ABSTRACT
The role played by antigenic peptides bound to major histocompatibility complex (MHC) molecules is evaluated with H2-DMalpha(-/)- mice. These mice have predominantly class II-associated invariant chain peptide (CLIP)-, not antigenic peptide-bound, MHC class II. H2-DMalpha(-/)- donor heart grafts survived three times longer than wild-type grafts and slightly longer than I-A(beta)(b)-(/)- grafts. Proliferative T cell response was absent, and cytolytic response was reduced against the H2-DMalpha(-/)- grafts in vivo. Residual cytolytic T cell and antibody responses against intact MHC class I lead to eventual rejection. Removal of both H2-DMalpha and beta2-microglobulin (beta2m) in cardiac grafts lead to greater (8-10 times) graft survival, whereas removal of beta2m alone did not have any effect. These results demonstrate the significance of peptide rather than just allogeneic MHC, in eliciting graft rejection.
Subject(s)
Graft Rejection/immunology , HLA-D Antigens/immunology , Heart Transplantation/immunology , Major Histocompatibility Complex , Th1 Cells/immunology , Th2 Cells/immunology , Animals , Antigens, Differentiation, B-Lymphocyte/genetics , Antigens, Differentiation, B-Lymphocyte/physiology , Cytokines/genetics , Graft Rejection/genetics , HLA-D Antigens/genetics , Histocompatibility Antigens Class I/analysis , Histocompatibility Antigens Class II/genetics , Histocompatibility Antigens Class II/physiology , Mice , Mice, Inbred BALB C , Mice, Inbred DBA , Mice, Inbred Strains , Mice, Knockout , Myocardium/immunology , Transplantation, HomologousABSTRACT
Class II transactivator (CIITA) is an unusual transcriptional coactivator in that it contains a functionally important, GTP-binding consensus domain. To assess the functional role of the GTP-binding domain of CIITA in vivo, we have generated knockout mice that bear a mutation in the CIITA gene spanning the GTP-binding domain. Upon analysis, these mice show no detectable CIITA mRNA; hence, they represent mice with deleted CIITA rather than mice with defects in the GTP-binding domain only. In these knockout mice, MHC class II expression is nearly eliminated, although a faint RT-PCR signal is visible in spleen, lymph node, and thymus, suggestive of the presence of CIITA-independent regulation of MHC class II expression. Invariant chain expression is also greatly reduced, but to a lesser extent than MHC class II. Serum IgM is not decreased, but the serum IgG level is greatly reduced, further confirming the absence of MHC class II Ag-dependent Ig class switching. Induction of MHC class II expression by IL-4 or LPS was absent on B cells, and Mac-1+ cells showed no detectable induction of MHC class II by either IL-4, LPS, or IFN-gamma. These findings demonstrate a requirement for CIITA in IFN-gamma-, IL-4-, and endotoxin-induced MHC class II expression as well as the possibility of rare CIITA-independent MHC class II expression.
Subject(s)
GTP-Binding Proteins/genetics , Histocompatibility Antigens Class II/biosynthesis , Interferon-gamma/pharmacology , Interleukin-4/physiology , Lipopolysaccharides/pharmacology , Nuclear Proteins , Peptide Fragments/genetics , Sequence Deletion , Trans-Activators/genetics , Animals , Antigens, Differentiation, B-Lymphocyte/biosynthesis , Antigens, Differentiation, B-Lymphocyte/genetics , CD4-CD8 Ratio , CD4-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/immunology , Cell Membrane/immunology , Cell Membrane/metabolism , Crosses, Genetic , Female , Gene Expression Regulation/genetics , Gene Expression Regulation/immunology , Gene Targeting , Histocompatibility Antigens Class II/genetics , IgG Deficiency/genetics , Immunoglobulin M/blood , Interferon-gamma/antagonists & inhibitors , Interleukin-4/antagonists & inhibitors , Lipopolysaccharides/antagonists & inhibitors , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Peptide Fragments/deficiency , Trans-Activators/deficiency , Up-Regulation/immunologyABSTRACT
The TAP1 and LMP2 genes are central for class I MHC function and share a common promoter. Here, we analyze the molecular mechanism of IFN gamma up-regulation of TAP1 and LMP2. In vivo footprinting indicates IFN gamma up-regulates protein-DNA contacts at an IRF-E that is essential for the up-regulation of TAP1 and LMP2 by IFN gamma. Gel shift analysis indicates that this site binds IRF-1. The expression of TAP1 and LMP2 are both greatly reduced in IRF-1-deficient mice. Surface class I MHC as well as CD8+ T cells are reduced in IRF-1-/- mice. The role of IRF-1 in the regulation of TAP1 and LMP2 suggests a mechanism for the antiviral properties of IRF-1 and the unexpected deficiency of CD8+ T cells observed in IRF-1-/- mice.
Subject(s)
ATP-Binding Cassette Transporters/genetics , CD8-Positive T-Lymphocytes/cytology , Cysteine Endopeptidases , DNA-Binding Proteins/physiology , Interferon-gamma/physiology , Phosphoproteins/physiology , Proteins/genetics , ATP Binding Cassette Transporter, Subfamily B, Member 2 , Animals , Cell Differentiation , DNA Footprinting , Gene Expression Regulation, Developmental , Genes, MHC Class I , Interferon Regulatory Factor-1 , Mice , Mice, Transgenic , Promoter Regions, Genetic , RNA, Messenger/genetics , Up-RegulationABSTRACT
The lpr gene encodes a defective form of the fas gene that mediates apoptosis, and its expression results in autoantibodies and massive lymphadenopathy. bcl-2, another gene locus that affects programmed cell death, acts to inhibit apoptosis. Since multiple mechanisms controlling programmed cell death may contribute to systemic autoimmunity, the effect of the bcl-2 transgene on the lpr model was examined by crossing bcl-2 transgenic and C57BL/6-lpr mice. Compared with bcl-2-/lpr mice, bcl-2+/lpr showed dramatic increases in lymphadenopathy and T cell accumulation, but not in autoantibodies or B cell numbers. Short term transfer studies demonstrated that double negative T cells normally have a limited lifespan, and their survival is enhanced by the bcl-2 transgene. Thus, defects in separate apoptosis mechanisms may combine to produce enhanced pathologic effects.
