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)

The expression of endogenous ecotropic viruses in radiation-induced thymomas of C57BL/6 mice was examined. Competition radioimmunoassays for AKR MuLV gp71, p30, and p12 were used for viral antigen expression. 3 of 40 lymphomas had readily detectable ecotropic gp71 at levels of 95-689 ng/mg protein; the remainder of the tumors had no detectable gp71 (less than 1.0 ng/mg protein). 30 thymomas were characterized by the presence of MuLV p30 at levels of 1-10 ng/mg protein, levels that were comparable to those found in thymus extracts from age-matched, nonirradiated control. 10 tumors were characterized by having p30 levels of 10-30 ng/mg protein. In one tumor significant levels of AKR MuLV p12 were detectable. Since B-tropic and N-tropic viruses from C57BL/6 mice have glycoproteins (gp71) indistinguishable from AKR MuLV gp71 and the N-tropic virus had a p12 serologically identical to AKR MuLV p12, these results demonstrate that overt endogenous B-tropic virus was detectable in 2 of 40 thymomas and endogenous N-tropic virus was detectable in 1 of 40 thymomas. The lack of overt expression of gp71 or p12 was also confirmed by cytotoxicity assays using monospecific antisera to these viral proteins. Radiation-induced lymphomas were also examined for the presence of reverse transcriptase after chromatography of tissue extracts on poly G-Sepharose. One tumor, which was characterized by the lack of gp71, also had no detectable reverse transcriptase; whereas one tumor with gp71 was characterized by readily detectable levels of reverse transcriptase in cellular extracts. The presence of viral RNA was examined using AKR cDNA. Low levels of RNA capable of hybridizing with AKR cDNA were found in age-matched, nonirradiated mice; these hybrids had Tm's of 72 degrees C, while hybrids with AKR MuLV 70S RNA had Tm's of 80 degrees C. In 1 of 12 thymomas the concentration of hybridizable RNA and the Tm of the hybrids were identical to control values. In 9 of 12 thymomas the concentration of hybridizable sequences increased approximately three-to fivefold and the Tm of these hybrids varied from 73 to 75 degrees C. In 1 of 12 thymomas the concentration of hybridizable sequences increased over 100-fold, hybridized completely with AKR MuLV cDNA, and the hybrids had Tm's of 79 degrees C. This thymoma was also characterized by the presence of the AKR MuLV type of gp71 and p12. One tumor was characterized by a 10-to 100-fold increase in hybridizable sequences, which only partially hybridized with AKR MuLV cDNA, and hybrids had a Tm of 73 degrees C. This tumor was characterized by the presence of AKR MuLV gp71 but not AKR MuLV p12. The results taken together demonstrate that overt endogenous ecotropic virus expression is only rarely detectable in radiation-induced thymomas of C57BL/6 mice.
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PMID:Radiation leukemia in C57BL/6 mice. II. Lack of ecotropic virus expression in the majority of lymphomas. 6 29

We investigated by molecular hybridization whether T cells contain RNA sequences homologous to RNA which codes for immunoglobulin kappa-chain (k-chain). A radioactive probe of complementary DNA (cDNA) was prepared by transcription of purified k-chain mRNA from mouse myeloma MOPC-41 with reverse transcriptase (RNA-dependent-DNA nucleotidyltransferase) from avian myeloblastosis virus. The cDNA probably corresponded only to the constant region and 3'-terminus of k-chain mRNA. Kappa-chain cDNA was found to hybridize efficiently with RNA from both thymus cells and an established culture of thymoma cells. The thymus and thymoma cells contained 99.8% and 100% theta-positive cells, respectively. Quantitatively the average thymus T cell (thymus derived lymphocyte) contained about one half as much k-chain mRNA as the average spleen B cell ("bursa" dependent lymphocyte), whereas the thymoma cells contained only 1/33 as much. Control hybridizations of k-chain cDNA with myeloma and liver RNA support the conclusion that T cells in the thymus and in the thymoma cell line synthesize k-chain mRNA-like molecules. The thermal stability of hybrids of k-chain cDNA with RNA from spleen, thymus, thymoma, and another k-chain producing myeloma tumor was lower than that with MOPC-41 RNA. This finding may be due to the existence of several slightly different ck genes in the mouse as suggested by various control experiments.
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PMID:Sequences related to immunoglobulin kappa chain messenger RNA in T cells. 82 Oct 55

A reverse transcriptase-polymerase chain reaction (RT-PCR) assay is described that allows the rapid detection and quantitation of mRNA encoding the cytokines interleukin-2 (IL-2), IL-4 and interferon-gamma (IFN-gamma). Analysis of cytokine production by defined CD4+ T cell clones and the thymoma cell line EL4, demonstrates that the oligonucleotide primers used in this assay are specific for the genes encoding the individual cytokines, generating PCR products of different sizes. This allows the simultaneous and unambiguous detection of all three cytokine mRNAs in the same cDNA sample. The assay is sensitive enough to reproducibly detect cytokine mRNA expressed in as few as ten cells and requires 10,000-fold less cells for the detection of IL-2 production than that required for its detection using a conventional bioassay. Reverse transcribed mRNA is quantitated in the PCR assay by amplifying in the presence of a known amount of competitive genomic DNA (gDNA) template containing a small intron using the same primers. The PCR products obtained form the target cDNA and gDNA templates, which are distinguished by size, are processed by Southern analysis and quantitated by scanning densitometry of autoradiographs. As little as two-fold differences in cytokine mRNA can be reliably detected using this assay. We have demonstrated the successful application of this assay to the quantitation of pg amounts of IL-2 mRNA that is constitutively produced at low levels by fetal thymocytes in vivo during T cell ontogeny. The sensitivity, specificity, reliability and speed of this assay will facilitate the analysis of cytokine production in in vivo-derived or, in vitro propagated cells which are not available in sufficient numbers for analysis using more conventional molecular and biochemical assays.
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PMID:A polymerase chain reaction assay for the detection and quantitation of cytokine gene expression in small numbers of cells. 162 16

The expression of the type-C retrovirus and the virus-related components in NFS mice were examined during preleukemic and leukemic phases after fractionated whole-body X irradiation. The NFS mice were highly susceptible to induction of thymoma by fractionated X irradiation. The leukemic tissues were negative for infectious type-C virus, as detected by both the XC-plaque test and mink S+ L- focus-inducing assays, but contained a substantially higher level of viral-specific RNA-dependent DNA polymerase activity and a major core protein p30 than the corresponding tissues from unirradiated age-control mice. In the preleukemic phase, the amount of p30-related antigen increased transiently in spleen. The leukemic cell lines established from radiation-induced lymphomas produced particulate entities with a buoyant density of about 1.15 g/ml. These virus-like particles lacked in vitro infectivity to mouse cells and mink lung cells and leukemogenicity in syngeneic mice. The p30-related antigens of these particles were immunologically similar to that of xenotropic virus derived from NZB mouse.
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PMID:Endogenous type-C viral expression during lymphoma development in irradiated NFS mice. 241 71

Aberrant elevation of serum IgA and induction of murine IgA nephropathy following dietary exposure to the naturally occurring trichothecene vomitoxin (VT or deoxynivalenol) may involve dysregulation of cytokine production at the T cell level. EL4.IL-2 (EL-4), a cloned thymoma that produces interleukins (IL)-2, 4, 5, and 6, was used as a T cell model to investigate the in vitro effects of VT on interleukin production and gene expression. When supernatants of cells stimulated with phorbol 12-myristate 13-acetate (PMA) were assessed by enzyme-linked immunosorbent assay, IL-2, 4, and 5 were increased in the presence of 50 and/or 100 ng/ml VT for 2 and/or 8 days of culture. IL-2, 5, and 6 were also significantly elevated in the presence of 10-100 ng/ml of cycloheximide (CHX), another protein synthesis inhibitor, after 8 days of culture. As demonstrated by Northern analysis, VT at the levels between 50 and 100 ng/ml superinduced IL-2, 4, 5, and 6 mRNAs in PMA-stimulated EL-4 cells during a 24 hr culture period. Similar effects in PMA-treated samples were observed for CHX at 50, 100, 250, 1000, and 10000 ng/ml. mRNA levels for both IL-4 and IL-5, but not IL-2 and IL-6, were increased in unstimulated EL-4 cultures exposed to 50 and 100 ng/ml VT for 48 hr when analyzed by reverse transcriptase-polymerase chain reaction. Using [3H]leucine incorporation as a measurement of protein synthesis, IC50s for VT and CHX were estimated to be 280 and 55 ng/ml, respectively. This study indicates that VT as well as CHX could increase production of several interleukins in the EL-4 model even when present at concentrations that partially inhibited protein synthesis, whereas IL mRNA superinduction occurred across a broader range of concentrations that included maximal protein synthesis inhibition.
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PMID:Elevated gene expression and production of interleukins 2, 4, 5, and 6 during exposure to vomitoxin (deoxynivalenol) and cycloheximide in the EL-4 thymoma. 804 72

Autoantibodies against both striated muscle proteins, particularly titin, and the acetylcholine receptor are a hallmark of thymoma-associated myasthenia gravis. However, the stimulus for these responses remains enigmatic as whole titin is not detectable in these tumors. This study reports that in thymomas with cortical differentiation many of the neoplastic epithelial cells expressed low and medium molecular weight neurofilaments detected with several antibodies (on selections and blots) and at the RNA level (by reverse transcriptase polymerase chain reaction). Moreover, higher molecular weight forms sharing at least one epitope with titin were detectable slightly less frequently, as were the more strongly phosphorylated epitopes. In stark contrast, in medullary and mixed thymomas, and especially in the normal thymus, immunoreactivity with anti-neurofilament antibodies was rare. This aberrant overexpression of a titin epitope by epithelial cells with antigen-presenting phenotype in an inappropriate cortical microenvironment suggests that they might autosensitize maturing T cells there and so initiate anti-titin autoimmunity in these patients.
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PMID:Expression of neurofilaments and of a titin epitope in thymic epithelial tumors. Implications for the pathogenesis of myasthenia gravis. 866 70

A xenograft model of the human disease Langerhans cell histiocytosis (LCH) was investigated with severe combined immunodeficiency (SCID) mice. Transplantation of human LCH biopsy material into SCID mice resulted in the generation of mouse tumors resembling lymphomas. A thymoma cell line (ThyE1M6) was generated from one of these mice and found to display significant levels of Mg2+-dependent reverse transcriptase activity. Electron microscopy revealed particles with type D retroviral morphology budding from ThyE1M6 cells at a high frequency, whereas control cultures were negative. Reverse transcription-PCR of virion RNA with degenerate primers for conserved regions of various mouse, human, and primate retroviruses amplified novel sequences related to primate type D retroviruses, murine intracisternal A particles, Jaagsiekte sheep retrovirus, and murine long interspersed nuclear elements but not other retroviral classes. We demonstrate that these sequences represent a novel group of endogenous retroviruses expressed at low levels in mice but expressed at high levels in the ThyE1M6 cell line. Furthermore, we propose that the activation of endogenous retroviral elements may be associated with a high incidence of thymomas in SCID mice.
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PMID:Novel endogenous type D retroviral particles expressed at high levels in a SCID mouse thymic lymphoma. 1023 25

[(111)In-DTPA0]octreotide scintigraphy allows the in vivo visualization of several types of SS receptor (SSR)-expressing tumors. Among these, thymomas have been recently detected. Here we report on 2 patients admitted for myasthenia gravis and radiological evidence of thymic mass. Although these patients had similar clinical presentation, in vivo SSR scintigraphy displayed a difference in the degree of the [(111)In-DTPA0]octreotide uptake. Considering that both thymic masses had comparable volume, [(111)In-DTPA0]octreotide level was significantly higher in one of the 2 tumors (tumor/background ratio of 5.7 vs 2.6). The SSR subtype expression pattern was studied in vitro on the surgically resected specimens by ligand binding techniques, quantitative reverse transcriptase polymerase chain reaction (RT-PCR) and immunohistochemistry. According to the recent World Health Organization classification, the 2 tumors were classified A and B2 thymomas respectively. In membrane homogenates, we found a higher number of high affinity [125I-Tyr11]-SS-14 binding sites in the B2 thymomas (23.5+/-2.5 vs 12.0+/-0.4 fmol/mg membrane protein; p<0.05). RT-PCR analysis showed sst1, sst2A and sst3 mRNA in the 2 thymoma tissues, whereas SS mRNA was detectable only in the A thymoma. Quantitative evaluation of RT-PCR data showed a comparable expression of the relative amount of sst2A mRNA in both tumors, whereas a significant higher expression of sst3 mRNA in the B2 thymoma. Sst2A immunoreactivity was localized mainly on the endothelium of intratumoral vessels, whereas sst3 was present on either tumoral epithelial cells or normal reactive thymocytes. The expression of sst2A receptors in these tumors is in line with the in vivo visualization by [(111)In-DTPA0]octreotide, which is considered a sst2-preferring ligand. However, since radioligand uptake was significantly higher in the B2 thymoma, which expressed the largest sst3 mRNA levels, it might be possible that this subtype is involved in determining the tumor visualization during SSR scintigraphy. Apart from the affinity of the radioligand for the receptor, also the efficacy of the internalization of the radioligand-receptor complex might play a role in radioactivity accumulation during in vivo SSR scintigraphy. In fact, although octreotide binds with lower affinity to sst3 receptors, this subtype displayed the highest amount of agonist-dependent receptor internalization compared to the other SSR subtypes. Moreover, sst3 was localized on both tumor cells and reactive thymocytes, and these latter cells are characterized by a very active turnover of membrane molecules. Finally, although more cases need to be evaluated, the lack of detection of SS mRNA in the tumor presenting a more aggressive phenotype (B2 thymoma) might have physiopathological or prognostic significance.
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PMID:In vivo and in vitro expression of somatostatin receptors in two human thymomas with similar clinical presentation and different histological features. 1150 87

Critical checkpoints controlling early thymic T-cell development and homeostasis are set by the proper signaling function of the interleukin 7 receptor (IL-7R) and the pre-T-cell antigen receptor. Although alpha beta T-cell development is observed in IL-7- and IL-7R alpha-deficient mice, the number of thymocytes is significantly reduced, implying a role for the IL-7R in controlling the size of the thymic T-cell compartment. Here, we report the overexpression of IL-7R alpha that occurs in the early T-cell compartment from AKR/J mice, animals that are highly susceptible to the spontaneous development of thymoma. Increased IL-7R alpha was revealed by surface staining, and increased IL-7R alpha mRNA was documented by using reverse transcriptase-polymerase chain reaction (RT-PCR). This resulted in increased survival of AKR/J early thymocytes, shown by the decreased frequency of TUNEL(+) (terminal deoxynucleotidyl transferase mediated deoxyuridine triphosphate [dUTP]-fluorescein nick end labeling) cells. In an in vivo thymocyte repopulation model, AKR/J thymocytes had a selective advantage over healthy thymocytes. This advantage occurred at early stages of T-cell development. Our findings support the model that overexpression of growth factor receptors can contribute to proliferation and malignancy.
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PMID:Overexpression of IL-7R alpha provides a competitive advantage during early T-cell development. 1459 27

Cell death caused by the accumulation of extracellular adenosine is believed to contribute to the profound loss of T lymphocytes in patients with severe combined immunodeficiency disease due to adenosine deaminase deficiency. Although adenosine is known to trigger apoptosis in thymocytes and peripheral T cells, the molecular basis of this effect is not understood. In this study, we show that adenosine-induced apoptosis in mouse EL-4 thymoma cells was associated with the generation of reactive oxygen species and a reduction in mitochondrial transmembrane potential. In addition, cell death was by a caspase-independent mechanism because caspase inhibitors did not protect EL-4 cells from adenosine-induced cytotoxicity. Although reverse transcriptase polymerase chain reaction revealed that EL-4 cells expressed A2b and A3 adenosine receptor subtypes, blockade of A2b and A3 adenosine receptors with receptor-selective antagonists did not attenuate adenosine-induced cell death. Nevertheless, the failure of nucleoside transport inhibitors to prevent adenosine cytotoxicity suggested that adenosine was acting through a cell-surface receptor. In addition, adenosine-induced apoptosis was not due to an accumulation of intracellular cyclic adenosine monophosphate (cAMP) since neither forskolin nor 8-Br-cAMP was cytotoxic for EL-4 cells. Adenosine therefore acts through a non-classical receptor at the cell surface to trigger caspase-independent apoptosis in mouse thymoma cells.
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PMID:Adenosine-induced apoptosis in EL-4 thymoma cells is caspase-independent and mediated through a non-classical adenosine receptor. 1616 10


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