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 Friend erythroleukemia cell line T3-C12, which produces Friend murine leukemia virus (F-MuLV) and can be induced to synthesize hemoglobin by dimethyl sulfoxide (DMSO), was monitored for viral RNA-dependent DNA polymerase reverse transcriptase (RT) activity. The amount of viral 60-70S RNA released from DMSO-treated cells was unaffected or increased compared to that from control cells, while RT activity from treated cells was decreased. Accordingly, the specific activity in F-MuLV from DMSO-treated cells expressed as RT/70S RNA was decreased to 8% of the control activity. The 5-bromo-2'-deoxyuridine added to cultures containing DMSO reversed the differentiation process, and the F-MuLV thus treated did not exhibit the reduced RT activity normally observed in DMSO-treated virus. Cell-free F-MuLV incubated with and without DMSO showed the same RT activity, indicating that DMSO itself did not inhibit RT activity. However, when F-MuLV-containing pellets from control and DMSO-treated culture fluids were mixed, there was marked inhibition of the control RT activity, suggesting that RNase hybrid activity was stimulated or that an inhibitor was produced. Assays of F-MuLV-RNase hybrid released from control and DMSO-treated cells showed no difference in activity, indicating that a specific inhibitor of RT was produced or activated. Additions of certain nucleotide triphosphates to RT incubation mixtures did not result in any stimulation of RT activity in DMSO-treated F-MuLV, suggesting that phosphatase was not responsible for the observed inhibition. The results suggested that DMSO treatment of T3-C12 cells caused a reduction in viral RT activity by stimulating the production of an inhibitor, the nature of which is unknown.
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PMID:Viral reverse transcriptase suppression associated with erythroid differentiation of Friend leukemia cells. 6 77

The drug 3'-azido-3'-deoxythymidine (AZT), a synthetic thymidine analogue, has been used clinically in the management of acquired immune deficiency syndrome (AIDS). The drug is an effective antiviral agent due to its ability to block reverse transcriptase activity. This action of AZT was demonstrated in the Rauscher leukemia virus (RLV)-induced murine erythroleukemia model system. Unfortunately, associated with AZT has been the development of hematopoietic toxicity manifested by anemia, neutropenia, and overall bone marrow suppression. Hematopoietic growth factors (GM-CSF, erythropoietin), cytokines (interleukin-1), and agents known to potentiate hematopoiesis (lithium) have been demonstrated to modulate drug and/or radiation-induced hematopoietic toxicity. We report the results of further studies designed to investigate the ability of GM-CSF, erythropoietin, interleukin-1, and lithium to modulate AZT toxicity on murine hematopoietic granulocyte-macrophage (CFU-GM), megakaryocytic (CFU-Meg), and erythroid (BFU-E) progenitors cultured from bone marrow and spleen cells from mice infected with RLV. Hematopoietic progenitors from either normal or RLV-infected animals when exposed to AZT demonstrated concentration-dependent toxicity and differed for each progenitor with BFU-E being the most sensitive (ID50 concentration, 5 x 10(-9) M) and CFU-GM the least sensitive (ID50 concentration, 5 x 10(-5) M). As has been previously demonstrated using normal murine hematopoietic progenitors, when cultured with RLV-infected marrow or spleen cells, addition of GM-CSF, Meg-CSF or erythropoietin failed to inhibit AZT toxicity in vitro on CFU-GM, CFU-Meg, and BFU-E, respectively. However, in the presence of interleukin-1 (recombinant human IL-1 alpha, 30 ngm) or lithium chloride (ultra-pure, 1.0 mM), AZT toxicity CFU-GM, CFU-Meg, and BFU-E cultured from RLV-infected marrow or spleen cells was reduced. These results further demonstrate interleukin-1 and lithium are effective in modulating the toxic action of AZT on hematopoietic progenitors and that RLV-infected animals serve as a useful viral model system to study the effect of agents capable of modulating hematopoiesis in the presence of the anti-viral drug AZT.
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PMID:Effect of interleukin-1, GM-CSF, erythropoietin, and lithium on the toxicity associated with 3'-azido-3'-deoxythymidine (AZT) in vitro on hematopoietic progenitors (CFU-GM, CFU-MEG, and BFU-E) using murine retrovirus-infected hematopoietic cells. 194 Jun 11

Fibroblast colony-forming units (CFU-F) in bone marrow and spleen were investigated after infection of BALB/c mice with the Rauscher leukemia virus complex (RLV) or with cellularly cloned helper virus (R-MuLV). Both viral preparations induced a transient suppression of CFU-F prior to the development of leukemia. A second CFU-F decrease was observed in the terminal phase of R-MuLV-induced chronic lymphoid or myeloid leukemias, but not in mice with a RLV-induced acute erythroleukemia, which are highly viremic. When a number of Rauscher virus transformed leukemic cell lines were injected intravenously an erythroid and a T-cell line suppressed CFU-F values, but a third non-virus producing B-cell line did not. In contrast to the in-vitro situation, in-vitro inhibition of CFU-F was only observed in conditioned medium of an erythroid cell line with undetectable reverse transcriptase activity, whereas conditioned medium of lymphoid and myeloid cell lines were not inhibitory irrespective of reverse transcriptase activity. Together these results indicate that murine leukemia viruses can suppress the numbers of detectable CFU-F in vivo in an early phase of the disease and that leukemic cells may inhibit CFU-F proliferation by a mechanism which is not related to viremic state of the animal or the production of virus by these cells.
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PMID:Effects of Rauscher murine leukemia virus on hemopoietic organ-derived clonogenic fibroblasts. 242 Nov 8

RNA amplification with transcript sequencing (RAWTS) is a rapid and sensitive method of direct sequencing that involves complementary DNA synthesis, polymerase chain reaction (PCR) with a primer or primers containing a phage promoter, transcription from the phage promoter, and reverse transcriptase-mediated sequencing. By means of RAWTS, it was possible to sequence each of four tissue-specific human messenger RNAs (blue pigment, factor IX, phenylalanine hydroxylase, and tyrosine hydroxylase) in four cell types examined (white blood cells, liver, K562 erythroleukemia cells, and chorionic villus cells). These results indicate that there is a basal rate of transcription, splicing, and polyadenylation of tissue-specific mRNAs in adult and embryonic tissues. In addition to revealing sequence information, it is possible to generate a desired in vitro translation product by incorporating a translation initiation signal into the appropriate PCR primer. RAWTS can be used to obtain novel mRNA sequence information from other species as illustrated with a segment of the catalytic domain of factor IX. In general, the ability to obtain mRNA sequences rapidly across species boundaries should aid both the study of protein evolution and the identification of sequences crucial for protein structure and function.
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PMID:Access to a messenger RNA sequence or its protein product is not limited by tissue or species specificity. 279 88

Rauscher murine leukemia virus (R-MuLV) induces a rapidly developing erythroleukemia in BALB/c mice. Previously, we have shown that mouse interferon-alpha/beta (Mu IFN-alpha/beta) applied shortly after virus inoculation efficiently inhibits the leukemic process (Hekman et al., 1981). Here we describe the effect of Mu IFN-alpha/beta on an established leukemia. Varying doses of Mu IFN-alpha/beta were injected over 3 days, starting 8 to 12 days after virus inoculation. The effect of Mu IFN-alpha/beta on the leukemic process was monitored by measuring the spleen weight, reverse transcriptase activity in the serum and, in selected experiments, by microscopic examination of sections of the spleen using standard histological and immunological staining techniques. Depending on the spleen weight at the start of its application (maximal about 450 mg), Mu IFN-alpha/beta caused a dramatic reduction in the number of virus-infected erythroleukemic cells in the spleen. Also, R-MuLV disappeared from the serum within 3 days. If Mu IFN-alpha/beta was injected into R-MuLV-infected mice with an already 10-fold enlarged spleen, it could only stop further development of leukemia. Results obtained with crude Mu IFN-alpha/beta preparations were confirmed with absolutely pure Mu IFN-beta.
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PMID:The effect of murine interferon-alpha/beta on an established Rauscher murine leukemia virus-induced erythroleukemia in BALB/c mice. 258 Aug 3

Studies of recombinants between murine leukemia viruses (MuLVs) that cause thymic or erythroid leukemias have shown that enhancer sequences in the long-terminal repeats (LTRs) can determine the target tissues for pathogenesis. It has been inferred that the enhancers may specifically target viral expression into the cells that then become neoplastic. However, the neoplasms in those studies formed after latencies and contained ultimate viruses (called MCFs) that differed from the injected viruses in their enhancer sequences and envelope (env) genes. Transcriptional activities of LTRs from these proximal and ultimate viruses have not been thoroughly analyzed in different hematopoietic lineages. We present evidence that the enhancer of Friend spleen focus-forming virus (SFFV), an ultimate erythroleukemogenic retrovirus, contains an unstable 42-nucleotide direct repeat. Other ultimate erythroleukemogenic MuLVs (Friend MCFs) contain an enhancer nearly identical to that of SFFV both in its sequence and in its specific instability. The instability occurs in sequences that contain inverted repeats and we propose that it occurs by a simple reverse transcriptase hop mechanism. We constructed plasmids that contain the two forms of the SFFV LTR linked to the bacterial chloramphenicol acetyltransferase (CAT) gene, and we compared these in transient transfection assays with LTR-CAT plasmids constructed from Friend and Moloney MuLVs. The assays employed erythroleukemia cells, thymic lymphoma cells, and fibroblasts. The tropisms of expression correlated only weakly with tissue specificities of pathogenesis and each LTR was active in all cells. The SFFV 42-nucleotide duplication reduced expression in erythroid cells and increased expression in fibroblasts. We conclude that retroviral enhancers do not stringently direct gene expression into specific cell lineages, but on the contrary they are leaky and contain replicative instabilities that also may facilitate viral entrenchment throughout the host. These results have important implications for understanding murine retroviral evolution and the multi-step process of leukemogenesis.
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PMID:An enhancer sequence instability that diversifies the cell repertoire for expression of a murine leukemia virus. 283 56

An erythroleukemia cell clone, 7C, which failed to produce reverse transcriptase-containing virions or infectious virus, was found to produce noninfectious virus particles by gradient banding of [3H]leucine- and [3H]uridine-labeled virions. The RNA from the 7C virus was shown to consist of the normal 70S size component, which converted to 35S upon heat denaturation. In contrast, the 7C virion proteins showed multiple defects. Analysis of the virion proteins by gel electrophoresis demonstrated that the pr65 gag precursor was incorporated into the 7C virus and that the processing of this precursor was severely diminished. Polymerase proteins pr180gag-pol and pr120pol were also detected in virions, and a third possible polymerase protein, p70, was reduced in size compared to its normal counterpart, p80. Incorporation of the viral gp70 glycoprotein into particles was also reduced 10-fold, despite synthesis and incorporation of gp70 into the 7C cell membrane in normal amounts. Pulse-chase analysis of the synthesis of the viral gag and env proteins in 7C cells showed greatly reduced amounts of pr180gag-pol, pr65gag, p80gag, and p42gag, whereas pr90env, gp70, and spleen focus-forming virus-specific gp55 were synthesized and processed normally. These results suggested that at least one defect in 7C virus was impaired cleavage of gag or pol proteins or both, most likely due to a lack of the appropriate viral protease, and that this lack of cleavage might affect incorporation of gp70 into virus particles.
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PMID:Replication-defective Friend murine leukemia virus particles containing uncleaved gag polyproteins and decreased levels of envelope glycoprotein. 616 68

The effect of high doses of poly(I).poly(C) induced mouse L-cell interferon on the development of Rauscher murine leukemia virus (R-MuLV)-induced erythroleukemia in BALB/c mice was determined. Female mice, 4 to 5 weeks old, were infected with R-MuLV and treated with interferon every 24 h starting 6 h after virus inoculation. Under these conditions injection of 3-5 X 10(4) units of interferon caused a partial inhibition of the leukemia process. Daily application of 3 X 10(5) units completely or almost completely inhibited the erythroleukemia. After 14 days of treatment with these high doses of interferon, spleen weights of interferon-treated infected mice were comparable to those of uninfected animals which received only interferon. Also, no Rauscher cells in spleens and livers of R-MuLV-infected interferon-treated infected animals could be demonstrated and the spleen structure was well preserved in these mice. In interferon-treated infected animals no virus could be detected in the serum as judged from the absence of reverse transcriptase activity in the serum. Moreover, no virus-infected cells could be demonstrated in spleen or liver as deduced from negative immunofluorescence data using anti-p30 and anti-gp70 sera. No virions budding from spleen cell membranes were seen by electron microscopic studies. However, when interferon treatment was stopped the leukemic process was reactivated and all the mice died. In control experiments interferon caused an inhibition of red blood cell formation and a 50 to 100% enlargement of the spleen. Pharmacokinetic data showed that, after intraperitoneal inoculation, maximum amounts of interferon were present in the peripheral blood after 1-2 h. After 12-24 h almost all interferon activity had disappeared from the blood.
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PMID:The effect of high doses of poly(I).poly(C) induced mouse L cell interferon on Rauscher leukemia virus induced erythroleukemia in BALB/c mice. 616 93

A new cell line designated SQ-A was established from the spleen of a leukemic DBA/2J mouse inoculated with the anemic strain of Friend erythroleukemia virus (FLV-A). The cells are similar in morphology, growth pattern, and tumorigenicity to our prototype erythroleukemia line 5-86 but are more sensitive to the cytotoxic effects of inducers of differentiation. The virus produced by SQ-A cells induces erythroleukemia associated with anemia in adult mice but has little activity when assayed on XC cells. It was characterized to determine what factors influence its leukemogenic potential. As compared to the attenuated virus from cultures of 5-86 and G-2 cells, the subunits of the RNA from the virions of SQ-A cells are the same size, and the amount of reverse transcriptase activity and RNase H present in the purified virions of the three lines are similar. However, differences are observed in levels of endonuclease and protein kinase. Both enzymes are increased in SQ-A virions. The activity of protein kinase in SQ-A virions is about 5 times higher than that in the attenuated virions. The number of polypeptides and their phosphorylation patterns also distinguish the virions of SQ-A. Whereas 5-86 virions contain seven proteins, three of which are phosphorylated in vitro, SQ-A virions contain eight proteins, all of which are phosphorylated. The extra protein in SQ-A virions has a molecular weight of 25,000 and is not glycosylated.
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PMID:Characterization of leukemogenic virus produced by a new line of Friend erythroleukemia virus-transformed cells. 632 17

Cultures of murine Friend erythroleukemia (FL) cells, which are chronically infected with leukemia virus, were inoculated with vaccinia virus. The yield of vaccinia virus was determined by assaying plaque-forming units in mouse L2 cells, and the yield of leukemia virus was determined by measuring reverse transcriptase (RNA-dependent DNA nucleotidyltransferase) activity released into the culture fluid. Although no facilitation of one virus by the other was detected, persistently infected cultures were established. Electron microscopic examination revealed the presence of vaccinia and leukemia viruses in the same cell. The permanent lines of cells persistently infected with vaccinia were designated FLvac. Their morphology, growth rate, cloning efficiency, and ability to respond to the induction of erythrodifferentiation by treatment with dimethyl sulfoxide were not appreciably altered as compared to the parental FL cells. However, the persistently infected cells showed a marked decrease in tumorigenicity when assayed in DBA/2 mice. The infectious virus produced by FLvac cells and by L2 cells were indistinguishable as judged by restriction endonuclease patterns of virion DNA, structural proteins, and the activities of two virion-associated DNases. The yield of infectious vaccinia virus from FLvac cells generally declined after about 60 serial passages. Although some cell lines no longer yield infectious virus, they are resistant to challenge with vaccinia at concentrations that are cytolytic for L2 cells. The mechanism responsible for the establishment of the persistent infection remains unclear because defective particles, interferon production, and temperature-sensitive mutants have not been detected.
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PMID:Persistent infection of Friend erythroleukemia cells with vaccinia virus. 695 93


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