ABSTRACT
Notch proteins regulate many developmental processes. Notch1 is highly expressed on thymocytes, but its role in regulating their development is not known. We show that activation of Notch1 signaling in CD4+CD8+ double positive thymocytes promotes the maturation of both CD4+ and CD8+ single positive thymocytes and that this occurs in the absence of interactions between the T cell receptor and MHC molecules expressed on thymic epithelial cells. We have also identified several genes that are transcriptionally regulated by Notch1 in T cells and show that they are upregulated during maturation into both single positive lineages. These observations suggest that Notch1 signaling plays a role in promoting maturation into both the CD4 and CD8 T cell lineages.
Subject(s)
CD4-Positive T-Lymphocytes/cytology , CD8-Positive T-Lymphocytes/cytology , Carrier Proteins , Disintegrins , Leukopoiesis , Membrane Proteins/metabolism , Metalloendopeptidases , Metalloproteases , Receptors, Cell Surface , Signal Transduction , Transcription Factors , ADAM Proteins , Animals , Basic Helix-Loop-Helix Transcription Factors , CD4-Positive T-Lymphocytes/metabolism , CD8-Positive T-Lymphocytes/metabolism , Epithelial Cells , Homeodomain Proteins/genetics , Hyaluronan Receptors/biosynthesis , Hypoxanthine Phosphoribosyltransferase/genetics , Major Histocompatibility Complex , Membrane Proteins/genetics , Mice , Mice, Inbred C57BL , Mice, Inbred DBA , Muscle Proteins/genetics , Nuclear Proteins/genetics , Phosphoproteins/genetics , Proteins/genetics , Receptor, Notch1 , Receptors, Interleukin-2/biosynthesis , Transcription Factor HES-1 , Up-RegulationABSTRACT
Thymocyte development is a tightly regulated process. CD4+CD8+ double-positive (DP) immature thymocytes exhibit distinct phenotypic features from mature T cells; they express only 10% of surface TCR that are found on mature T cells and do not proliferate and produce IL-2 in response to stimulation. In this report we show that transgenic expression of the orphan nuclear receptor ROR gamma t in mature T cells down-regulates their surface TCR expression. The ROR gamma t transgene inhibits IL-2 production by mature T cells, and this inhibition may be partially due to the inhibitory effect of ROR gamma t on c-Rel transcription. Furthermore, ectopic expression of ROR gamma t inhibits the proliferation of mature and immature T cells. These results, together with its predominant expression in DP thymocytes, suggest that ROR gamma t controls these distinct phenotypic features of DP thymocytes. Our data suggest that down-regulation of ROR gamma t expression in thymocytes is essential for their maturation.
Subject(s)
Down-Regulation/immunology , Receptors, Cytoplasmic and Nuclear/biosynthesis , Receptors, Retinoic Acid , Receptors, Thyroid Hormone , T-Lymphocyte Subsets/cytology , T-Lymphocyte Subsets/metabolism , Animals , Cell Differentiation/genetics , Cell Differentiation/immunology , Crosses, Genetic , Down-Regulation/genetics , Fas Ligand Protein , Gene Expression Regulation/immunology , Immunosuppressive Agents/pharmacology , Interleukin-2/antagonists & inhibitors , Interleukin-2/biosynthesis , Ligands , Lymphocyte Activation/immunology , Membrane Glycoproteins/biosynthesis , Mice , Mice, Inbred C57BL , Mice, Inbred CBA , Mice, Transgenic , Nuclear Receptor Subfamily 1, Group F, Member 3 , Proto-Oncogene Proteins c-rel/antagonists & inhibitors , Proto-Oncogene Proteins c-rel/biosynthesis , Receptors, Antigen, T-Cell/antagonists & inhibitors , Receptors, Antigen, T-Cell/biosynthesis , Receptors, Cytoplasmic and Nuclear/genetics , Receptors, Cytoplasmic and Nuclear/physiology , T-Lymphocyte Subsets/immunology , Thymus Gland/cytology , Thymus Gland/metabolism , Transgenes/immunology , Up-Regulation/genetics , Up-Regulation/immunology , fas Receptor/metabolismABSTRACT
The Notch receptor and its ligands are involved in many developmental processes. They are highly expressed in the thymus and have been implicated in the CD4 versus CD8 lineage decision. We identified the constitutively active intracellular fragment of murine Notch-1 as capable of rendering thymomas resistant to glucocorticoid-induced apoptosis. This effect was confirmed in other T cell lines and in CD4+ CD8+ DP thymocytes. Activation of the Notch signaling pathway also upregulated a number of other markers that, like steroid resistance, correlate with DP maturation into both the CD4 and CD8 lineages. These results suggest that Notch signaling is critically involved in the maturation of DP thymocytes into both CD4+ and CD8+ SP thymocytes.
Subject(s)
CD4-Positive T-Lymphocytes/metabolism , CD8-Positive T-Lymphocytes/metabolism , Drosophila Proteins , Leukopoiesis , Membrane Proteins/metabolism , Receptors, Cell Surface/metabolism , Signal Transduction , Transcription Factors , Animals , Binding Sites , CD4-Positive T-Lymphocytes/cytology , CD4-Positive T-Lymphocytes/drug effects , CD8-Positive T-Lymphocytes/cytology , CD8-Positive T-Lymphocytes/drug effects , Drug Resistance , Glucocorticoids/pharmacology , Insect Proteins/genetics , Intracellular Fluid , Mice , Peptide Mapping , Proto-Oncogene Proteins c-bcl-2/biosynthesis , Receptor, Notch1 , Receptors, Antigen, T-Cell, alpha-beta/biosynthesis , Thymus Gland , Tumor Cells, Cultured , Up-RegulationABSTRACT
We have identified RORgamma t, a novel, thymus-specific isoform of the orphan nuclear receptor RORgamma that is expressed predominantly in CD4+ CD8+ double-positive thymocytes. Ectopic expression of RORgamma t protects T cell hybridomas from activation-induced cell death by inhibiting the upregulation of Fas ligand. Following hybridoma stimulation, RORgamma t also inhibits IL-2 production but does not affect the induction of Nur-77 and Egr-3 nor the upregulation of CD69. Both the ligand-binding and DNA-binding domains of RORgamma t are required for this effect. We propose that the role of RORgamma t expression in immature thymocytes is to inhibit Fas ligand expression and cytokine secretion following engagement of their TCR during positive or negative selection.
