RESUMO
Phosphatidylinositol-3-kinase gamma (PI3Kγ) is a leukocyte-specific lipid kinase with signaling function downstream of G protein-coupled receptors to regulate cell trafficking, but its role in T cells remains unclear. To investigate the requirement of PI3Kγ kinase activity in T-cell function, we studied T cells from PI3Kγ kinase-dead knock-in (PI3Kγ(KD/KD)) mice expressing the kinase-inactive PI3Kγ protein. We show that CD4(+) and CD8(+) T cells from PI3Kγ(KD/KD) mice exhibit impaired TCR/CD28-mediated activation that could not be rescued by exogenous IL-2. The defects in proliferation and cytokine production were also evident in naïve and memory T cells. Analysis of signaling events in activated PI3Kγ(KD/KD) T cells revealed a reduction in phosphorylation of protein kinase B (AKT) and ERK1/2, a decrease in lipid raft formation, and a delay in cell cycle progression. Furthermore, PI3Kγ(KD/KD) CD4(+) T cells displayed compromised differentiation toward Th1, Th2, Th17, and induced Treg cells. PI3Kγ(KD/KD) mice also exhibited an impaired response to immunization and a reduced delayed-type hypersensitivity to Ag challenge. These findings indicate that PI3Kγ kinase activity is required for optimal T-cell activation and differentiation, as well as for mounting an efficient T cell-mediated immune response. The results suggest that PI3Kγ kinase inhibitors could be beneficial in reducing the undesirable immune response in autoimmune diseases.
Assuntos
Diferenciação Celular/imunologia , Classe Ib de Fosfatidilinositol 3-Quinase/imunologia , Ativação Linfocitária/imunologia , Linfócitos T Auxiliares-Indutores/imunologia , Animais , Antígenos/imunologia , Doenças Autoimunes/tratamento farmacológico , Doenças Autoimunes/genética , Doenças Autoimunes/imunologia , Doenças Autoimunes/patologia , Diferenciação Celular/genética , Classe Ib de Fosfatidilinositol 3-Quinase/genética , Inibidores Enzimáticos/uso terapêutico , Hipersensibilidade Tardia/genética , Hipersensibilidade Tardia/imunologia , Hipersensibilidade Tardia/patologia , Ativação Linfocitária/genética , Microdomínios da Membrana/genética , Microdomínios da Membrana/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Transgênicos , Proteína Quinase 3 Ativada por Mitógeno/genética , Proteína Quinase 3 Ativada por Mitógeno/imunologia , Inibidores de Fosfoinositídeo-3 Quinase , Fosforilação/genética , Fosforilação/imunologia , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/imunologiaRESUMO
The innate immune system is equipped with sensitive and efficient machineries to provide an immediate, first line defense against infections. Toll-like receptors (TLRs) detect pathogens and the IL-1 receptor (IL-1R) family enables cells to quickly respond to inflammatory cytokines by mounting an efficient protective response. Interleukin-1 receptor activated kinases (IRAKs) are key mediators in the signaling pathways of TLRs/IL-1Rs. By means of their kinase and adaptor functions, IRAKs initiate a cascade of signaling events eventually leading to induction of inflammatory target gene expression. Due to this pivotal role, IRAK function is also highly regulated via multiple mechanisms. In this review, we focus on IRAK1, the earliest known and yet the most interesting member of this family. An overview on its structure, function and biology is given, with emphasis on the different novel mechanisms that regulate IRAK1 function. We also highlight several unresolved questions in this field and evaluate the potential of IRAK1 as a target for therapeutic intervention.
Assuntos
Imunidade Inata/imunologia , Quinases Associadas a Receptores de Interleucina-1/metabolismo , Transdução de Sinais , Animais , Doenças Autoimunes/enzimologia , Humanos , Quinases Associadas a Receptores de Interleucina-1/química , Quinases Associadas a Receptores de Interleucina-1/genética , Receptores Imunológicos/imunologia , UbiquitinaçãoRESUMO
Notch1 specifically upregulates expression of the cytokine interferon-gamma in peripheral T cells through activation of NF-kappaB. However, how Notch mediates NF-kappaB activation remains unclear. Here, we examined the temporal relationship between Notch signaling and NF-kappaB induction during T-cell activation. NF-kappaB activation occurs within minutes of T-cell receptor (TCR) engagement and this activation is sustained for at least 48 h following TCR signaling. We used gamma-secretase inhibitor (GSI) to prevent the cleavage and subsequent activation of Notch family members. We demonstrate that GSI blocked the later, sustained NF-kappaB activation, but did not affect the initial activation of NF-kappaB. Using biochemical approaches, as well as confocal microscopy, we show that the intracellular domain of Notch1 (N1IC) directly interacts with NF-kappaB and competes with IkappaBalpha, leading to retention of NF-kappaB in the nucleus. Additionally, we show that N1IC can directly regulate IFN-gamma expression through complexes formed on the IFN-gamma promoter. Taken together, these data suggest that there are two 'waves' of NF-kappaB activation: an initial, Notch-independent phase, and a later, sustained activation of NF-kappaB, which is Notch dependent.
Assuntos
Núcleo Celular/metabolismo , Regulação da Expressão Gênica , Proteínas I-kappa B/metabolismo , Interferon gama/genética , NF-kappa B/metabolismo , Receptor Notch1/metabolismo , Animais , Anticorpos/farmacologia , Antígenos CD28/imunologia , Complexo CD3/imunologia , Núcleo Celular/química , Citosol/química , Citosol/metabolismo , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos C57BL , Inibidor de NF-kappaB alfa , NF-kappa B/análise , Subunidade p50 de NF-kappa B/genética , Regiões Promotoras Genéticas , Proteínas Proto-Oncogênicas c-rel/genética , Transdução de Sinais , Baço/citologia , Baço/efeitos dos fármacos , Baço/imunologia , Linfócitos T/imunologiaRESUMO
Notch receptors are processed by gamma-secretase acting in synergy with T cell receptor signaling to sustain peripheral T cell activation. Activated CD4+ T cells differentiate into T helper type 1 (TH1) or TH2 subsets. Molecular cues directing TH1 differentiation include expression of the TH1-specific transcription factor T-bet, encoded by Tbx21. However, the regulation of Tbx21 remains incompletely defined. Here we report that Notch1 can directly regulate Tbx21 through complexes formed on the Tbx21 promoter. In vitro, gamma-secretase inhibitors extinguished expression of Notch, interferon-gamma and Tbx21 in TH1-polarized CD4+ cells, whereas ectopic expression of activated Notch1 restored Tbx21 transcription. In vivo, administration of gamma-secretase inhibitors substantially impeded TH1-mediated disease progression in the mouse experimental autoimmune encephalomyelitis model of multiple sclerosis. Thus, using gamma-secretase inhibitors to modulate Notch signaling may prove beneficial in treating TH1-mediated autoimmunity.
Assuntos
Proteínas de Ligação a DNA/antagonistas & inibidores , Endopeptidases/imunologia , Inibidores de Proteases/farmacologia , Receptores de Superfície Celular/antagonistas & inibidores , Células Th1/imunologia , Fatores de Transcrição/antagonistas & inibidores , Secretases da Proteína Precursora do Amiloide , Animais , Ácido Aspártico Endopeptidases , Citocinas/imunologia , Proteínas de Ligação a DNA/biossíntese , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/imunologia , Encefalomielite Autoimune Experimental/tratamento farmacológico , Encefalomielite Autoimune Experimental/enzimologia , Encefalomielite Autoimune Experimental/imunologia , Ensaio de Imunoadsorção Enzimática , Feminino , Hipersensibilidade Tardia/tratamento farmacológico , Hipersensibilidade Tardia/imunologia , Immunoblotting , Camundongos , Camundongos Endogâmicos C57BL , Receptor Notch1 , Receptores de Superfície Celular/imunologia , Proteínas com Domínio T , Células Th1/efeitos dos fármacos , Células Th1/enzimologia , Fatores de Transcrição/biossíntese , Fatores de Transcrição/genética , Fatores de Transcrição/imunologia , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/imunologiaRESUMO
Truncated Notch receptors have transforming activity in vitro and in vivo. However, the role of wild-type Notch signaling in neoplastic transformation remains unclear. Ras signaling is deregulated in a large fraction of human malignancies and is a major target for the development of novel cancer treatments. We show that oncogenic Ras activates Notch signaling and that wild-type Notch-1 is necessary to maintain the neoplastic phenotype in Ras-transformed human cells in vitro and in vivo. Oncogenic Ras increases levels and activity of the intracellular form of wild-type Notch-1, and upregulates Notch ligand Delta-1 and also presenilin-1, a protein involved in Notch processing, through a p38-mediated pathway. These observations place Notch signaling among key downstream effectors of oncogenic Ras and suggest that it might be a novel therapeutic target.