor XBP1. Previously described ER-stress target genes of XBP1 were not induced by TLR signaling. Instead, TLR-activated XBP1 was required for optimal and sustained production of proinflammatory cytokines in macrophages. Consistent with that finding, activation of IRE1alpha by ER stress acted in synergy with TLR activation for cytokine production. Moreover, XBP1 deficiency resulted in a much greater bacterial burden in mice infected with the TLR2-activating human intracellular pathogen Francisella tularensis. Our findings identify an unsuspected critical function for XBP1 in mammalian host defenses.
Assuntos
Proteínas de Ligação a DNA/metabolismo , Francisella tularensis/imunologia , Imunidade Inata , Macrófagos/metabolismo , Fatores de Transcrição/metabolismo , Tularemia/imunologia , Animais , Linhagem Celular , Citocinas/biossíntese , Citocinas/genética , Citocinas/imunologia , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/imunologia , Endorribonucleases/genética , Endorribonucleases/imunologia , Endorribonucleases/metabolismo , Francisella tularensis/patogenicidade , Lipopeptídeos/farmacologia , Lipopolissacarídeos/farmacologia , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Macrófagos/patologia , Macrófagos/virologia , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/imunologia , Glicoproteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C3H , Camundongos Knockout , Camundongos Mutantes , Fator 88 de Diferenciação Mieloide/genética , Fator 88 de Diferenciação Mieloide/metabolismo , NADPH Oxidase 2 , NADPH Oxidases/genética , NADPH Oxidases/imunologia , NADPH Oxidases/metabolismo , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/imunologia , Proteínas Serina-Treonina Quinases/metabolismo , Processamento de Proteína/efeitos dos fármacos , Processamento de Proteína/genética , Processamento de Proteína/imunologia , RNA Interferente Pequeno/genética , Fatores de Transcrição de Fator Regulador X , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Estresse Fisiológico/efeitos dos fármacos , Estresse Fisiológico/genética , Estresse Fisiológico/imunologia , Fator 6 Associado a Receptor de TNF/genética , Fator 6 Associado a Receptor de TNF/metabolismo , Receptor 2 Toll-Like/genética , Receptor 2 Toll-Like/imunologia , Receptor 2 Toll-Like/metabolismo , Receptor 4 Toll-Like/genéticaRESUMO
Recognition of pathogens and altered self must be efficient and highly specific to orchestrate appropriate responses while limiting excessive inflammation and autoimmune reaction to normal self. AIM2 is a member of innate immune sensors that detects the presence of DNA, arguably the most conserved molecules in living organisms. However, AIM2 achieves specificity by detecting altered or mislocalized DNA molecules. It can detect damaged DNA, and the aberrant presence of DNA within the cytosolic compartment such as genomic DNA released into the cytosol upon loss of nuclear envelope integrity. AIM2 is also a key sensor of pathogens that detects the presence of foreign DNA accumulating in the cytosol during the life cycle of intracellular pathogens including viruses, bacteria, and parasites. AIM2 activation initiates the assembly of the inflammasome, an innate immune complex that leads to the activation of inflammatory caspases. This triggers the maturation and secretion of the cytokines IL-1ß and IL-18. It can also initiate pyroptosis, a proinflammatory form of cell death. The AIM2 inflammasome contributes to physiological responses and diseases. It is a key player in host defenses, but its deregulation can contribute immune-linked diseases, such as autoinflammatory and autoimmune pathologies. Moreover, AIM2 may play a role in cancer development. Recent studies have shown that the detection of self-DNA species by AIM2 is an important factor that contributes to diseases associated with perturbation of cellular homeostasis. Thus, in addition of being a sensor of pathogen associated molecular patterns (PAMPs), the AIM2 inflammasome is emerging as a key guardian of cellular integrity.
Assuntos
Dano ao DNA/imunologia , Proteínas de Ligação a DNA/metabolismo , Inflamassomos/metabolismo , Animais , Caspases/metabolismo , Morte Celular , DNA/imunologia , Homeostase , Interações Hospedeiro-Patógeno , Humanos , Imunidade Inata , Moléculas com Motivos Associados a Patógenos/imunologiaRESUMO
Danger signals released by damaged organelles can promote inflammation. In this issue of Immunity, Shimada et al. (2012) report that oxidized DNA, released by mitochondria, directly binds and activates the NLRP3 inflammasome.
RESUMO
The inflammasome is a major component of the innate immune system, and its main function is to activate caspase-1, a cysteine protease that promotes inflammation by inducing interleukin-1ß (IL-1ß) maturation and release into the extracellular milieu. To prevent uncontrolled inflammation, this complex is highly regulated. When it is assembled, the inflammasome is insoluble, which has long precluded the analysis of its interactions with other proteins. Here we used the proximity-dependent biotinylation assay (BioID) to identify proteins associated with caspase-1 during inflammasome activation. Using the BioID in a cell-free system in which the inflammasome had been activated, we found that a caspase-1-biotin ligase fusion protein selectively labeled 111 candidates, including the p62/sequestosome-1 protein (p62). Using co-immunoprecipitation experiments, we demonstrated that p62 interacts with caspase-1. This interaction promoted caspase-1-mediated cleavage of p62 at Asp-329. Mechanistic and functional analyses revealed that caspase-1-mediated cleavage of p62 leads to loss of its interaction with the autophagosomal protein microtubule-associated protein 1 light chain 3 ß (LC3B). Strikingly, overexpression of a p62 N-terminal fragment generated upon caspase-1 cleavage decreased IL-1ß release, whereas overexpression of p62's C-terminal portion enhanced IL-1ß release, by regulating pro-IL1ß levels. Overall, the overexpression of both fragments together decreased IL-1ß release. Taken together, our results indicate that caspase-1-mediated p62 cleavage plays a complex role in balancing caspase-1-induced inflammation.
Assuntos
Apoptose , Caspase 1/metabolismo , Inflamassomos , Interleucina-1beta/metabolismo , Proteína Sequestossoma-1/metabolismo , Coloração e Rotulagem/métodos , Animais , Bioensaio , Biotinilação , Caspase 1/genética , Células HEK293 , Humanos , Camundongos , Proteína Sequestossoma-1/genéticaRESUMO
A variety of stimuli, including monosodium urate (MSU) crystals, activate the NLRP3 inflammasome, and this activation involves several molecular mechanisms including xanthine oxidase (XO) up-regulation and mitochondrial dysfunction. Upon oligomerization of apoptosis-associated speck-like protein containing a CARD (ASC), caspase-1 becomes active and cleaves the proinflammatory cytokine IL-1ß into its active secreted form. Hydrogen sulfide (H2S), a gasotransmitter mainly produced by cystathionine γ-lyase (CSE) in macrophages, could modulate inflammation. Here, we sought to investigate the effects of exogenous and endogenous H2S on NLRP3 inflammasome activation in vitro and in vivo Primed bone marrow-derived macrophages (BMDM) isolated from wildtype (wt) or CSE-deficient mice and human macrophages (THP1 cells and primary macrophages), were stimulated with MSU crystals in the presence or absence of a H2S donor, sodium thiosulfate (STS) or GYY4137 (GYY). In murine and human macrophages in vitro, both STS and GYY inhibited MSU crystal-induced IL-1ß secretion in a dose-dependent manner. Moreover, the H2S donors inhibited MSU crystal-induced XO/caspase-1 activities, mitochondrial reactive oxygen species (ROS) generation, and ASC oligomerization. Accordingly, IL-1ß secretion and XO/caspase-1 activities were higher in CSE-deficient BMDMs than in wt BMDMs. For in vivo studies, we experimentally induced peritonitis by intraperitoneal injection of MSU crystals into mice. GYY pretreatment ameliorated inflammation, evidenced by decreased IL-6/monocyte chemoattractant protein-1 (MCP-1) released into peritoneal lavages. Taken together, our results suggest that both exogenous (via H2S donors) and endogenous (via CSE) H2S production may represent approaches for managing, for example, acute gout or other inflammation conditions.
Assuntos
Sulfeto de Hidrogênio/imunologia , Inflamassomos/imunologia , Inflamação/imunologia , Macrófagos/imunologia , Proteína 3 que Contém Domínio de Pirina da Família NLR/imunologia , Animais , Humanos , Inflamassomos/genética , Inflamação/genética , Interleucina-1beta/genética , Interleucina-1beta/imunologia , Interleucina-6/genética , Interleucina-6/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Proteína 3 que Contém Domínio de Pirina da Família NLR/genéticaRESUMO
The endoplasmic reticulum kinase inositol-requiring enzyme 1 (IRE1) and its downstream target X-box-binding protein 1 (XBP1) drive B-cell differentiation toward plasma cells and have been shown to contribute to multiple myeloma development; yet, little is known of the role of this pathway in diffuse large B-cell lymphoma (DLBCL). Here, we show that in the germinal center B-cell-like (GCB) DLBCL subtype, IRE1 expression is reduced to a level that prevents XBP1 activation. Gene expression profiles indicated that, in GCB DLBCL cancer samples, expression of IRE1 messenger RNA was inversely correlated with the levels and activity of the epigenetic repressor, histone methyltransferase enhancer of zeste homolog 2 (EZH2). Correspondingly, in GCB-derived cell lines, the IRE1 promoter carried increased levels of the repressive epigenetic mark histone 3 lysine 27 trimethylation. Pharmacological inhibition of EZH2 erased those marks and restored IRE1 expression and function in vitro and in vivo. Moreover, reconstitution of the IRE1-signaling pathway, by expression of the XBP1-active form, compromised GCB DLBCL tumor growth in a mouse xenograft cancer model. These findings indicate that IRE1-XBP1 downregulation distinguishes GCB DLBCL from other DLBCL subtypes and contributes to tumor growth.
Assuntos
Linfócitos B/imunologia , Endorribonucleases/genética , Regulação Neoplásica da Expressão Gênica , Centro Germinativo/imunologia , Linfoma Difuso de Grandes Células B/imunologia , Proteínas Serina-Treonina Quinases/genética , RNA Mensageiro/genética , Proteína 1 de Ligação a X-Box/genética , Animais , Linfócitos B/patologia , Diferenciação Celular , Linhagem Celular Tumoral , Proliferação de Células , Endorribonucleases/antagonistas & inibidores , Endorribonucleases/imunologia , Proteína Potenciadora do Homólogo 2 de Zeste/antagonistas & inibidores , Proteína Potenciadora do Homólogo 2 de Zeste/genética , Proteína Potenciadora do Homólogo 2 de Zeste/imunologia , Epigênese Genética , Centro Germinativo/patologia , Histonas/genética , Histonas/imunologia , Humanos , Indazóis/farmacologia , Leupeptinas/farmacologia , Linfoma Difuso de Grandes Células B/genética , Linfoma Difuso de Grandes Células B/patologia , Linfoma Difuso de Grandes Células B/terapia , Camundongos , Camundongos da Linhagem 129 , Plasmócitos/imunologia , Plasmócitos/patologia , Regiões Promotoras Genéticas , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/imunologia , Piridonas/farmacologia , RNA Mensageiro/antagonistas & inibidores , RNA Mensageiro/imunologia , Transdução de Sinais , Proteína 1 de Ligação a X-Box/antagonistas & inibidores , Proteína 1 de Ligação a X-Box/imunologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Periodic fever with aphthous stomatitis, pharyngitis, and cervical adenitis (PFAPA) is a relatively common autoinflammatory condition that primarily affects children. Although tendencies were reported for this syndrome, genetic variations influencing risk and disease progression are poorly understood. In this study, we performed next-generation sequencing for 82 unrelated PFAPA patients and identified a frameshift variant in the CARD8 gene (CARD8-FS). Subsequently, we compared the frequency of CARD8-FS carriers in our PFAPA cohort (13.9%) with a healthy local population group (3.2%) and found a significant association between the CARD8-FS polymorphism and risk for PFAPA syndrome (p = 0.012; odds ratio: 4.96 [95% confidence interval, 1.33-18.47]). Moreover, CARD8-FS carriers display a distinct PFAPA phenotype that is characterized by a higher prevalence of symptoms out of flares and oral aphthosis (both p = 0.02 compared with PFAPA patients without the frameshift variant). CARD8 encodes a protein component of the NLRP3 inflammasome, which plays an important role in inflammation and contributes to the pathology of various autoinflammatory diseases. We found that the CARD8-FS variant led to a truncated CARD8 protein lacking the FIIND and CARD domains. As a result, the mutant CARD8 protein lost the ability to interact with the NOD domain of NLRP3. In summary, these results identify a new CARD8 variant associated with PFAPA and further suggest that disruption of the interaction between CARD8 and NLRP3 can regulate autoinflammation in patients.
Assuntos
Proteínas Adaptadoras de Sinalização CARD/genética , Febre/genética , Mutação da Fase de Leitura/genética , Doenças Hereditárias Autoinflamatórias/genética , Proteínas de Neoplasias/genética , Proteínas Adaptadoras de Sinalização CARD/metabolismo , Criança , Análise Mutacional de DNA , Frequência do Gene , Estudos de Associação Genética , Predisposição Genética para Doença , Genótipo , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Inflamassomos/metabolismo , Linfadenite , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Proteínas de Neoplasias/metabolismo , Faringite , Polimorfismo de Nucleotídeo Único , Ligação Proteica/genética , Risco , Estomatite Aftosa , SíndromeRESUMO
Inhibitors of the HIV aspartyl protease [HIV protease inhibitors (HIV-PIs)] are the cornerstone of treatment for HIV. Beyond their well-defined antiretroviral activity, these drugs have additional effects that modulate cell viability and homeostasis. However, little is known about the virus-independent pathways engaged by these molecules. Here we show that the HIV-PI Nelfinavir decreases translation rates and promotes a transcriptional program characteristic of the integrated stress response (ISR). Mice treated with Nelfinavir display hallmarks of this stress response in the liver, including α subunit of translation initiation factor 2 (eIF2α) phosphorylation, activating transcription factor-4 (ATF4) induction, and increased expression of known downstream targets. Mechanistically, Nelfinavir-mediated ISR bypassed direct activation of the eIF2α stress kinases and instead relied on the inhibition of the constitutive eIF2α dephosphorylation and down-regulation of the phophatase cofactor CReP (Constitutive Repressor of eIF2α Phosphorylation; also known as PPP1R15B). These findings demonstrate that the modulation of eIF2α-specific phosphatase cofactor activity can be a rheostat of cellular homeostasis that initiates a functional ISR and suggest that the HIV-PIs could be repositioned as therapeutics in human diseases to modulate translation rates and stress responses.
Assuntos
Fator de Iniciação 2 em Eucariotos/metabolismo , Inibidores da Protease de HIV/farmacologia , Protease de HIV/metabolismo , Estresse Fisiológico/efeitos dos fármacos , Fator 4 Ativador da Transcrição/genética , Fator 4 Ativador da Transcrição/metabolismo , Animais , Células HeLa , Humanos , Fígado/efeitos dos fármacos , Fígado/metabolismo , Proteínas de Membrana/deficiência , Proteínas de Membrana/metabolismo , Camundongos , Nelfinavir/farmacologia , Fosforilação/efeitos dos fármacos , Proteína Fosfatase 1/metabolismo , Proteínas Serina-Treonina Quinases/deficiência , Proteínas Serina-Treonina Quinases/metabolismo , RNA Interferente Pequeno/metabolismo , Transcrição Gênica/efeitos dos fármacos , Resposta a Proteínas não Dobradas/efeitos dos fármacos , eIF-2 Quinase/metabolismoRESUMO
Inflammasomes are critical sensors that convey cellular stress and pathogen presence to the immune system by activating inflammatory caspases and cytokines such as IL-1ß. The nature of endogenous stress signals that activate inflammasomes remains unclear. Here we show that an inhibitor of the HIV aspartyl protease, Nelfinavir, triggers inflammasome formation and elicits an IL-1R-dependent inflammation in mice. We found that Nelfinavir impaired the maturation of lamin A, a structural component of the nuclear envelope, thereby promoting the release of DNA in the cytosol. Moreover, deficiency of the cytosolic DNA-sensor AIM2 impaired Nelfinavir-mediated inflammasome activation. These findings identify a pharmacologic activator of inflammasome and demonstrate the role of AIM2 in detecting endogenous DNA release upon perturbation of nuclear envelope integrity.
Assuntos
Inflamassomos/efeitos dos fármacos , Nelfinavir/farmacologia , Membrana Nuclear/efeitos dos fármacos , Animais , Proteínas Adaptadoras de Sinalização CARD/fisiologia , Caspase 1/metabolismo , DNA/metabolismo , Inflamassomos/fisiologia , Interleucina-1beta/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Proteína 3 que Contém Domínio de Pirina da Família NLR/fisiologia , Membrana Nuclear/fisiologia , Receptores de Interleucina-1/fisiologiaRESUMO
Stress induced by accumulation of unfolded proteins at the endoplasmic reticulum (ER) is a classic feature of secretory cells and is observed in many tissues in human diseases including cancer, diabetes, obesity, and neurodegeneration. Cellular adaptation to ER stress is achieved by the activation of the unfolded protein response (UPR), an integrated signal transduction pathway that transmits information about the protein folding status at the ER to the nucleus and cytosol to restore ER homeostasis. Inositol-requiring transmembrane kinase/endonuclease-1 (IRE1α), the most conserved UPR stress sensor, functions as an endoribonuclease that processes the mRNA of the transcription factor X-box binding protein-1 (XBP1). IRE1α signaling is a highly regulated process, controlled by the formation of a dynamic scaffold onto which many regulatory components assemble, here referred to as the UPRosome. Here we provide an overview of the signaling and regulatory mechanisms underlying IRE1α function and discuss the emerging role of the UPR in adaptation to protein folding stress in specialized secretory cells and in pathological conditions associated with alterations in ER homeostasis.
Assuntos
Endorribonucleases/fisiologia , Proteínas Serina-Treonina Quinases/fisiologia , Desdobramento de Proteína , Transdução de Sinais/fisiologia , Estresse Fisiológico/fisiologia , Proteínas de Ligação a DNA/fisiologia , Retículo Endoplasmático/fisiologia , Homeostase/fisiologia , Humanos , Fatores de Transcrição de Fator Regulador X , Fatores de Transcrição/fisiologia , Proteína 1 de Ligação a X-BoxRESUMO
Activation of the elongation factor 2 kinase (eEF2K) leads to the phosphorylation and inhibition of the elongation factor eEF2, reducing mRNA translation rates. Emerging evidence indicates that the regulation of factors involved in protein synthesis may be critical for controlling diverse biological processes including cancer progression. Here we show that inhibitors of the HIV aspartyl protease (HIV-PIs), nelfinavir in particular, trigger a robust activation of eEF2K leading to the phosphorylation of eEF2. Beyond its anti-viral effects, nelfinavir has antitumoral activity and promotes cell death. We show that nelfinavir-resistant cells specifically evade eEF2 inhibition. Decreased cell viability induced by nelfinavir is impaired in cells lacking eEF2K. Moreover, nelfinavir-mediated anti-tumoral activity is severely compromised in eEF2K-deficient engrafted tumors in vivo Our findings imply that exacerbated activation of eEF2K is detrimental for tumor survival and describe a mechanism explaining the anti-tumoral properties of HIV-PIs.
Assuntos
Quinase do Fator 2 de Elongação/metabolismo , Neoplasias/metabolismo , Neoplasias/patologia , Proteínas Quinases Ativadas por AMP/metabolismo , Animais , Morte Celular/efeitos dos fármacos , Morte Celular/genética , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Modelos Animais de Doenças , Progressão da Doença , Relação Dose-Resposta a Droga , Resistência a Medicamentos/genética , Quinase do Fator 2 de Elongação/genética , Feminino , Expressão Gênica , Humanos , Alvo Mecanístico do Complexo 1 de Rapamicina , Camundongos , Camundongos Knockout , Complexos Multiproteicos/metabolismo , Nelfinavir/química , Nelfinavir/farmacologia , Neoplasias/genética , Fator 2 de Elongação de Peptídeos/metabolismo , Fosforilação , Biossíntese de Proteínas , Serina-Treonina Quinases TOR/metabolismo , Carga TumoralRESUMO
Spontaneous CD8 T-cell responses occur in growing tumors but are usually poorly effective. Understanding the molecular and cellular mechanisms that drive these responses is of major interest as they could be exploited to generate a more efficacious antitumor immunity. As such, stimulator of IFN genes (STING), an adaptor molecule involved in cytosolic DNA sensing, is required for the induction of antitumor CD8 T responses in mouse models of cancer. Here, we find that enforced activation of STING by intratumoral injection of cyclic dinucleotide GMP-AMP (cGAMP), potently enhanced antitumor CD8 T responses leading to growth control of injected and contralateral tumors in mouse models of melanoma and colon cancer. The ability of cGAMP to trigger antitumor immunity was further enhanced by the blockade of both PD1 and CTLA4. The STING-dependent antitumor immunity, either induced spontaneously in growing tumors or induced by intratumoral cGAMP injection was dependent on type I IFNs produced in the tumor microenvironment. In response to cGAMP injection, both in the mouse melanoma model and an ex vivo model of cultured human melanoma explants, the principal source of type I IFN was not dendritic cells, but instead endothelial cells. Similarly, endothelial cells but not dendritic cells were found to be the principal source of spontaneously induced type I IFNs in growing tumors. These data identify an unexpected role of the tumor vasculature in the initiation of CD8 T-cell antitumor immunity and demonstrate that tumor endothelial cells can be targeted for immunotherapy of melanoma.
Assuntos
Células Endoteliais/metabolismo , Imunidade , Proteínas de Membrana/metabolismo , Neoplasias/imunologia , Neoplasias/terapia , Animais , Antígenos de Neoplasias/imunologia , Linfócitos T CD8-Positivos/imunologia , Antígeno CTLA-4/imunologia , Proliferação de Células/efeitos dos fármacos , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/imunologia , Modelos Animais de Doenças , Relação Dose-Resposta Imunológica , Células Endoteliais/efeitos dos fármacos , Injeções Intralesionais , Interferon Tipo I/metabolismo , Linfócitos do Interstício Tumoral/efeitos dos fármacos , Linfócitos do Interstício Tumoral/imunologia , Melanoma/imunologia , Melanoma/patologia , Melanoma Experimental/imunologia , Melanoma Experimental/patologia , Camundongos Endogâmicos C57BL , Neoplasias/patologia , Nucleotídeos Cíclicos/administração & dosagem , Nucleotídeos Cíclicos/farmacologia , Receptor de Interferon alfa e beta/metabolismo , Transdução de Sinais/efeitos dos fármacosRESUMO
Adaptation to endoplasmic reticulum (ER) stress depends on the activation of an integrated signal transduction pathway known as the unfolded protein response (UPR). Bax inhibitor-1 (BI-1) is an evolutionarily conserved ER-resident protein that suppresses cell death. Here we have investigated the role of BI-1 in the UPR. BI-1 expression suppressed IRE1alpha activity in fly and mouse models of ER stress. BI-1-deficient cells displayed hyperactivation of the ER stress sensor IRE1alpha, leading to increased levels of its downstream target X-box-binding protein-1 (XBP-1) and upregulation of UPR target genes. This phenotype was associated with the formation of a stable protein complex between BI-1 and IRE1alpha, decreasing its ribonuclease activity. Finally, BI-1 deficiency increased the secretory activity of primary B cells, a phenomenon regulated by XBP-1. Our results suggest a role for BI-1 in early adaptive responses against ER stress that contrasts with its known downstream function in apoptosis.
Assuntos
Proteínas Reguladoras de Apoptose/metabolismo , Apoptose/fisiologia , Retículo Endoplasmático/fisiologia , Endorribonucleases/metabolismo , Proteínas de Membrana/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Sequência de Aminoácidos , Animais , Proteínas Reguladoras de Apoptose/genética , Linfócitos B/metabolismo , Células Cultivadas , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/metabolismo , Endorribonucleases/genética , Fibroblastos/citologia , Fibroblastos/metabolismo , Humanos , Imunoglobulina M/metabolismo , Proteínas de Membrana/genética , Camundongos , Camundongos Knockout , Dados de Sequência Molecular , Proteínas Serina-Treonina Quinases/genética , Splicing de RNA , Fatores de Transcrição de Fator Regulador X , Homologia de Sequência de Aminoácidos , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteína 1 de Ligação a X-BoxRESUMO
Gout is an arthritis characterized by elevated uric acid in the bloodstream. In this condition, crystals of uric acid are formed and accumulate in the synovial fluids. Crystal deposition leads to acute inflammation, which is associated with the spontaneous resolution of the disease. Recent studies have led to significant advances in the understanding of the basic biology of crystal-mediated inflammation. Uric acid has been identified as a danger signal that triggers a cytosolic sensor, the inflammasome. This signaling platform is required for the activation of interleukin-1, a cytokine that is critical to the initiation of acute inflammation in gout. Importantly, both molecular and pathological evidence support the notion that gout is a prototypical member of the growing family of autoinflammatory diseases. This review discusses the role of the inflammasome in gout and the emerging new therapeutic strategies aimed at controlling inflammation in crystal arthritis.
Assuntos
Artrite Gotosa/imunologia , Autoimunidade , Imunidade Inata , Mediadores da Inflamação/imunologia , Transdução de Sinais/imunologia , Ácido Úrico/imunologia , Animais , Artrite Gotosa/terapia , Cristalização , HumanosRESUMO
Gain-of-function mutations in NLRP3 are linked to cryopyrin-associated periodic syndromes (CAPS). Although NLRP3 autoinflammasome assembly triggers inflammatory cytokine release, its activation mechanisms are not fully understood. Our study used a functional genetic approach to identify regulators of NLRP3 inflammasome formation. We identified the HSP90ß-SGT1 chaperone complex as crucial for autoinflammasome activation in CAPS. A deficiency in HSP90ß, but not in HSP90α, impaired the formation of ASC specks without affecting the priming and expression of inflammasome components. Conversely, activating NLRP3 with stimuli such as nigericin or alum bypassed the need for SGT1 and HSP90ß, suggesting the existence of alternative inflammasome assembly pathways. The role of HSP90ß was further demonstrated in PBMCs derived from CAPS patients. In these samples, the pathological constitutive secretion of IL-1ß could be suppressed using a pharmacological inhibitor of HSP90ß. This finding underscores the potential of SGT1-HSP90ß modulation as a therapeutic strategy in CAPS while preserving NLRP3's physiological functions.