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)

Cell banks were created by controlled rate freezing of pooled cell suspensions. Each line was characterized for freedom from adventitious agents, identity and biological behavior in immunosuppressed animals. Pharmacopeal culture methods were employed to search for bacterial, fungal and mycoplasma contamination. Conditioned medium was inoculated into a battery of 6 cell lines to search for acute viral contamination. Recombinant cell lines were also examined for retroviruses by electron microscopy, reverse transcriptase assay, XC plaque assay, tritiated uridine labelling, chemical induction techniques, and co-cultivation with three indicator cell lines. Further characterization was obtained from inoculation of 4 animal species in vivo, mouse antibody production test, and inoculation of immunosuppressed rodents. Finally, cytogenetic evaluation including banding methods and isoenzyme analysis were employed to verify cell bank identity. These methods have generated useful data concerning the identity, safety, and biological purity of recombinant DNA mammalian cell lines.
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PMID:Cell bank characterization for recombinant DNA mammalian cell lines. 404 32

A late-passage derivative of SC-1 cells with stable relative resistance to N- and B-tropic viruses but with retained sensitivity to NB-tropic viruses has been identified. Dual resistance was seen with the XC plaque assay, as well as reverse transcriptase assay and radioimmunoassay for viral p30. Evidence was found for resistance in clonal derivatives. These findings suggest a possible instability in determinants of cellular resistance to endogenous retroviruses and may be useful in further analysis of resistance to retrovirus infection.
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PMID:Late-passage SC-1 cells resistant to N- and B-tropic viruses but sensitive to NB-tropic viruses. 615 76

A persistent infection by human coronavirus 229E (HCV/229E) was established in a human continuous cell line (L132). Following the initial infection with stock HCV/229E, several cultures were established of which two (HV1 and HV4) have been maintained by continuous passage for two years. These cultures have shed high titres of infectious virus continuously into the supernatant fluid since their initiation. The persistently infected cells were resistant to homologous super-infection but supported polio virus replication to normal titres. Preliminary tests indicated that 50-100 percent of the cells contain virus. Neither interferon nor reverse transcriptase could be detected in these cultures and the presence of defective interfering particles could not be demonstrated. VH1 and VH4 coronaviruses, isolated from these persistently infected cultures (HV) and identified by 229E antiserum neutralization, were more cytocidal than the parent virus as judged by plaque characteristics and CPE, however they were indistinguishable on the basis of density, EM morphology, and genome size. Present evidence indicated that temperature plays an important but as yet undetermined role in the establishment and maintenance of stable 229E persistently infected cell cultures.
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PMID:Establishment and maintenance of a persistent infection of L132 cells by human coronavirus strain 229E. 617 Dec 37

We have constructed a series of deletion mutations in the p30 and p10 domains of the gag gene of Moloney murine leukemia virus. Mutants with deletions in P30 were completely defective in virion particle production even though an altered gag precursor protein is synthesized. This domain is apparently critical for particle formation. A mutant in P10 was able to release virion particles into the medium, and low levels of reverse transcriptase activity could be detected in these virions. To explore the effects of these mutations on the utilization of the gag-pol precursor, we have introduced these mutants into cells already releasing defective particles from an endogenous provirus which directs the synthesis of gag gene products and not pol gene products. The P10 mutant was capable of providing pol function as judged by the incorporation of high levels of reverse transcriptase into the particles and complete complementation for XC plaque formation. In contrast, the mutants in P30 were negative in this complementation test. Thus, those gag mutants which were unable on their own to assemble virion particles were also unable to contribute the gag-pol precursor to these particles. These mutations are the first to be mapped to the gag region which affect pol function, suggesting that the gag-pol precursor must be assembled before pol is functionally separated from the gag domain. The concordance of the effects of different mutations on both particle formation and gag-pol utilization suggests that similar domains of gag (namely, domains in the P30 region) are needed for these two processes.
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PMID:Mutations in the gag gene of Moloney murine leukemia virus: effects on production of virions and reverse transcriptase. 619 13

The integrated proviral genome of Rauscher murine leukemia virus was molecularly cloned in a bacteriophage Charon 4A vector after the proviral sequences were enriched by sequential RPC-5 column chromatography and sucrose gradient centrifugation. A recombinant DNA clone, lambda-RV-1, possessing a 12-kilobase-pair EcoRI insert, was shown to contain the entire 8.8-kilobase-pair leukemia virus genome flanked by rat cellular sequences at the 5' and 3' ends. This DNA fragment was biologically active, inducing the release of virion-associated reverse transcriptase activity with as little as 10 ng of DNA insert. The virus induced XC plaque formation at high titers on NIH/3T3 and BALB/3T3 cells and demonstrated identity with the parental virus in radioimmunoassays for the highly type-specific gag gene-coded p12 protein. The molecularly cloned Rauscher murine leukemia virus should be useful in studying the molecular mechanisms involved in the transformation of specific lymphoid target cells by chronic mouse leukemia viruses.
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PMID:Rauscher murine leukemia virus: molecular cloning of infectious integrated proviral DNA. 629 29

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

Synchronized mouse cells (JLS-V9) chronically infected with Rauscher murine leukemia virus were used to study virus production, the synthesis of gag and env precursor proteins, and the expression of env protein on the cell surface during the cell cycle. The amount of virus released into the medium by synchronized cells during a 30-min interval was determined by using the XC plaque assay and by measuring reverse transcriptase activity. The results show that virus production occurs during mitosis. Labeling of the cell surface of synchronized cells with 125I or with fluorescein-conjugated antiserum shows that the amount of gp 70env on the cell surface parallels cellular growth. Therefore, the cell cycle-dependent release of virus is not accompanied by similar variations in the amount of viral envelope protein on the cell surface. Immunoprecipitation of cells labeled with [35S]methionine, followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, was used to measure viral protein synthesis during the cell cycle. The rate of synthesis of gag precursor proteins show three maximums corresponding to the G1, middle S, and late S to G2 phases of the cell cycle. The rate of synthesis of env precursor proteins does not change, suggesting that in these cells the synthesis of these two gene products is controlled separately.
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PMID:Retrovirus gene expression during the cell cycle. I. Virus production, synthesis, and expression of viral proteins in Rauscher murine leukemia virus-infected mouse cells. 728 18

The replication of foot and mouth disease virus (FMDV) was studied in isolated bovine skin Langerhans cells (LC), in keratinocytes from epidermal cell suspension, and in migrating LC obtained from cultured bovine epidermal sheets in vitro. Viral RNA replication in infected cells was determined by the reverse transcriptase-polymerase chain reaction (RT-PCR) of the negative FMDV RNA strand and by the plaque forming assay of FMDV. It was established that bovine skin LC, keratinocytes, and migratory bovine LC infected with FMDV strain 01 Geshur supported virus replication. This RT-PCR method to detect the negative strand of FMDV RNA in migratory bovine skin LC may be useful for determining FMD virus replication in tissue cells.
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PMID:Foot and mouth disease virus replication in bovine skin Langerhans cells under in vitro conditions detected by RT-PCR. 748 89

We have selected and plaque purified zidovudine (3'-azido-3'-deoxythymidine [AZT])-resistant mutants from an infectious molecular clone of feline immunodeficiency virus (FIV). The patterns of cross-resistance and drug susceptibilities of these mutants were similar to those of the AZT-resistant FIV that we previously selected in vitro from a wild-type FIV population and to those of the most common AZT-resistant clinical isolates of human immunodeficiency virus type 1. Two AZT-resistant mutants of FIV, one selected from a normal population and one selected from the molecular clone, each reverted rapidly to an AZT-sensitive phenotype when passaged in the absence of drug. Sequence analysis of the reverse transcriptase (RT)-encoding region from the plaque-purified AZT-resistant FIV revealed a single base change at position 2939, resulting in a Glu-to-Lys substitution at amino acid 202 of the RT. Similar analyses of plaque-purified revertants showed that the phenotypic reversion was not the result of a genotypic reversion at this position and that no additional mutations existed within the RT-encoding region of the revertants. Moreover, RTs purified from the mutant and revertant were both resistant to the 5'-triphosphate of AZT. These results indicate the complexity of AZT resistance and suggest the presence of additional factors, outside the RT-encoding region, which may contribute to AZT resistance.
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PMID:Rapid phenotypic reversion of zidovudine-resistant feline immunodeficiency virus without loss of drug-resistant reverse transcriptase. 750 82

R82913 and R86183, two derivatives of tetrahydroimidazo[4,5,1-jk][1,4]-benzodiazepin-2(1H)-thione (TIBO), were found to potently and selectively inhibit the replication and cell killing effects of a panel of biologically diverse laboratory and clinical strains of HIV-1. The two compounds exhibited significant activity in all human cell lines tested, as well as in fresh human peripheral blood lymphocytes and macrophages. One of these two compounds (R82913) was found to significantly inhibit the replication of a murine retrovirus (Rauscher murine leukemia virus) in both UV-XC plaque formation and virus yield reduction assays. R86183, despite differing from R82913 only in the positioning of a single chlorine molecule, was not active against the murine retrovirus but was 10-fold more potent in inhibiting HIV-1 replication. Combination antiviral assays with other reverse transcriptase inhibitors, including AZT, ddC, and carbovir, yielded synergistic anti-HIV activity with both TIBO derivatives. Additive to slightly synergistic results were obtained in combinations with ddI and phosphonoformic acid whereas additive to antagonistic activity was detected in combination with dextran sulfate.
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PMID:Differential antiviral activity of two TIBO derivatives against the human immunodeficiency and murine leukemia viruses alone and in combination with other anti-HIV agents. 750 23


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