Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.7.49 (reverse transcriptase)
31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We previously demonstrated that the MHC class II molecule on murine sperm and CD4-like molecule on the egg vitelline membrane are involved in fertilization. In this study, the RNA transcripts in murine eggs corresponding to parts of the CD4 gene and lck gene in thymocytes were demonstrated by means of the reverse transcriptase (RT)/polymerase chain reaction (PCR) followed by the sequencing of the PCR products. The deduced sequences potentially encode the amino acid sequence of the transmembrane to the cytoplasmic domain of CD4 and that of the N-terminal domain of p56lck respectively. These findings indicate that a signal transducing complex similar to that in immune T cells is expressed at the transcriptional level in murine eggs.
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PMID:Expression of the signal transducing regions of CD4-like and lck genes in murine egg. 137 Aug 84

We previously identified an immunodominant CD4+ T cell determinant in the carboxy-terminal region of HIV-1 reverse transcriptase (RT528-543). The present study aimed at enumerating all the potential sites of HIV-1 RT recognized by Th cells in the BALB/c (H-2d) mouse model. To achieve this we used a panel of 62 overlapping 15-mer synthetic peptides covering the whole RT sequence to assay the following parameters: (i) immunogenicity in naive BALB/c mice injected either with peptides pools or individual peptides; (ii) antigenicity, as detected by their ability to restimulate in vitro T cells from BALB/c mice primed with native RT; (iii) MHC class II (Ad)-binding capacity as measured by the inhibition of the antigen-specific, Ad-restricted presentation of unfolded apamin (4-Acm) by fixed antigen-presenting cells to Ad/4-Acm-specific, interleukin-2-producing T hybridoma cells; and (iv) the presence of typical or degenerate consensus Ad-binding motifs. The results in this study permitted identification of three novel immunodominant RT mouse CD4+ T cell sites (RT276-290, RT375-389 and RT411-425) located in regions of limited polymorphism among RT from several HIV isolates. Some of these RT segments were found to be in the vicinity of B cell or H-2Kk- or HLA-A2-restricted cytotoxic T lymphocyte epitopes. Finally, the approach used in this study was found to be very efficient for enumerating most T cell recognition sites in a complex protein, a result that would have not been achieved by a single parameter-based analysis.
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PMID:Human immunodeficiency virus-1 reverse transcriptase immunodominant CD4+ T cell epitopes: a peptide-based multiparametric assessment in the mouse. 751 71

Dendritic cells (DC) act as potent primary antigen-presenting cells in many immune responses and therefore may have a role in the initiation and perpetuation of the synovial inflammation in chronic inflammatory arthritis. To examine their function, it is important to isolate fresh DC from arthritic joints without aberrant activation. We have developed a technique using minimal cell manipulation to isolate DC from the synovial fluid of chronic arthritic patients. Using this method, DC were shown to be potent allostimulatory cells, with 63-90% of cells lacking lineage-specific markers (lin-), but positive for MHC class II molecules. Two morphologically distinct populations of these cells were identified in 10 out of 13 DC preparations. Both populations expressed CD40, intercellular adhesion molecule-1 (ICAM-1), ICAM-2, ICAM-3 and leucocyte function associated antigen-3 (LFA-3), but the predominant population, which was larger and more typical of cultured blood DC, had a higher density of these antigens compared with the minor population, which were smaller and morphologically similar to lymphocytes. Two new MoAbs which label activated human blood DC, HB15 (CD83) and CMRF-44, were tested. CD83 labelled very weakly or not at all, whereas CMRF-44 was positive on the larger cells only. Likewise, the costimulator molecule, B7/BB1 (CD80), was not detected on the surface of either synovial lin- cell population, reverse transcriptase polymerase chain reaction (RT-PCR) showed little or no CD80 mRNA, and no binding of the CTLA-4Ig fusion protein was found. These results suggest that synovial DC are not, despite the inflammatory environment, in a fully activated state.
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PMID:Dendritic cells in synovial fluid of chronic inflammatory arthritis lack CD80 surface expression. 753 11

We have studied the effect of poly(ADP-ribose) synthetase on the interferon-gamma (IFN-gamma)-inducible expression of major histocompatibility complex (MHC) class II molecules. We constructed an expression plasmid capable of expressing either a sense RNA (MT-ARS) or an antisense RNA (pAS-FL or pAS-5') for poly(ADP-ribose) synthetase. We transfected the plasmid into mouse or human macrophage tumor cells and examined the effect on the expression of MHC class II molecules. The IFN-gamma-inducible expression of MHC class II gene was considerably reduced in transformant clones (A-2, B-2), in which the synthetase was highly expressed, whereas the depletion of the synthetase due to the expression of antisense RNA for the synthetase amplified the expression of MHC class II molecules. The results indicate that the level of the synthetase critically regulates the IFN-gamma-inducible MHC class II molecules. Next, we analyzed DNase I hypersensitive sites (DHS) of mouse MHC class II, I-A beta gene and found two sites, one in the promoter region and the other one in the first intron. The DHS in first intron was less sensitive towards DNase I attack in transformant clones (A-2, B-2) in which the synthetase was synthesized in a large quantity. Thus we constructed two beta-galactosidase reporter genes, one (A beta 2.0kb-lac z) containing the promoter region to a part of the second exon of the class II gene, and the other (A beta pro-lac z) containing the promoter region of the class II gene alone. The expression of the reporter gene was analyzed by reverse transcriptase-polymerase chain reaction (RT-PCR) and found that the expression of A beta 2.0kb-lac z was suppressed in the transformant clones (A-2, B-2) relevant to control cells but the expression of A beta pro-lac z was the same level among those cells.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Effect of poly(ADP-ribose) synthetase on the expression of major histocompatibility complex (MHC) class II genes. 757 32

In the present study, the expression of the CD4 molecule on murine egg plasma membrane was confirmed by the indirect immunofluorescence (IIF) method. The full-length CD4 cDNA from murine eggs was synthesised by the reverse transcriptase-polymerase chain reaction (RT-PCR) method and its authenticity verified by Southern blot hybridisation using an end-labelled internal oligonucleotide. The results of DNA sequencing showed that the nucleotide sequence of the cDNA of CD4 from murine egg mRNA was identical to that of immune T cells. To demonstrate the direct interaction of CD4 from murine egg with murine sperm cells bearing MHC (major histocompatibility complex) class II molecule, we employed a baculovirus expression system to generate CD4 on the surface of Spodoptera frugiperda (Sf9) cells. Expression of CD4 on Sf9 cells infected with Autographa californica nuclear polyhedrosis virus (AcNPV)-CD4 was demonstrated by IIF and immunoblotting. The CD4-expressing Sf9 cells adhered to MHC class II-bearing sperm cells since the adhesion was specifically blocked by anti-CD4 monoclonal antibody (mAb) or anti-monomorphic region of MHC class II mAb. Taking our previous and present experimental results together, they strongly suggest that intercellular membrane adhesion between two gametes at the fusion step in fertilisation is mediated by the MHC class II molecule located on the posterior region of the sperm head and the CD4 molecule on egg plasma membrane.
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PMID:Molecular structure and function of CD4 on murine egg plasma membrane. 761 76

Major histocompatibility complex (MHC) class II deficiency is an inherited autosomal recessive combined immunodeficiency. The disease is known as bare lymphocyte syndrome (BLS). BLS is characterized by a lack of constitutive MHC class II expression on macrophages and B cells as well as a lack of induced MHC class II expression on cells other than professional antigen-presenting cells (APCs) due to the absence of mRNA and protein of the human leukocyte antigen (HLA) class II molecules, designated HLA-DR, -DQ, and -DP. The defect in gene expression is located at the transcriptional level and affects all class II genes simultaneously. Here we have analyzed transcription and protein expression of class II antigens in Epstein-Barr virus (EBV)-transformed B lymphoblastoid cell lines and mononuclear cells (MNCs) of twin brothers. Whereas flow cytometric analysis failed to detect class II antigens on the cell surface of the patients' EBV-B cells and MNCs, examination of the genes coding for HLA-DR, -DQ, -DP, and the invariant chain (Ii) by reverse transcriptase-polymerase chain reaction amplification resulted in an unusual mRNA pattern in the B cell lines of the patients (HLA-DR alpha +, -DR beta, -DQ alpha +, -DQ beta -, -DP alpha -; -DP beta +, Ii+). In accordance with these findings no HLA-DR beta-specific protein was detected by immunoblotting, whereas low levels of HLA-DR alpha and normal levels of Ii were present. In contrast to EBV-B cells, the MNCs of both patients displayed a residual HLA-DR beta, -DQ beta, and -DP alpha mRNA signal. Furthermore, HLA-DR beta-specific protein was found in addition to HLA-DR alpha by immunoblotting of cell lysates, even though it was clearly decreased as compared with controls. Our results indicate that the defect in class II antigen expression is not necessarily present to the same extent in B cells and cells of other lineages. mRNA levels of HLA-DR beta were found to be enriched in adherent cells within the MNC fraction. Further investigations indicated that the MHC class II expressed is functional in antigen presentation, as the two boys' CD4+ T cells became activated and expressed interleukin-2R after stimulation of peripheral blood mononuclear cell cultures with recall antigen (tetanus toxoid). Furthermore, T cells tested in one of the two patients responded to both MHC class I and II allostimulation, and this response was inhibited by monoclonal antibodies of the respective specificity.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Molecular characterization of major histocompatibility complex class II gene expression and demonstration of antigen-specific T cell response indicate a new phenotype in class II-deficient patients. 769 27

Syngeneic graft-vs-host disease (SGVHD) is a MHC class II-restricted T cell-mediated autoimmune syndrome that occurs following syngeneic bone marrow transplantation and the administration of cyclosporin (CsA). The present studies evaluated the V beta repertoire of T lymphocytes that mediate SGVHD. To facilitate analysis, SGVHD effector cells were adoptively transferred into thymectomized syngeneic recipients reconstituted with T cell-depleted bone marrow to provide an environment that allows for the selective clonal expansion of autoreactive T cells. Analysis of target tissues and PBL by reverse transcriptase PCR using oligonucleotide V beta-specific primers revealed a predominance of V beta 8.5+ T cells and a minor population expressing V beta 10. The majority of infiltrating lymphocytes in target tissues was confirmed to be V beta 8.5+ by in situ hybridization and by immunoperoxidase staining. A small population of V beta 10+ cells could also be detected. Furthermore, SGVHD effector T splenocytes depleted of lymphocytes expressing either the TCR-alpha beta or the V beta 8.5 determinant could not adoptively transfer SGVHD. Depletion of T cells expressing the V beta 10 determinant delayed the onset of this autoaggression syndrome. Subset analysis of the autoreactive T cell compartment revealed that the V beta 8.5 determinant was expressed on both CD4+ and CD8+ lymphocytes whereas the V beta 10 determinant was principally expressed on a minor population of CD4+ autoreactive T cells. These data were confirmed by limiting dilution analysis. Additional studies examining the effect of CsA on thymic differentiation revealed that although V beta 8.5 is not normally clonally deleted, there was a pronounced shift in the expression of this determinant between CD4 and CD8 single positive thymocytes, suggesting that CsA may inhibit normal positive selection processes for MHC class I and class II reactive T cells.
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PMID:Characterization of the autoreactive T cell repertoire in cyclosporin-induced syngeneic graft-versus-host disease. A highly conserved repertoire mediates autoaggression. 770 14

Despite the important role played by endogenous superantigen in shaping the T cell repertoire, little is known concerning the expression of the different Mtv loci in cells of the thymic microenvironment involved in repertoire selection. Here we have examined the expression of a panel of Mtv Ags by different MHC class II+ stromal cel types using reverse transcriptase-PCR and monitored the effects of these stromal cells on the development of cells expressing Mtv-reactive TCR V beta elements in closed thymic organ culture systems. Although Mtv-6 and Mtv-8/9 mRNAs are expressed in normal thymus lobe organ cultures, no Mtv expression was detected in MHC class II+ thymic epithelial cells. In contrast a striking pattern of differential expression was observed in dendritic cells of thymic origin that were devoid of Mtv-8/9 but expressed readily detectable levels of Mtv-6. This pattern of Mtv gene expression correlated well with TCR V beta repertoire development. TCRV beta 3+ T cells, normally deleted in response to Mtv-6, were virtually absent from the single positive thymocyte compartment in thymic organ cultures where dendritic cells are present but were present in reaggregate cultures where the only MHC class II-positive cells were thymic epithelial cells. On the other hand, V beta 11+ T-cells were not deleted in organ cultures, possibly reflecting the absence of Mtv-8/9 expression in dendritic cells. Our studies suggest that the influence Mtvs have on shaping the T cell repertoire not only depends on their expression within a particular strain but also on their tissue specific expression in relation to MHC class II, which is necessary for their presentation.
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PMID:Differential expression of Mtv loci in MHC class II-positive thymic stromal cells. 817 5

Langerhans cells (LC) are Ag-presenting cells required for induction of primary immune responses in skin. After activation by Ag, LC express increased levels of MHC class II Ag, exhibit increased accessory cell activity, and migrate to regional lymph nodes where they stimulate T cells. One of the earliest manifestations of LC activation is the accumulation of increased amounts of IL-1 beta mRNA in LC within 15 min after exposure to contact allergens in vivo. To determine if enhanced IL-1 beta production by LC could be causally linked to epicutaneous sensitization, we injected IL-1 beta intradermally into the ears of BALB/c mice and extracted total epidermal RNA 4 h later. A quantitative reverse transcriptase-polymerase chain reaction technique was used to compare changes in IL-1 alpha, IL-1 beta, macrophage inflammatory protein 2, IL-10, TNF-alpha, and 1-A alpha chain mRNA signals caused by intradermally-injected IL-1 beta to those caused by intradermal IL-1 alpha or TNF alpha, or by topical application of the contact allergen trinitrochlorobenzene (3% TNCB). Intradermal injection of 25 ng IL-1 beta resulted in 5-to 100-fold enhancement of mRNA signals for IL-1 alpha, IL-1 beta, MIP-2, IL-10, TNF alpha, and class II I-A alpha, mimicking the changes caused by allergen. In contrast, injection of equivalent amounts of IL-1 alpha or TNF alpha did not significantly alter the epidermal cytokine pattern. Simulating the effects of topically applied TNCB, intradermally-injected IL-1 beta (but not IL-1 alpha or TNF alpha) also caused enhancement of LC MHC class II expression. In addition, LC derived from IL-1 beta-injected skin were 2 to 3 times more potent accessory cells in an anti-CD3 proliferation assay than LC from IL-1 alpha or sham-injected skin. Finally, injection of hamster anti-mIL-1 beta mAb into the skin prior to TNCB treatment completely prevented sensitization to this allergen, although injections of similar amounts of hamster anti-mIL-1 alpha mAb or PBS were without effect. Taken together, our data indicate that dendritic cell-derived IL-1 beta may be a critical molecule required for initiation of primary immune responses in skin.
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PMID:An essential role for Langerhans cell-derived IL-1 beta in the initiation of primary immune responses in skin. 847 27

Allograft rejection is dependent on T cell activation, which requires both the engagement of the T cell receptor by antigen in the context of the MHC molecules and costimulatory signals delivered by cell surface molecules such as B7-CD28/CTLA4 pathway. CTLA4-Ig is a fusion protein that blocks this pathway and has previously been shown to prolong both allograft and xenograft survival. The current study demonstrates markedly prolonged murine cardiac allograft survival and specific prolongation of secondary skin grafts using a combination of CTLA4-Ig plus donor bone marrow. A role for hematopoietic chimerism in the establishment of CTLA4-Ig-induced transplantation tolerance was investigated using reverse transcriptase polymerase chain reaction analysis of recipient tissues. Expression of donor-specific MHC class II transcripts in both peripheral and lymphoid tissues was demonstrated at greater than 200 days after transplant. To investigate the functional significance of this observation, heart donors, and donor bone marrow were irradiated before transplantation in CTLA4-Ig-treated recipients. A reduction in allograft survival was associated with irradiation of both the donor heart and the bone marrow. These results suggest that there may be a donor-derived radiosensitive element that enhances allograft survival in this model. Reverse transcriptase polymerase chain reaction analysis of allografts of tolerant and control animals at days 5, 8, and 12 after transplantation failed to demonstrate a dramatic difference in the expression of interleukin (IL)-2, IL-4, IL-10, and interferon-gamma message. Cytotoxicity effector transcripts were largely intact in CTLA4-Ig + bone marrow-treated recipients as they showed no decrease in intragraft granzyme, perforin, Fas, or Fas ligand transcripts during thr first 8 days after transplant. These results imply that complex mechanisms may be important for the induction and maintenance of transplantation tolerance in the CTLA4-Ig plus bone marrow murine cardiac allograft model.
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PMID:CTLA4-Ig plus bone marrow induces long-term allograft survival and donor specific unresponsiveness in the murine model. Evidence for hematopoietic chimerism. 862 6


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