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)

Friend virus (FV) is a murine leukemia virus that infects progenitor red blood cells and causes an erythroleukemia in susceptible mouse strains, resulting in splenomegaly. Several genetic loci of the host have been identified that affect erythroleukemia development, differentiation status of target cells and virus replication. Since age may change expression of these loci, age may affect FV disease. To explore this possibility, FV expression in four genetically diverse strains of mice of different ages was examined. Extent of viral replication and of disease were evaluated by measuring spleen focus forming units (SFFU), spleen weight and reverse transcriptase (RT) activity in target organs. Young DBA/2 and (C57BL/6 x DBA/2)F1 mice exhibited a greater level of virus expression than their aged counterparts in all parameters investigated. Young CBA/Ca mice had slightly higher spleen weights and SFFU values than aged CBA/Ca mice, but a definitive age-related change was not observed in the RT activity of the target organs. C57BL/6 mice, which are genetically resistant to the development of FV-induced erythroleukemia, exhibited a limited degree of virus replication that was not effected by the age of the animal. Our results indicate that the age of the mouse, as well as the genetic background, can contribute to the level of susceptibility to FV.
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PMID:Differences between young and aged mice in susceptibility to Friend virus. 751 1

A 29 kD protein previously isolated from murine erythroleukemia (MEL) cells and shown to enhance the rate of differentiation of these cells has now been demonstrated to possess an amino acid sequence identical to that reported for the 30 kD heparin-binding protein from developing rat brain, named amphoterin after its highly dipolar structure. The identity between the two proteins has been established on the basis of a strong heparin binding affinity and a complete homology in the amino acid sequences of N-terminal region as well as of several tryptic peptides. Furthermore, the cDNA encoding this protein has been isolated from MEL cell mRNA, by means of reverse transcriptase-polymerase chain reaction, and its sequence was found to correspond to that of amphoterin. The MEL cell differentiation enhancing factor, previously abbreviated as DEF, is again confirmed to reduce the latent period preceding the appearance of hexamethylenebisacetamide induced cell commitment and to stimulate the catalytic activity of alpha-protein kinase C. Thus, here we demonstrate that a protein expressed in MEL cells, whose sequence is identical to that previously reported for amphoterin, plays an essential role in promoting cell differentiation, thereby indicating a new relevant function of amphoterin.
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PMID:Identity in molecular structure between "differentiation enhancing factor" of murine erythroleukemia cells and the 30 kD heparin-binding protein of developing rat brain. 774 53

The presence of alpha, delta, epsilon, theta, and zeta protein kinase C isoforms in DS19 murine erythroleukemia cells has been established in this study. In addition, the mRNA levels of these isozymes have been measured by quantitative reverse transcriptase-polymerase chain reaction. Isoform delta has been found to be the most abundant isotype, whereas isoform zeta resulted to be present in only few copies. Furthermore, the expression levels of all five protein kinase C isozymes have been studied in three cell clones, derived from parental DS19 cells and characterized by different susceptibilities to differentiation. This comparative analysis indicated that the calcium-independent isozymes (delta, epsilon, zeta, and theta) display significantly higher expression levels in cells less prone to differentiation. On the other hand, the mRNA levels of the only calcium-dependent isoform present (alpha) fluctuate poorly from one cell clone to the other, but are the highest in the cell clone characterized by the fastest rate of differentiation. This study represents the first complete characterization of the basal levels of specific protein kinase C isotypes in different murine erythroleukemia cell clones and provides further evidence for the role of individual isozymes in the early events that trigger chemical induced murine erithroleukemia cell differentiation.
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PMID:Differential expression of protein kinase C isoform genes in three murine erythroleukemia cell variants: implication for chemical induced differentiation. 798 May 1

A cis-acting DNA regulatory element 3' to the A gamma-globin gene contains eight distinct regions of DNA-protein interaction distributed over 750 bp of DNA. The sequences of two foot-printed regions (sites I and IV) are A-T rich and generate a highly retarded complex on gel shift analysis with nuclear extract from human erythroleukemia (K562) cells. We have purified a 98-kD protein that reproduces this gel shift. Tryptic cleavage and peptide sequence analysis demonstrated that the 98-kD protein is identical to a recently cloned protein, special A-T-rich binding protein 1 (SATB1), that binds selectively to nuclear matrix/scaffold-associated regions of DNA (MARs/SARs). We have shown by functional analysis that the 3' A gamma regulatory element associates with the nuclear matrix. SATB1 mRNA was identified in K562 cells, and reverse transcriptase-polymerase chain reaction (RT-PCR) demonstrated its transcript in several other hematopoietic lines. Antisera to SATB1 caused ablation of the gel shift complex generated by both the crude nuclear extract and the purified 98-kD protein with the site I oligonucleotide. Furthermore, oligonucleotides that bind SATB1 inhibited formation of the site I gel shift complex when added as excess unlabeled competitor. An immunoblot analysis of the site I gel shift complex documented the presence of SATB1. Binding of SATB1 to two sites within the 3' A gamma regulatory element and its MAR/SAR activity suggests that this element may influence gene expression through interaction with the nuclear matrix.
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PMID:The regulatory element 3' to the A gamma-globin gene binds to the nuclear matrix and interacts with special A-T-rich binding protein 1 (SATB1), an SAR/MAR-associating region DNA binding protein. 804 44

Thromboxane A2, a potent platelet agonist and vasoconstrictor, exerts its actions via specific G protein-coupled receptors. cDNAs encoding the full length thromboxane receptor have been isolated from human placenta mRNA by reverse transcriptase-polymerase chain reaction. An expression construct, under control of the cytomegalovirus promoter, was introduced into human embryonic kidney 293 cells. Membranes from transfected cells bound the thromboxane antagonist SQ29,548 and the agonist [15-(1 alpha,2 beta(5z)-3 alpha(1E,3S)-4 alpha)]-7-[3-(3-hydroxy-4-(p- iodophenoxy)-1-butenyl)-7-oxabicyclo[2,2,1]hept-2-yl]-5-heptenoic acid) with high affinities, and significantly more receptors were expressed in these cells, compared with platelet preparations. The putative seventh transmembrane segment is highly related in all cloned members of the eicosanoid receptor family and forms a critical portion of the ligand binding pocket for G protein-coupled receptors. Several point mutations in this segment were generated. Binding of SQ29,548 was virtually abolished in cells transfected with all the variant receptor constructs. However, one receptor variant (TxR-W299L), in which a tryptophan at position 299 was substituted for a leucine residue, allowed a definite discrimination between agonist and antagonist binding sites in competition and saturation binding experiments. An antibody directed toward the third intracellular loop of the thromboxane receptor was able to immunoprecipitate native thromboxane receptor in solubilized membranes from human erythroleukemia cells and transfected cells.
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PMID:Point mutation in the seventh hydrophobic domain of the human thromboxane A2 receptor allows discrimination between agonist and antagonist binding sites. 824 16

The 12-lipoxygenase (LO) pathway of arachidonic acid plays an important role in angiotensin II (AII)-mediated aldosterone synthesis. Several distinct isoforms of 12-LO have been cloned. However, in humans only the platelet form of 12-LO has been reported to be present. Western immunoblotting analysis in cultured human adrenal glomerulosa cells using polyclonal antibodies to porcine leukocyte 12-LO enzyme or peptide showed a specific 72-kilodalton band, which is identical to the molecular size of the porcine leukocyte form of 12-LO. In addition, AII (10(-7)) increased the intensity of the 72-kilodalton band nearly 2-fold over basal. In situ hybridization analysis indicated a strong positive reaction with the porcine leukocyte type of 12-LO antisense riboprobe in the zona glomerulosa of the adrenal cortex. The 12-LO probe also recognized a near 4-kilobase messenger RNA (mRNA) from human glomerulosa cells in Northern blots. Since the leukocyte type of 12-LO is highly homologous to human 15-LO, a reverse transcriptase and polymerase chain reaction was used to analyze the specific type of 12-LO mRNA in human cells. The mRNA for a porcine leukocyte type of 12-LO was detected in human adrenal glomerulosa cells, and the level of 12-LO transcripts was increased approximately 60-fold by AII (10(-7) M). The leukocyte type of 12-LO also was detected in human monocyte-like U937 cells, but not in IM-9 lymphocytes or human erythroleukemia cells. These results suggest that human adrenal glomerulosa cells and human monocyte-like U937 cells express a 12-LO which has immunological and molecular biological characteristics similar to the porcine leukocyte 12-LO.
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PMID:Evidence that a leukocyte type of 12-lipoxygenase is expressed and regulated by angiotensin II in human adrenal glomerulosa cells. 827 71

The p53 gene is currently thought to be a tumor suppressor gene, and its alterations have been suggested to be involved in the pathogenesis of several human malignancies, including some leukemias and lymphomas. We present here evidence for the possible involvement of p53 gene mutations in the myelodysplastic syndrome (MDS), although the incidence is relatively low. Forty-four patients with MDS and six patients with overt leukemias that developed from MDS were studied for p53 gene alterations using reverse transcriptase-polymerase chain reaction, single-strand conformation polymorphism analysis, and nucleotide sequencing. Three patients with MDS (2 RAEB and 1 RAEB in T) had missense point mutations in the conserved regions of the p53 coding sequence. Furthermore, expression of the wild-type p53 mRNA was not detected in these three patients. The probable absence of normal p53 function in the three cases studied here suggests that alterations in the p53 gene may occasionally play a role in MDS. These three MDS patients with p53 gene mutations and an MDS-derived erythroleukemia cell line that we had previously reported to carry a p53 gene mutation showed no N-ras gene mutations, suggesting heterogeneity in the oncogenic mechanism of MDS.
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PMID:Mutations of the p53 gene in myelodysplastic syndrome (MDS) and MDS-derived leukemia. 849 37

Differentiation inhibitory factor (nm23 protein) inhibited the induction of differentiation of mouse myeloid leukemia M1 and WEHI-3BD+ and human erythroleukemia HEL, KU812, and K562 cells. Block of differentiation may be associated with the aggressive behavior of leukemia. To examine the role of nm23 in human myeloid leukemia, we investigated the relative levels of nm23-H1, nm23-H2, and c-myc transcripts in 42 patients with acute myelogenous leukemia (AML), and in 5 with chronic myelogenous leukemia at chronic phase by reverse transcriptase polymerase chain reaction. The expression of nm23-H1 and -H2 but not of c-myc in AML was significantly higher than that in normal blood cells. Among AMLs, acute monocytic leukemia (presentation with AML-M5 morphology) was especially associated with elevated nm23-H1 and -H2 mRNA levels. On the other hand, the elevated levels of c-myc expression in AML-M5 were less evident. An analysis of correlation between nm23 expression and clinicopathological parameters showed that resistance to initial chemotherapy is associated with increased nm23-H1 mRNA levels and that a high initial white blood cell count is associated with increased nm23-H2 mRNA levels. Elevated nm23-H1 mRNA levels were associated with significantly reduced the overall survival of AML, especially of AML-M5 patients. The present results indicate that nm23-H1 and -H2 are overexpressed in AML and especially nm23-H1 gene expression predicts the prognosis of AML, especially of AML-M5.
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PMID:Differentiation inhibitory factor nm23 as a new prognostic factor in acute monocytic leukemia. 889 23

Rauscher murine leukemia virus induces an erythroleukemia in susceptible strains of mice that is associated with splenomegaly and viremia. This animal model has been used for evaluating the in vivo efficacy of potential anti-HIV agents. The in vivo antiviral activity of therapeutic agents has usually been determined by measuring a reduction in the spleen weights of compound-treated mice or by quantitating viremia with the UV-XC plaque assay. The UV-XC assay, however, is time-consuming and labor-intensive. Virions of Rauscher murine leukemia virus, like other retroviruses, contain the enzyme reverse transcriptase. Quantitating the level of this enzyme in infected mouse sera provides a more rapid measure of viremia in the animal. We have examined the effects of several reagents, including detergent, KCl, EGTA, dGMP, spermine, as well as protease and RNase inhibitors, on the reverse transcriptase assay. The optimized assay method was effective in evaluating the antiviral activity of AZT in the Rauscher murine leukemia virus in vivo model. The assay is also amenable to automation if large numbers of assays are required.
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PMID:Optimization of the reverse transcriptase assay for the detection of viral burden in mice infected with Rauscher murine leukemia virus. 891 Jun 49

Platelet basic protein (PBP) is a chemokine family member that is only found in platelets and their precursors megakaryocytes. The PBP gene is physically linked to the gene for another platelet-specific chemokine, platelet factor 4. While the biological basis of platelet factor 4 expression has been pursued by others, the regulatory features controlling the platelet-specific expression of PBP have not been investigated. In this article, we examined the molecular basis by which this megakaryocyte-specific gene is regulated. Transient expression studies of truncated reporter constructs containing from 4.5 to 0.1 kilobases of the functional PBP gene 5'-flanking region, demonstrated that the proximal 0.1 kilobases of the promoter was sufficient for high levels of expression in human erythroleukemia and CHRF-288 cells, two megakaryocytic cell lines. However, none of these constructs was expressed above background levels in HeLa and 293 cells, two non-megakaryocytic cell lines. Further truncation of this promoter suggested that there was an important regulatory element(s) within a pyrimidine-rich tract. Mobility shift analysis of the pyrimidine-rich tract defined a region between -85 and -64 which bound to a nuclear factor(s). This region contains sequences matching the consensus Ets-binding site from -78 to -75 base pairs. In particular, we noted that this site matched a PU.1 consensus sequence known as a PU box. Mobility shift and supershift studies with nuclear extracts as well as recombinant PU.1 protein and anti-PU.1 antibody further confirmed that PU.1 was the specific Ets family factor that bound to this site. Transient expression assays using reporter constructs which contained point mutations that abrogated PU.1 binding also significantly reduced PBP promoter activity in human erythroleukemia and CHRF cells. In addition, while all reporter gene constructs containing PBP promoters were completely inactive in HeLa cells, transactivation experiments using a PU.1 expression construct demonstrated that exogenous expression of PU.1 could increase reporter gene expression up to 8-fold in these cells. Finally, the role of PU.1 in PBP gene expression was compared between wild-type and PU.1-null embryonic stem (ES) cells that were differentiated in vitro into cells that resembled megakaryocytes both morphologically and immunologically. We found that PBP gene expression in the differentiated PU.1(-/-) null ES cells (as determined by semi-quantitative reverse transcriptase-polymerase chain reaction) was more than four times lower than that in the wild-type ES cells, while other platelet-specific genes were expressed equally or similarly in the two ES cell lines. Previous reports have shown that PU.1 is expressed in several hematopoietic lineages, including megakaryocytes. However, the functional role of PU.1 has only been previously demonstrated in the myeloid and lymphoid lineages. Therefore, our studies are the first to show the biological importance of this nuclear factor in the regulated expression of a megakaryocyte-specific gene.
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PMID:Activation of the megakaryocyte-specific gene platelet basic protein (PBP) by the Ets family factor PU.1. 933 92


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