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)

An RNA-directed DNA polymerase was found to be associated with intracytoplasmic A-particles from DBA/2 mouse leukemia cells. The enzyme activity was detected after disrupting the purified particles with 2 M NaCl-20 mM dithiothreitol. The presence of a divalent cation and all four deoxyribonucleoside triphosphates was essential for this enzyme activity. The enzyme had a clear preference for Mg2+ over Mn2+. Cesium sulfate isopycnic gradient centrifugation of the DNA product synthesized in the actinomycin D-containing reaction revealed the presence of DNA-RNA hybrid. Furthermore, the purified DNA product was found to hybridize with RNA isolated from A-particles. These observations strongly indicate that the endogenous A-particle RNA serves as the template for the DNA polymerase.
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PMID:Characterization of an RNA-directed DNA polymerase found in association with murine intracytoplasmic A-particles. 6 24

Strains of Friend leukemia virus (FLV) that are associated with polycythemia contain the defective spleen focus-forming virus (SFFV). To determine whether the transforming ability of FLV was affected by the presence of this second agent, DBA/2J mouse bone marrow cells were infected in vitro. Criteria for transformation were the establishment of permanent lines, growth on semisolid agarose, and the production of tumors at the site of inoculation in syngeneic hosts. Two lines of immature hematopoietic cells that grow in suspension originated from the infected cultures. Each has an almost diploid karyotype (38-39 chromosomes) and 3-4 metacentric chromosomes. These transformed cells express gp71 viral envelope glycoprotein and p30 viral core protein antigens. Virus production was measured by reverse transcriptase (RNA-dependent DNA polymerase) activity of the virions released into the medium. The virus, assayed in vivo for infectivity, has SFFV activity but is attenuated for leukemogenicity. The stimulation of hemoglobin synthesis in the cells grown in medium supplemented with dimethyl sulfoxide or hexamethylene bisacetamide indicates that the cells are erythroid in origin. SFFV may have a function analogous to erythropoietin in influencing the process of transformation by FLV.
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PMID:In vitro transformation of mouse bone marrow cells by the polycythemic strain of Friend leukemia virus. 8 91

Delineation of major T helper cell recognition sites of human immunodeficiency virus (HIV-1) proteins represents one important step in the design of an efficient acquired immune deficiency syndrome (AIDS) vaccine. Towards this end, we have explored the immunogenicity of HIV-1BRU proteins in the mouse model. Preliminary experiments revealed that inbred mice primed with whole inactivated HIV-1 developed strong CD4+ T cell proliferative responses to a variety of recombinant viral proteins including reverse transcriptase (RT). To characterize further the mouse T cell responses to this protein, several Ad- or Ed-restricted T hybridoma cells (THC) were established from BALB/c or DBA/2 mice. These THC were tested for their capacity to recognize a series of 15-mer synthetic overlapping peptides spanning three segments of HIV-1 RT that had been preselected on the basis of either alpha-helicity, amphipaticity, and/or for containing rare amino acid sequence patterns. Peptides corresponding to a C-terminal region (residues 528-560) of RT were recognized by several of the THC established from RT-primed mice. Furthermore, a non-alpha-helical peptide from this region (A3, 528-543) was capable of priming mice with different H-2 haplotypes for both peptide A3 and native RT CD4+ T cell recognition. In addition to the recently identified RT determinant 203-219 capable of triggering both mouse and human CD8+ CTL, the present results identify a good candidate for an immunodominant RT epitope capable of eliciting RT-specific T helper cell responses.
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PMID:Identification of a major human immunodeficiency virus-1 reverse transcriptase epitope recognized by mouse CD4+ T lymphocytes. 171 May 63

The characteristics of tumor antigens of the transplantation type, TATA, were studied in a series of methylcholanthrene-induced sarcomas in pedigreed BALB/cAN mice. Each of the six sarcomas exhibited unique TATA when assayed for tumor rejection in syngeneic hosts. In three instances, however, in sarcomas CI-I, CII-5 and CII-7, TATA activity was lost during early in vivo passages. This activity was reestablished invivo from the in vitro-passaged lines in CI-I and CII-7 and these sarcomas along with CI-3, CI-4 and CII-10 maintained a stable TATA phenotype throughout the 50 transplant passages; TATA was lost permanently, however, in CII-5. These results indicate a dimorphic nature of early-passage neoplasms. No cross-reacting antigens were detected among these sarcomas, including also Meth A, a sarcoma well characterized as to its membrane antigens; nor was any evidence obtained for the existence of alien (inappropriate) H-2 antigens employing tumor rejection assays in F1 hybrids of BALB/c with strains bearing several haplotypes and also with studies of H-2 serology. In three sarcomas, some evidence was obtained for the existence of alloantigens of the non-H-2 type since it was found that (BALB/c X DBA/2)F1 hybrids could not be immunized by respective sarcomas. These findings suggest the existence on these sarcomas of a tumor antigen that is expressed normally in DBA/2 mice, although no definitive evidence has been obtained. Assays for MuLV, for the viral structural proteins gp70 and p30 and for reverse transcriptase showed that three sarcomas were positive and three others, as well as Meth A, were negative. MuLV and its antigens had no influence on tumor rejection activity nor could the cross-reactivity observed in the radioisotopic footpad assay (IFP) be related to MuLV.
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PMID:Changes in tumor-specific antigen expression during passage in vitro and in vivo of newly derived methylcholanthrene-induced sarcomas of BALB/C mice. 615 30

The properties of the virus synthesized by each of three morphologically different cell lines originating from DBA/2J fetal liver cells transformed by the anemic strain of Friend leukemia virus in vitro were analyzed. The cells of line G-1 are malignant in syngeneic DBA/2 mice, grow in suspension, and are erythroid in origin. Cells of lines G-2 and G-3 are adherent, are epithelial in appearance, and produce no tumors in DBA/2J mice. Higher reverse transcriptase activity was detected in the culture fluid of lines G-2 and G-3, although virus from G-1 cells was more leukemogenic. Differences were also found in the virion density and size of the viral genome. RNA from the virions produced by G-2 and G-3 cells sedimented at 75 S in a sucrose gradient; virion RNA from G-1 cells sedmiented at 60 S. However, when subjected to polyacrylamide gel electrophoresis, all three virus strains showed identical RNA subunits with an estimated molecular weight of 2.6 X 10(6). Analysis of virion proteins by slab gel electrophoresis showed differences in envelope protein (gp71) components but not in the major core protein (p30). The properties of these viruses are stable and remain unchanged after passage in 3T3 cells.
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PMID:Variations in properties of virus released from morphologically different cell lines transformed in vitro by Friend leukemia virus. 616 May 80

The biological properties of the virus synthesized by 18 clones of a line of mouse bone-marrow hematopoietic cells transformed in vitro by the polycythemic strain of Friend leukemia virus (FLV-P) were compared. In vitro assays were performed to determine whether the virus released into the culture fluids was ecotropic or xenotropic, and in vivo assays were carried out to determine spleen focus formation and leukemogenicity in susceptible DBA/2J and BALB/c mice. A number of clones released virus which reproduced the entire range of effects typical of the FLY-P complex. However, in other clones, there appeared to be no correlation between the assays for leukemogenicity and the assays for either ecotropic virus, reverse transcriptase activity, or virus antigens. Xenotropic virus was not detected in any of the cultures. These results suggest that the FLV-P complex contains a heterogeneous population of viruses, but the possibility that the differences observed may be due to the inability of FLV-P to be expressed fully in some clones cannot be excluded.
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PMID:Lack of correlation between in vivo and in vitro assays for the detection of virus released from clones of Friend erythroleukemia cells. 616 81

Using partially purified mouse liver 23S mRNA known to be associated with the Ah locus and 3-methylcholanthrene-induced cytochrome P(1)-450, we synthesized double-stranded cDNA by the successive action of reverse transcriptase (RNA-directed DNA nucleotidyltransferase) and the Klenow A fragment of Escherichia coli DNA polymerase I. The double-stranded cDNA was inserted into pBR322 plasmid DNA by Pst I cleavage and homopolymeric "tailing" and cloned in E. coli LE392. Clone 46 hybridized with [(32)P]cDNA made from 23S mRNA from "Ah-responsive" C57BL/6N mice but did not hybridize with similarly prepared [(32)P]cDNA from "Ah-nonresponsive" DBA/2N mice. Clone 30 was positive, and clone 7 was negative, with both C57BL/6N and DBA/2N [(32)P]cDNA probes; these two clones were therefore used as "positive" and "negative" control clones, respectively. By translation-arrest experiments, clone 46 DNA and clone 30 DNA were shown to be associated with anti-P(1)-450- and anti-albumin-precipitable material, respectively. By agarose gel electrophoresis of Pst I digests, the clone 46 DNA insert was shown to be 1100 base pairs in total length and to contain one internal Pst I site. The cDNA made from total mRNA isolated from 3-methylcholanthrene-treated C57BL/6N mice hybridized to the two fragments of Pst I-digested DNA from clone 46, whereas similarly prepared cDNA from 3-methylcholanthrene-treated DBA/2N and control C57BL/6N and DBA/2N mice did not. Of 11 restriction endonucleases used, two (Pst I and Xba I) had sites within the clone 46 DNA insert. After hybridization of clone 46 (32)P-labeled nick-translated DNA to EcoRI fragments from A/HeJ mouse genomic DNA and fractionation by RPC-5 chromatography and gel electrophoresis, only one positive band (3-4 kilobase pairs appeared. These data demonstrate conclusively that pBR322 clone 46 DNA is associated with mRNA controlled by the murine Ah locus, presumably the structural gene encoding 3-methylcholanthrene-induced P(1)-450.
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PMID:Isolation and characterization of a cloned DNA sequence associated with the murine Ah locus and a 3-methylcholanthrene-induced form of cytochrome P-450. 626 72

The Myeloproliferative Sarcoma Virus (MPSV) induces an increase in the number and concentration of pluripotent stem cells in long-term murine bone marrow cultures. This is followed by an increased number of precursor cells of the granulocyte and macrophage lines (GM-CFC). This increase is comparable to that observed in DBA/2 mouse spleens in vivo two to three weeks after viral infection. Proliferation of CFUs and GM-CFC decreases five weeks after infection with MPSV, in parallel to the gradual decline of reverse transcriptase activity in the culture medium. GM-CFC which can proliferate in the absence of added colony stimulating factor (CSF) were detected at week 6 post MPSV infection. Adherent tumor cells were observed nine weeks after infection. These fibroblast type cells gave rise to a permanent line which produced a CSF-like activity. Our results show that MPSV causes the tumoral transformation of fibroblast-like cells of the bone-marrow hematopoietic microenvironment. In addition, MPSV also strongly stimulates the proliferation of hematopoietic stem cells. MPSV is, until now, the first murine retrovirus which exhibits such properties.
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PMID:Myeloproliferative Sarcoma Virus stimulates pluripotent hematopoietic stem cells and provokes tumoral transformation of the hematopoietic microenvironment in vitro. 630 May 64

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

Antibodies raised in several mouse and rat strains against the hapten 2-phenyloxazolone (phOx, "oxazolone") regularly contain a fraction recognized by antiidiotypic reagents. We have studied this response in BALB/c and DBA/2 mice by generating over fifty anti-phOx antibody-secreting hybridoma clones. The hybridization was performed either 7 or 14 days after a primary immunization with phOx-protein conjugate. Most of the hybrids secreted IgG1. Whereas over 80% (17/21) of IgG-producing hybrids from day-7 fusions secreted oxazolone-idiotype positive immunoglobulin, all hybridomas originating from day-14 fusions were idiotype negative. The mRNA for heavy (H) and light (L) chains of three idiotype-positive and one idiotype-negative IgG1 hybridomas were sequenced by a modification of Sanger's dideoxynucleotide method of DNA sequencing, using crude mRNA as template, synthetic oligonucleotides as primers, and reverse transcriptase to incorporate both dideoxynucleotides and labeled deoxynucleotides. The sequence of the mRNA coding for the whole variable region of each chain was established using primers complementary to the constant region near the V-C boundary and another two that coded for a framework segment in either VH or VL. This method not only provided more information than protein sequencing but was also faster and simpler. The mRNA preparation did not need fractionation beyond the poly A-containing fraction. The sequences of the H and L chain mRNA of the three idiotype-positive anti-oxazolone antibodies were extremely similar or identical, and from them a tentative oxazolone-idiotype basic sequence was derived. Only three nucleotide differences were detected; these occurred in the D segment of one H chain mRNA, in the V-J boundary of one of the light chain mRNA, and in the first hypervariable region of another. The idiotype-negative antibody had a totally different H chain mRNA and a light chain mRNA that differed by 21 bases, almost all affecting the amino acid sequence.
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PMID:Anti-oxazolone hybridomas and the structure of the oxazolone idiotype. 640 10


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