RESUMO
The a disintegrin and metalloproteinase (ADAM) family of proteinases alter the extracellular environment and are involved in the development of T cells and autoimmunity. The role of ADAM family members in Th17 cell differentiation is unknown. We identified ADAM9 to be specifically expressed and to promote Th17 differentiation. Mechanistically, we found that ADAM9 cleaved the latency-associated peptide to produce bioactive transforming growth factor ß1, which promoted SMAD2/3 phosphorylation and activation. A transcription factor inducible cAMP early repressor was found to bind directly to the ADAM9 promoter and to promote its transcription. Adam9-deficient mice displayed mitigated experimental autoimmune encephalomyelitis, and transfer of Adam9-deficient myelin oligodendrocyte globulin-specific T cells into Rag1-/- mice failed to induce disease. At the translational level, an increased abundance of ADAM9 levels was observed in CD4+ T cells from patients with systemic lupus erythematosus, and ADAM9 gene deletion in lupus primary CD4+ T cells clearly attenuated their ability to differentiate into Th17 cells. These findings revealed that ADAM9 as a proteinase provides Th17 cells with an ability to activate transforming growth factor ß1 and accelerates its differentiation, resulting in aberrant autoimmunity.
Assuntos
Proteínas ADAM/genética , Autoimunidade/genética , Proteínas de Homeodomínio/genética , Proteínas de Membrana/genética , Linfócitos T/imunologia , Fator de Crescimento Transformador beta1/genética , Adulto , Animais , Autoimunidade/imunologia , Linfócitos T CD4-Positivos/imunologia , Diferenciação Celular/genética , AMP Cíclico/genética , Feminino , Humanos , Lúpus Eritematoso Sistêmico , Masculino , Camundongos , Pessoa de Meia-Idade , Bainha de Mielina/genética , Oligodendroglia/metabolismo , Fosforilação/genética , Proteína Smad2/genética , Proteína Smad3/genética , Linfócitos T/patologia , Células Th17/imunologia , Adulto JovemRESUMO
OBJECTIVE: Glutaminase (GLS) isoenzymes GLS1 and GLS2 catalyze the first step of glutaminolysis. GLS1 is requisite for Th17 cell differentiation, and its inhibition suppresses autoimmune disease in animals, but the function of GLS2 is not known. The aim of this study was to investigate the role of GLS2 in CD4+ T cell function and systemic lupus erythematosus (SLE) pathogenesis. METHODS: We measured reactive oxygen species (ROS) levels, lipid peroxidation, and mitochondrial mass and polarization by flow cytometry, interleukin-2 (IL-2) production by a dual luciferase assay, and CpG DNA methylation of Il2 by a real-time polymerase chain reaction system. The impact of the overexpression of wild-type GLS1, wild-type GLS2, or mutated GLS2 at the PDZ domain-binding motif in CD4+ T cells was examined. Furthermore, GLS2 expression in CD4+ T cells from lupus-prone mice and patients with SLE was analyzed by Western blotting. RESULTS: GLS2, but not GLS1, reduced ROS levels and lipid peroxidation and restored mitochondrial function in T cells. GLS2 promoted IL-2 production through the demethylation of the Il2 promoter. Mutation of the PDZ domain-binding motif abated the ability of GLS2 to regulate IL-2 and ROS levels. In lupus-prone mice and patients with SLE, the expression of GLS2 was decreased in CD4+ T cells. Finally, GLS2 overexpression corrected ROS levels and restored IL-2 production by CD4+ T cells from lupus-prone mice and SLE patients. CONCLUSION: Our findings suggest that GLS2 has a crucial role in IL-2 production by CD4+ T cells by supporting antioxidant defense, and they offer a new approach to correcting IL-2 production by T cells in SLE.
Assuntos
Antioxidantes , Linfócitos T CD4-Positivos , Glutaminase , Interleucina-2 , Lúpus Eritematoso Sistêmico , Animais , Antioxidantes/metabolismo , Linfócitos T CD4-Positivos/metabolismo , Glutaminase/metabolismo , Interleucina-2/metabolismo , Lúpus Eritematoso Sistêmico/imunologia , Camundongos , Espécies Reativas de Oxigênio/metabolismoRESUMO
Th1 and Th17 are important in the pathogenesis of autoimmune diseases and they depend on glycolysis as a source of energy. T cell antigen receptor signaling phosphorylates a serine/threonine kinase, calcium/calmodulin-dependent protein kinase IV (CaMK4), and promotes glycolysis. Based on these findings we hypothesized that CaMK4 promotes glycolysis. Camk4-deficient CD4+ T cells and cells treated with a CaMK4 inhibitor had less glycolysis compared with their counterparts. Pull-down of CaMK4 and mass spectrometry identified pyruvate kinase muscle isozyme (PKM), the final rate-limiting enzyme in glycolysis, as a binding partner. Coimmunoprecipitation and Western blotting showed that CaMK4 interacts directly with PKM2. Camk4-deficient CD4+ T cells displayed decreased pyruvate kinase activity. Silencing or pharmacological inhibition of PKM2 reduced glycolysis and in vitro differentiation to Th1 and Th17 cells, while PKM2 overexpression restored Th17 cell differentiation. Treatment with a PKM2 inhibitor ameliorated experimental autoimmune encephalomyelitis and CD4+ T cells treated with PKM2 inhibitor or Pkm2-shRNA caused limited disease activity in an adoptive cell transfer model of experimental autoimmune encephalomyelitis. Our data demonstrate that CaMK4 binds to PKM2 and promotes its activity, which is requisite for Th1 and Th17 differentiation in vitro and in vivo. PKM2 represents a therapeutic target for T cell-dependent autoimmune diseases.