ABSTRACT
Regulatory T (Treg) cells maintain immune homeostasis by preventing abnormal or excessive immune responses. Histone deacetylase 6 (HDAC6) regulates expression of Foxp3, and thus, Treg cell differentiation; however, its role in Treg cell differentiation is unclear and somewhat controversial. Here, we investigated the role of HDAC6 in TGF-ß-induced murine Treg cells. HDAC6 expression was higher in Treg cells than in other T helper cell subsets. Pharmacological inhibitors of HDAC6 selectively inhibited Treg cell differentiation and suppressive function. A specific HDAC6 inhibitor induced changes in global gene expression by Treg cells. Of these changes, genes related to cell division were prominently affected. In summary, HDAC6 plays an important role in TGF-ß-induced murine Treg cell differentiation by regulating cell proliferation.
Subject(s)
Histone Deacetylase 6 , T-Lymphocytes, Regulatory , Transforming Growth Factor beta , Animals , Mice , Cell Differentiation , Cell Proliferation , Histone Deacetylase 6/metabolism , Transforming Growth Factor beta/metabolismABSTRACT
Regulatory T (Treg) cells play an essential role in maintaining immune homeostasis, but the suppressive function of Treg cells can be an obstacle in the treatment of cancer and chronic infectious diseases. Here, we identified the homeobox protein Hhex as a negative regulator of Treg cells. The expression of Hhex was lower in Treg cells than in conventional T (Tconv) cells. Hhex expression was repressed in Treg cells by TGF-ß/Smad3 signaling. Retroviral overexpression of Hhex inhibited the differentiation of induced Treg (iTreg) cells and the stability of thymic Treg (tTreg) cells by significantly reducing Foxp3 expression. Moreover, Hhex-overexpressing Treg cells lost their immunosuppressive activity and failed to prevent colitis in a mouse model of inflammatory bowel disease (IBD). Hhex expression was increased; however, Foxp3 expression was decreased in Treg cells in a delayed-type hypersensitivity (DTH) reaction, a type I immune reaction. Hhex directly bound to the promoters of Foxp3 and other Treg signature genes, including Il2ra and Ctla4, and repressed their transactivation. The homeodomain and N-terminal repression domain of Hhex were critical for inhibiting Foxp3 and other Treg signature genes. Thus, Hhex plays an essential role in inhibiting Treg cell differentiation and function via inhibition of Foxp3.
Subject(s)
Forkhead Transcription Factors/metabolism , Homeodomain Proteins/metabolism , T-Lymphocytes, Regulatory/immunology , T-Lymphocytes, Regulatory/metabolism , Transcription Factors/metabolism , Animals , CTLA-4 Antigen/metabolism , Cell Differentiation , Colitis/metabolism , Colitis/pathology , Disease Models, Animal , Forkhead Transcription Factors/genetics , Gene Expression Regulation , Homeodomain Proteins/genetics , Inflammatory Bowel Diseases/genetics , Inflammatory Bowel Diseases/metabolism , Inflammatory Bowel Diseases/pathology , Interleukin-2 Receptor alpha Subunit/metabolism , Mice , Signal Transduction , Skin/pathology , Smad3 Protein/metabolism , Transcription Factors/genetics , Transforming Growth Factor beta/metabolismABSTRACT
Th9 cells preferentially produce IL-9 and participate in allergic responses and asthma. Differentiation of Th9 cells is induced by IL-4 and TGF-ß, and then the cells are amplified by OX40 signals. The transcription factors PU.1, IRF4, and BATF are required for Th9 differentiation. BATF3 is an AP-1 family transcription factor that is highly homologous to BATF; however, its role in Th9 cells is poorly defined. Here, we show that OX40 signaling induced the expression of Batf3 and that its overexpression in the presence or absence of OX40 signaling increased the expression of IL-9 in Th9 cells. BATF3 physically interacted with IRF4 and was bound to the Il9 locus. A transient reporter assay revealed that the BATF3-IRF4 complex induced Il9 promoter activity. BATF3 rescued Il9 expression and restored the capacity to induce the airway inflammation in Batf KO Th9 cells. Thus, BATF3 itself is sufficient for the induction of Th9 cell differentiation and can substitute for BATF during Th9 cell differentiation.
Subject(s)
Basic-Leucine Zipper Transcription Factors/metabolism , CD4-Positive T-Lymphocytes/metabolism , Interleukin-9/metabolism , Repressor Proteins/metabolism , Animals , Basic-Leucine Zipper Transcription Factors/genetics , Cell Differentiation/genetics , Cell Differentiation/physiology , Chromatin Immunoprecipitation , Enzyme-Linked Immunosorbent Assay , Immunoblotting , Interferon Regulatory Factors/genetics , Interferon Regulatory Factors/metabolism , Lung , Mice , Mice, Inbred C57BL , Mice, Knockout , Proto-Oncogene Proteins/genetics , Proto-Oncogene Proteins/metabolism , Repressor Proteins/genetics , Reverse Transcriptase Polymerase Chain Reaction , Trans-Activators/genetics , Trans-Activators/metabolismABSTRACT
Regulatory T (Treg) cells are a CD4 T-cell subset with an important role in immune tolerance; however, the mechanisms underlying Treg cell differentiation and function are incompletely understood. Here, we show that NFIL3/E4BP4, a transcription factor, plays a key role in Treg cell differentiation and function. Microarray analysis showed that Treg cells had lower Nfil3 expression than all other CD4 T-cell subsets. Overexpression of Nfil3 in Treg cells led to diminished expression of Foxp3 and other signature Treg genes, including Il2ra, Icos, Tnfrsf18, and Ctla4. Furthermore, Nfil3-overexpressing Treg cells exhibited impaired immunosuppressive activity in vitro and in vivo. We discovered that NFIL3 directly binds to and negatively regulates the expression of Foxp3. In addition, bisulfite sequencing revealed that NFIL3 induces methylation at Foxp3 locus regulatory CpG sites, which contributes to the control of Treg cell stability. Together, these data indicate that NFIL3 impairs Treg cell function through the downregulation of Foxp3 expression.
Subject(s)
Basic-Leucine Zipper Transcription Factors/metabolism , Forkhead Transcription Factors/metabolism , Gene Expression Regulation , Immune Tolerance , T-Lymphocytes, Regulatory/metabolism , Animals , Basic-Leucine Zipper Transcription Factors/genetics , Cell Differentiation , DNA Methylation , Down-Regulation , Female , Forkhead Transcription Factors/genetics , Gene Expression , Genes, Reporter , HEK293 Cells , Humans , Mice , Promoter Regions, Genetic/genetics , T-Lymphocytes, Regulatory/immunologyABSTRACT
T helper 17 (Th17) cells are a CD4+ T cell subset that produces IL-17A to mediate inflammation and autoimmunity. IL-2 inhibits Th17 cell differentiation. However, the mechanism by which IL-2 is suppressed during Th17 cell differentiation remains unclear. Here, we show that phosphatase and tensin homologue (PTEN) is a key factor that regulates Th17 cell differentiation by suppressing IL-2 production. Th17-specific Pten deletion (Ptenfl/flIl17acre ) impairs Th17 cell differentiation in vitro and ameliorated symptoms of experimental autoimmune encephalomyelitis (EAE), a model of Th17-mediated autoimmune disease. Mechanistically, Pten deficiency up-regulates IL-2 and phosphorylation of STAT5, but reduces STAT3 phosphorylation, thereby inhibiting Th17 cell differentiation. PTEN inhibitors block Th17 cell differentiation in vitro and in the EAE model. Thus, PTEN plays a key role in Th17 cell differentiation by blocking IL-2 expression.
Subject(s)
Cell Differentiation/immunology , Interleukin-2/immunology , PTEN Phosphohydrolase/immunology , Th17 Cells/immunology , Animals , Cell Differentiation/genetics , Cells, Cultured , Encephalomyelitis, Autoimmune, Experimental/genetics , Encephalomyelitis, Autoimmune, Experimental/immunology , Encephalomyelitis, Autoimmune, Experimental/metabolism , Gene Expression Profiling/methods , Interleukin-2/genetics , Interleukin-2/metabolism , Mice, Inbred C57BL , Mice, Knockout , PTEN Phosphohydrolase/genetics , PTEN Phosphohydrolase/metabolism , Phosphorylation , RNA Interference , STAT3 Transcription Factor/metabolism , STAT5 Transcription Factor/metabolism , Th17 Cells/metabolism , Up-RegulationABSTRACT
Th17 cells promote inflammatory reactions, whereas regulatory T (Treg) cells inhibit them. Thus, the Th17/Treg cell balance is critically important in inflammatory diseases. However, the molecular mechanisms underlying this balance are unclear. Here, we demonstrate that casein kinase 2 (CK2) is a critical determinant of the Th17/Treg cell balance. Both the inhibition of CK2 with a specific pharmacological inhibitor, CX-4945, and its small hairpin RNA (shRNA)-mediated knockdown suppressed Th17 cell differentiation but reciprocally induced Treg cell differentiation in vitro. Moreover, CX-4945 ameliorated the symptoms of experimental autoimmune encephalomyelitis and reduced Th17 cell infiltration into the central nervous system. Mechanistically, CX-4945 inhibited the IL-6/STAT3 and Akt/mTOR signaling pathways. Thus, CK2 has a crucial role in regulating the Th17/Treg balance.
Subject(s)
Casein Kinase II/metabolism , Cell Differentiation , T-Lymphocytes, Regulatory/cytology , T-Lymphocytes, Regulatory/metabolism , Th17 Cells/cytology , Th17 Cells/metabolism , Animals , Casein Kinase II/antagonists & inhibitors , Casein Kinase II/genetics , Cell Differentiation/genetics , Cell Differentiation/immunology , Encephalomyelitis, Autoimmune, Experimental , Female , Gene Expression , Gene Knockdown Techniques , Lymphocyte Activation/genetics , Lymphocyte Activation/immunology , Lymphocyte Count , Mice , Phosphorylation , Proto-Oncogene Proteins c-akt/metabolism , RNA Interference , Receptors, Interleukin-6/metabolism , STAT3 Transcription Factor/metabolism , Signal Transduction , T-Lymphocyte Subsets/cytology , T-Lymphocyte Subsets/immunology , T-Lymphocyte Subsets/metabolism , T-Lymphocytes, Regulatory/immunology , TOR Serine-Threonine Kinases , Th17 Cells/immunologyABSTRACT
Subset-specific gene expression is a critical feature of CD4 T cell differentiation. Th2 cells express Th2 cytokine genes including Il4, Il5, and Il13 and mediate the immune response against helminths. The expression of Th2 cytokine genes is regulated by Rad50 hypersensitive site 6 (RHS6) in the Th2 locus control region; however, the molecular mechanisms of RHS6 action at the chromatin level are poorly understood. Here, we demonstrate that RHS6 is crucial for chromosomal interactions and nuclear substructure binding of the Th2 cytokine locus. RHS6-deficient cells had a marked reduction in chromatin remodeling and in intrachromosomal interactions at the Th2 locus. Deficiency of RHS6-binding transcription factors GATA3, SATB1, and IRF4 also caused a great reduction in chromatin remodeling and long-range chromosomal interactions involving the Th2 locus. RHS6 deficiency abrogated association of the Th2 locus with the nuclear substructure and RNA polymerase II. Therefore, RHS6 serves as a crucial cis-acting hub for coordinate regulation of Th2 cytokine genes by forming chromosomal loops and binding to a nuclear substructure.
Subject(s)
Chromatin Assembly and Disassembly/immunology , Chromosomes, Mammalian/immunology , Cytokines/immunology , Locus Control Region/immunology , Th2 Cells/immunology , ATP-Binding Cassette Transporters/genetics , ATP-Binding Cassette Transporters/immunology , Acid Anhydride Hydrolases , Animals , Chromatin Assembly and Disassembly/genetics , Chromosomes, Mammalian/genetics , Cytokines/genetics , DNA-Binding Proteins , Mice , Mice, Transgenic , Th2 Cells/cytology , Transcription Factors/genetics , Transcription Factors/immunologyABSTRACT
Regulatory T (T(reg)) cells are essential for maintenance of immune homeostasis. Foxp3 is the key transcription factor for T(reg)-cell differentiation and function; however, molecular mechanisms for its negative regulation are poorly understood. Here we show that YY1 expression is lower in T(reg) cells than T(conv) cells, and its overexpression causes a marked reduction of Foxp3 expression and abrogation of suppressive function of Treg cells. YY1 is increased in T(reg) cells under inflammatory conditions with concomitant decrease of suppressor activity in dextran sulfate-induced colitis model. YY1 inhibits Smad3/4 binding to and chromatin remodelling of the Foxp3 locus. In addition, YY1 interrupts Foxp3-dependent target gene expression by physically interacting with Foxp3 and by directly binding to the Foxp3 target genes. Thus, YY1 inhibits differentiation and function of T(reg) cells by blocking Foxp3.
Subject(s)
Cell Differentiation , Colitis/metabolism , Forkhead Transcription Factors/metabolism , T-Lymphocytes, Regulatory/cytology , YY1 Transcription Factor/metabolism , Animals , Colitis/genetics , Female , Forkhead Transcription Factors/genetics , Humans , Male , Mice, Inbred C57BL , Mice, Knockout , Protein Binding , T-Lymphocytes, Regulatory/metabolism , YY1 Transcription Factor/geneticsABSTRACT
The Th2 locus control region (LCR) has been shown to be important in efficient and coordinated cytokine gene regulation during Th2 cell differentiation. However, the molecular mechanism for this is poorly understood. To study the molecular mechanism of the Th2 LCR, we searched for proteins binding to it. We discovered that transcription factor YY1 bound to the LCR and the entire Th2 cytokine locus in a Th2-specific manner. Retroviral overexpression of YY1 induced Th2 cytokine expression. CD4-specific knockdown of YY1 in mice caused marked reduction in Th2 cytokine expression, repressed chromatin remodeling, decreased intrachromosomal interactions, and resistance in an animal model of asthma. YY1 physically associated with GATA-binding protein-3 (GATA3) and is required for GATA3 binding to the locus. YY1 bound to the regulatory elements in the locus before GATA3 binding. Thus, YY1 cooperates with GATA3 and is required for regulation of the Th2 cytokine locus and Th2 cell differentiation.
