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:3.1.27.5 (RNase)
17,967 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The binding of [3H]dexamethasone to the cytosol fraction prepared from the human leukemia cell line K562 was studied with a competitive binding assay. Specific, saturable binding was identified by incubating cytosol with increasing concentrations of [3H]dexamethasone in the presence and absence of nonlabeled dexamethasone. A Scatchard plot of the data was linear, suggesting the presence of a single class of binding sites with KD = 2.49 +/- 0.23 x 10(-8)M. The binding sites appear to be protein in nature, since specific binding was reduced by treatment of the cytosol with trypsin, pronase, and heat; neither DNase nor RNase affected the binding. Binding was also reduced in the absence of alpha-thioglycerol and in the presence of p-chloromercuribenzoate and N-ethylamaleimide, suggesting that optimal binding activity requires reduced sulfhydryl groups. The binding site appears to be specific for glucocorticoids as evaluated in competition studies. Finally, glucocorticoids were found to inhibit the clonal growth of K562 cells in vitro, suggesting a potential role for glucocorticoid binding sites in the modulation of K562 cell proliferation.
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PMID:Identification of a glucocorticoid receptor in the human leukemia cell line K562. 720 98

In the presence of Mn2+, reverse transcriptase of both human immunodeficiency virus and murine leukemia virus hydrolyzes duplex RNA. However, designating this novel activity RNase D conflicts with Escherichia coli RNase D, which participates in tRNA processing. On the basis of its location in the RNase H domain, we propose that this novel retroviral activity be redesignated RNase H*.
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PMID:Redesignation of the RNase D activity associated with retroviral reverse transcriptase as RNase H. 750 4

Replication complexes that contained either murine leukemia virus reverse transcriptase (MLV RT) or a variant reverse transcriptase without a ribonuclease (RNase) H domain (delta RH MLV RT) were visualized by enzymatic footprinting. Wild-type MLV RT protected template nucleotides +6 to -27, and primer nucleotides -1 to -26 of primers that had first been extended by one or four nucleotides. Although it catalyzed DNA synthesis, delta RH MLV RT stably bound template-primer only under conditions of reduced ionic strength and protected the duplex portion only as far as position -15. Despite altered hydrolysis profiles, both enzymes covered primarily the template-primer duplex, contradicting recent predictions based on the structure of rat DNA polymerase beta.
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PMID:Footprint analysis of replicating murine leukemia virus reverse transcriptase. 752 42

The reverse transcriptase of retroviruses contains an RNase H activity essential for the proper synthesis of the viral DNA copy of the RNA genome. We have previously characterized a number of point mutations altering the RNase domain of the Moloney murine leukemia virus reverse transcriptase (S. W. Blain and S. P. Goff, J. Biol. Chem. 268:23585-23592, 1993). One such mutation, Y586F (a Y-to-F change at position 586), reduced RNase H activity, as assayed by in situ gel analysis, to about 5% of the wild-type level and prevented viral replication. We have now recovered a revertant virus with near-normal infectivity and in vitro enzymatic activity. The revertant contains a single substitution, N613H, distant in the primary sequence of the protein, but modeling with the Escherichia coli RNase H structure suggests that the reverted residue is close in space to the original substituted residue. Examination of the structure permits some suggestions as to how this second-site revertant restores enzyme activity.
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PMID:Reversion of a Moloney murine leukemia virus RNase H mutant at a second site restores enzyme function and infectivity. 754 47

Previously, we showed that in vitro resistance to daunorubicin (DNR) at initial diagnosis was related to a poor long-term clinical outcome in childhood acute lymphoblastic leukemia (ALL), and that cells of relapsed ALL were in vitro more resistant to DNR than cells of untreated ALL. Topoisomerase II (Topo II) is an intracellular target for anthracyclines and epipodophyllotoxins. Decreased levels and/or activity of Topo II have been associated with multidrug resistance in cell lines. We investigated Topo II alpha gene expression in fresh leukemic samples from 19 children with untreated and 14 children with relapsed ALL using a sensitive RNase protection assay. The in vitro cytotoxicity of the Topo II inhibitors DNA and teniposide (VM26) was measured using the MTT assay, and the cell cycle distribution of leukemic samples was analyzed by DNA flow cytometry. Results showed that (1) relapsed ALL samples were more resistant to DNR, but not to VM26 compared to untreated samples; (2) large interpatient variations existed in both Topo II alpha gene expression and in vitro cytotoxicity results; (3) Topo II alpha gene expression was detectable in 29/33 childhood ALL samples with a median expression of 5% the level of a relatively chemosensitive human small cell lung cancer cell line; (4) Topo II alpha gene expression did not differ between untreated and relapsed ALL; (5) Topo II alpha gene expression was positively correlated with the percentage of ALL cells in S- and G2M-phase, but not with the in vitro cytotoxicity of the drugs tested. In conclusion, resistance to DNR in childhood ALL can not be explained by decreased levels of Topo II alpha gene expression, but additional Topo II activity studies in fresh leukemia samples may need further exploration.
Leukemia 1995 Oct
PMID:Topoisomerase II alpha gene expression in childhood acute lymphoblastic leukemia. 756 5

In this paper we report the presence and function of the 5' untranslated region (5'UTR) from the mRNA encoding human gamma-glutamyltransferase (GGT) in three different hematopoietic cell lines (HL-60, U-937 and K-562) as well as in the RNA of the leukocyte fraction from six acute lymphoblastic leukemias (ALL). Results obtained by RNase protection analysis demonstrate the presence of a unique form of 5'UTR expressed in most human tissues. In order to investigate the possible role of this type of sequence on regulation of GGT in hematopoietic cells, plasmid constructs carrying human hepatoma GGT 5'UTR and a luciferase reporter gene were transfected into the three blood cell lines. Compared to control untransfected cells, transfected HL-60 and K-562 showed a decrease in reporter gene activity of 51 and 73%, respectively. In contrast, transfected U-937 showed a 139% increase of reporter gene activity. Results were compared to GGT activity in the relevant cells and we concluded that the 5'UTR appears to have a regulatory role in GGT expression as a tissue-specific modulator of translation.
Leukemia 1995 Aug
PMID:Characterization and regulatory effect of gamma-glutamyltransferase messenger RNA untranslated regions in human leukemia. 764 21

HERV-K is a 50-copy, human endogenous, class 1 retroviral element that contains some polycistrons with gag, pol, and env open reading frames. Although expression of HERV-K proviruses has been shown in cultured human cell lines, expression of these elements has not been shown in human blood leukocytes. Using both reverse transcriptase-polymerase chain reaction and ribonuclease protection techniques, we show HERV-K pol gene expression in human blood leukocytes. Expression in blood leukocytes from 7 normal individuals was from a variety of different HERV-K proviruses, while restricted expression was observed in blood cells of 5 leukemia patients and 3 polycythemia vera patients. Evidence is presented suggesting that the restricted expression in leukemia blood cells is a result of gene regulation, not gene amplification.
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PMID:Expression of HERV-K proviruses in human leukocytes. 768 17

Nuclear RNA synthesis can be analysed by flow cytometry of cells labelled with 5-bromouridine (BrUrd) and stained with anti-bromodeoxyuridine (BrdUrd) antibody and FITC-conjugated secondary antibody. A panel of 5 different commercially available anti-BrdUrd antibodies was tested on cells of a HL-60 human leukemia cell line, stained as a methanol-fixed nuclear suspension. The BrUrd-induced fluorescence signals were highest with the antibody ABDM (Partec), moderate but reproducible with B-44 (Becton Dickinson), variable or low with BR-3 and IU-4 (Caltag), and not detectable with Bu20a (DAKO). Treatment of BrUrd-labelled nuclei with ribonuclease before staining with antibodies indicated that ABDM and B-44 antibodies specifically recognized BrUrd-substituted RNA, whereas BR-3 and IU-4 antibodies also bound to BrUrd-unlabelled RNA. Combined analysis of BrUrd and DNA contents demonstrated the variation of RNA synthesis during the cell cycle. The BrUrd incorporation was high in the S and G2 phase, variable in G1, and negligible in mitosis. Similar results were obtained using other cell types.
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PMID:Flow cytometric measurement of RNA synthesis using bromouridine labelling and bromodeoxyuridine antibodies. 768 81

Because mutations in receptor tyrosine kinases may contribute to cellular transformation, studies were undertaken to examine c-kit in human leukemia. Isoforms of c-kit have been characterized in the human megakaryoblastic leukemia cell line M-07. Deletion of the four amino acids Gly-Asn-Asn-Lys in the extracellular domain represents an alternatively spliced isoform that has been shown by others, in mice, to be associated with constitutive receptor autophosphorylation (Reith et al, EMBO J 10:2451, 1991). Additional isoforms differ in the inclusion or exclusion of a serine residue in the interkinase domain, a region that contains the binding site for phosphatidylinositol 3-kinase. By RNase protection analysis, we have shown coexpression of the Gly-Asn-Asn-Lys+ and Gly-Asn-Asn-Lys- isoforms, with dominance of the Gly-Asn-Asn-Lys- transcript, in normal human bone marrow, normal melanocytes, a range of tumor cell lines, and the blasts of 23 patients with acute myeloid leukemia. Analysis of transcripts for the Ser+ and Ser- isoforms also showed coexpression in all normal and leukemic cells examined. The ratios of isoform expression for both the Gly-Asn-Asn-Lys and Ser variants were relatively constant, providing no evidence in the tumors examined that upregulation of one isoform contributes to the neoplastic process.
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PMID:Expression of isoforms of the human receptor tyrosine kinase c-kit in leukemic cell lines and acute myeloid leukemia. 768 88

Differentiation inhibiting activity/leukaemia inhibitory factor (DIA/LIF) is a pleiotropic cytokine which has been implicated in a variety of developmental and physiological processes in mammals due to its broad range of biological activities in vitro. A role in very early development is suggested by the requirement for DIA/LIF to support the self-renewal of cultured embryonic stem (ES) cells. Other data point to potential roles in the establishment and maintenance of primordial germ cells, in osteogenesis and in haematopoiesis, and possibly in neuronal specification. DIA/LIF may also act as a mediator of the hepatic acute phase response. In the present study the expression of DIA/LIF transcripts during murine development and in adult mice has been determined using a highly sensitive ribonuclease protection analysis. In contrast to previous reports, it is apparent that DIA/LIF transcripts are present at low levels in many adult mouse tissues. Higher levels of expression are observed in skin, lung, intestine, and uterus. Elevated amounts of mRNA are also found in certain foetal tissue during late gestation and neonatally. In earlier embryogenesis, however, DIA/LIF mRNA is produced primarily in extraembryonic tissues. The alternative transcripts which produce either soluble or matrix-associated DIA/LIF exhibit overlapping but non-identical patterns of expression, consistent with the proposition that the two isoforms may have distinct biological functions. These findings are suggestive of widespread roles for DIA/LIF in vivo and are discussed in the light of available data on the phenotype of homozygous DIA/LIF-deficient mice.
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PMID:Expression of alternative forms of differentiation inhibiting activity (DIA/LIF) during murine embryogenesis and in neonatal and adult tissues. 768 30


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