RESUMO
Although the function of the circulating immune cell compartment has been studied in detail for decades, limitations in terms of access and cell yields from peripheral tissues have restricted our understanding of tissue-based immunity, particularly in humans. Recent advances in high-throughput protein analyses, transcriptional profiling, and epigenetics have partially overcome these obstacles. Innate lymphoid cells (ILCs) are predominantly tissue-resident, and accumulating data indicate that they have significant tissue-specific functions. We summarize current knowledge of ILC phenotypes in various tissues in mice and humans, aiming to clarify ILC immunity in distinct anatomical locations.
Assuntos
Imunidade Inata , Linfócitos , Animais , Humanos , Imunidade Inata/imunologia , Linfócitos/citologia , Linfócitos/imunologiaRESUMO
In mast cells, IL-33 typically induces the activation of NF-κB, which results in the production of cytokines such as IL-6 and IL-2. Here, we demonstrate that the IL-33-induced IL-6 production in murine mast cells and the formation of RORγt+ Tregs essentially depends on the MAPKAPs, MK2, and MK3 (MK2/3) downstream of MyD88. In contrast to this, the IL-33-induced and MyD88-dependent IL-2 production in mast cells contributes to the maintenance of Helios+ Tregs . Thereby, the IL-33-induced IL-2 response and, thus, the maintenance of Helios+ Tregs are limited by an IL-6-mediated autocrine negative feedback stimulation acting on mast cells. Collectively, we present MK2/3 in IL-33-activated mast cells as a signaling node, which controls the dichotomy between RORγt+ Treg and Helios+ Treg in vitro.
Assuntos
Interleucina-33/imunologia , Interleucina-6/imunologia , Peptídeos e Proteínas de Sinalização Intracelular/imunologia , Sistema de Sinalização das MAP Quinases/imunologia , Mastócitos/imunologia , Proteínas Serina-Treonina Quinases/imunologia , Linfócitos T Reguladores/imunologia , Animais , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/imunologia , Interleucina-33/genética , Interleucina-6/genética , Peptídeos e Proteínas de Sinalização Intracelular/genética , Sistema de Sinalização das MAP Quinases/genética , Mastócitos/citologia , Camundongos , Camundongos Knockout , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/genética , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/imunologia , Proteínas Serina-Treonina Quinases/genética , Linfócitos T Reguladores/citologia , Fatores de Transcrição/genética , Fatores de Transcrição/imunologiaRESUMO
Constitutive NF-κB signaling represents a hallmark of chronic inflammation and autoimmune diseases. The E3 ligase TNF receptor-associated factor 6 (TRAF6) acts as a key regulator bridging innate immunity, pro-inflammatory cytokines, and antigen receptors to the canonical NF-κB pathway. Structural analysis and point mutations have unraveled the essential role of TRAF6 binding to the E2-conjugating enzyme ubiquitin-conjugating enzyme E2 N (Ubc13 or UBE2N) to generate Lys63-linked ubiquitin chains for inflammatory and immune signal propagation. Genetic mutations disrupting TRAF6-Ubc13 binding have been shown to reduce TRAF6 activity and, consequently, NF-κB activation. However, to date, no small-molecule modulator is available to inhibit the TRAF6-Ubc13 interaction and thereby counteract NF-κB signaling and associated diseases. Here, using a high-throughput small-molecule screening approach, we discovered an inhibitor of the TRAF6-Ubc13 interaction that reduces TRAF6-Ubc13 activity both in vitro and in cells. We found that this compound, C25-140, impedes NF-κB activation in various immune and inflammatory signaling pathways also in primary human and murine cells. Importantly, C25-140 ameliorated inflammation and improved disease outcomes of autoimmune psoriasis and rheumatoid arthritis in preclinical in vivo mouse models. Hence, the first-in-class TRAF6-Ubc13 inhibitor C25-140 expands the toolbox for studying the impact of the ubiquitin system on immune signaling and underscores the importance of TRAF6 E3 ligase activity in psoriasis and rheumatoid arthritis. We propose that inhibition of TRAF6 activity by small molecules represents a promising novel strategy for targeting autoimmune and chronic inflammatory diseases.
Assuntos
Artrite Reumatoide/tratamento farmacológico , Doenças Autoimunes/tratamento farmacológico , Inflamação/tratamento farmacológico , Psoríase/tratamento farmacológico , Bibliotecas de Moléculas Pequenas/farmacologia , Fator 6 Associado a Receptor de TNF/antagonistas & inibidores , Enzimas de Conjugação de Ubiquitina/antagonistas & inibidores , Animais , Artrite Reumatoide/metabolismo , Artrite Reumatoide/patologia , Doenças Autoimunes/metabolismo , Doenças Autoimunes/patologia , Células HEK293 , Ensaios de Triagem em Larga Escala , Humanos , Inflamação/metabolismo , Inflamação/patologia , Peptídeos e Proteínas de Sinalização Intracelular , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Mapas de Interação de Proteínas , Psoríase/metabolismo , Psoríase/patologia , Peptídeos e Proteínas Associados a Receptores de Fatores de Necrose Tumoral/antagonistas & inibidoresRESUMO
The IL-1R family member IL-33R mediates Fcε-receptor-I (FcεRI)-independent activation of mast cells leading to NF-κB activation and consequently the production of cytokines. IL-33 also induces the activation of MAPKs, such as p38. We aimed to define the relevance of the p38-targets, the MAPK-activated protein kinases 2 and 3 (MK2 and MK3) in IL-33-induced signaling and the resulting mast cell effector functions in vitro and in vivo. We demonstrate that the IL-33-induced IL-6 and IL-13 production strongly depends on the MK2/3-mediated activation of ERK1/2 and PI3K signaling. Furthermore, in the presence of the stem cell factors, IL-33 did induce an MK2/3-, ERK1/2- and PI3K-dependent production of TNF-α. In vivo, the loss of MK2/3 in mast cells decreased the IL-33-induced leukocyte recruitment and the resulting skin inflammation. Therefore, the MK2/3-dependent signaling in mast cells is essential to mediate IL-33-induced inflammatory responses. Thus, MK2/3 are potential therapeutic targets for suppression of IL-33-induced inflammation skin diseases such as psoriasis.
Assuntos
Inflamação/imunologia , Interleucina-33/imunologia , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Leucócitos/imunologia , Mastócitos/imunologia , Proteínas Serina-Treonina Quinases/metabolismo , Psoríase/imunologia , Pele/imunologia , Animais , Movimento Celular , Células Cultivadas , Mediadores da Inflamação/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/genética , Sistema de Sinalização das MAP Quinases , Camundongos , Camundongos Knockout , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Serina-Treonina Quinases/genéticaRESUMO
The CARMA1-BCL10-MALT1 (CBM) signalosome triggers canonical NF-κB signaling and lymphocyte activation upon antigen-receptor stimulation. Genetic studies in mice and the analysis of human immune pathologies unveiled a critical role of the CBM complex in adaptive immune responses. Great progress has been made in elucidating the fundamental mechanisms that dictate CBM assembly and disassembly. By bridging proximal antigen-receptor signaling to downstream signaling pathways, the CBM complex exerts a crucial scaffolding function. Moreover, the MALT1 subunit confers a unique proteolytic activity that is key for lymphocyte activation. Deregulated 'chronic' CBM signaling drives constitutive NF-κB signaling and MALT1 activation, which contribute to the development of autoimmune and inflammatory diseases as well as lymphomagenesis. Thus, the processes that govern CBM activation and function are promising targets for the treatment of immune disorders. Here, we summarize the current knowledge on the functions and mechanisms of CBM signaling in lymphocytes and how CBM deregulations contribute to aberrant signaling in malignant lymphomas.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Caspases/metabolismo , Guanilato Ciclase/metabolismo , Ativação Linfocitária , Linfócitos/citologia , Linfócitos/imunologia , Transdução de Sinais , Proteínas Adaptadoras de Transdução de Sinal/química , Animais , Caspases/química , Guanilato Ciclase/química , HumanosRESUMO
Alternative splicing plays key roles for cell type-specific regulation of protein function. It is controlled by cis-regulatory RNA elements that are recognized by RNA binding proteins (RBPs). The MALT1 paracaspase is a key factor of signaling pathways that mediate innate and adaptive immune responses. Alternative splicing of MALT1 is critical for controlling optimal T cell activation. We demonstrate that MALT1 splicing depends on RNA structural elements that sequester the splice sites of the alternatively spliced exon7. The RBPs hnRNP U and hnRNP L bind competitively to stem-loop RNA structures that involve the 5' and 3' splice sites flanking exon7. While hnRNP U stabilizes RNA stem-loop conformations that maintain exon7 skipping, hnRNP L disrupts these RNA elements to facilitate recruitment of the essential splicing factor U2AF2, thereby promoting exon7 inclusion. Our data represent a paradigm for the control of splice site selection by differential RBP binding and modulation of pre-mRNA structure.
Assuntos
Ribonucleoproteínas Nucleares Heterogêneas Grupo L , Precursores de RNA , Processamento Alternativo , Sítios de Ligação , Éxons , Ribonucleoproteínas Nucleares Heterogêneas Grupo L/genética , Ribonucleoproteínas Nucleares Heterogêneas Grupo L/metabolismo , Ribonucleoproteínas Nucleares Heterogêneas Grupo U/genética , Ribonucleoproteínas Nucleares Heterogêneas Grupo U/metabolismo , Ribonucleoproteínas Nucleares Heterogêneas/genética , Ribonucleoproteínas Nucleares Heterogêneas/metabolismo , Proteína de Translocação 1 do Linfoma de Tecido Linfoide Associado à Mucosa/genética , Proteína de Translocação 1 do Linfoma de Tecido Linfoide Associado à Mucosa/metabolismo , Precursores de RNA/genética , Sítios de Splice de RNA , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismoRESUMO
The CARMA1/CARD11-BCL10-MALT1 (CBM) complex bridges T and B cell antigen receptor (TCR/BCR) ligation to MALT1 protease activation and canonical nuclear factor κB (NF-κB) signaling. Using unbiased mass spectrometry, we discover multiple serine phosphorylation sites in the MALT1 C terminus after T cell activation. Phospho-specific antibodies reveal that CBM-associated MALT1 is transiently hyper-phosphorylated upon TCR/CD28 co-stimulation. We identify a dual role for CK1α as a kinase that is essential for CBM signalosome assembly as well as MALT1 phosphorylation. Although MALT1 phosphorylation is largely dispensable for protease activity, it fosters canonical NF-κB signaling in Jurkat and murine CD4 T cells. Moreover, constitutive MALT1 phosphorylation promotes survival of activated B cell-type diffuse large B cell lymphoma (ABC-DLBCL) cells addicted to chronic BCR signaling. Thus, MALT1 phosphorylation triggers optimal NF-κB activation in lymphocytes and survival of lymphoma cells.
Assuntos
Ativação Linfocitária , Linfoma Difuso de Grandes Células B/imunologia , Proteína de Translocação 1 do Linfoma de Tecido Linfoide Associado à Mucosa/metabolismo , Transdução de Sinais , Linfócitos T/imunologia , Motivos de Aminoácidos , Animais , Proteína 10 de Linfoma CCL de Células B/metabolismo , Proteínas Adaptadoras de Sinalização CARD/metabolismo , Antígenos CD28/metabolismo , Caseína Quinase Ialfa/metabolismo , Células Cultivadas , Guanilato Ciclase/metabolismo , Células HEK293 , Humanos , Células Jurkat , Camundongos , Camundongos Endogâmicos C57BL , Proteína de Translocação 1 do Linfoma de Tecido Linfoide Associado à Mucosa/química , NF-kappa B/metabolismo , FosforilaçãoRESUMO
This research explores the first design and synthesis of macrocyclic peptide aldehydes as potent inhibitors of the 20S proteasome. Two novel macrocyclic peptide aldehydes based on the ring-size of the macrocyclic natural product TMC-95 were prepared and evaluated as inhibitors of the 20S proteasome. Both compounds inhibited in the low nanomolar range and proved to be selective for the proteasome over other serine and cysteine proteases, particularly when compared to linear analogues with similar amino acid sequences. In HeLa cells, both macrocycles efficiently inhibited activation of nuclear factor-κB (NF-κB) transcription factor by blocking proteasomal degradation of the inhibitor protein IκBα after cytokine stimulation. Due to their covalent mechanism of binding these compounds represent a 1000-fold increase in inhibitory potency over previously reported noncovalently binding TMC-95 analogues. Molecular modeling of the macrocyclic peptides confirms the preference of the large S3 pocket for large, hydrophobic residues and the ability to exploit this to improve selectivity of proteasome inhibitors.
RESUMO
MALT1 channels proximal T-cell receptor (TCR) signalling to downstream signalling pathways. With MALT1A and MALT1B two conserved splice variants exist and we demonstrate here that MALT1 alternative splicing supports optimal T-cell activation. Inclusion of exon7 in MALT1A facilitates the recruitment of TRAF6, which augments MALT1 scaffolding function, but not protease activity. Naive CD4(+) T cells express almost exclusively MALT1B and MALT1A expression is induced by TCR stimulation. We identify hnRNP U as a suppressor of exon7 inclusion. Whereas selective depletion of MALT1A impairs T-cell signalling and activation, downregulation of hnRNP U enhances MALT1A expression and T-cell activation. Thus, TCR-induced alternative splicing augments MALT1 scaffolding to enhance downstream signalling and to promote optimal T-cell activation.
Assuntos
Processamento Alternativo/genética , Linfócitos T CD4-Positivos/imunologia , Caspases/genética , Ativação Linfocitária/imunologia , Proteínas de Neoplasias/genética , Transdução de Sinais , Animais , Caspases/metabolismo , Regulação para Baixo , Ativação Enzimática , Éxons/genética , Células HEK293 , Ribonucleoproteínas Nucleares Heterogêneas Grupo U/metabolismo , Humanos , Interleucina-2/biossíntese , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Células Jurkat , Camundongos Endogâmicos C57BL , Proteína de Translocação 1 do Linfoma de Tecido Linfoide Associado à Mucosa , NF-kappa B/metabolismo , Proteínas de Neoplasias/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Fator 6 Associado a Receptor de TNF/metabolismo , Células Th17/imunologia , Regulação para CimaRESUMO
Mast cell differentiation and proliferation depends on IL-3. IL-3 induces the activation of MAP-kinases and STATs and consequently induces proliferation and survival. Dysregulation of IL-3 signaling pathways also contribute to inflammation and tumorigenesis. We show here that IL-3 induces a SFK- and Ca²âº-dependent activation of the inhibitor of κB kinases 2 (IKK2) which results in mast cell proliferation and survival but does not induce IκBα-degradation and NFκB activation. Therefore we propose the term "subthreshold IKK activation".This subthreshold IKK activation also primes mast cells for enhanced responsiveness to IL-33R signaling. Consequently, co-stimulation with IL-3 and IL-33 increases IKK activation and massively enhances cytokine production induced by IL-33.We further reveal that in neoplastic mast cells expressing constitutively active Ras, subthreshold IKK activation is associated with uncontrolled proliferation. Consequently, pharmacological IKK inhibition reduces tumor growth selectively by inducing apoptosis in vivo.Together, subthreshold IKK activation is crucial to mediate the full IL-33-induced effector functions in primary mast cells and to mediate uncontrolled proliferation of neoplastic mast cells. Thus, IKK2 is a new molecularly defined target structure.