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:5.99.1.2 (topoisomerase)
9,166 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In order to characterize more fully the mechanism by which casein kinase II is regulated in mammalian cells, the effect of epidermal growth factor (EGF) on the activity of the kinase in human A-431 carcinoma cells was examined. Treatment of cells with EGF prior to lysis consistently resulted in a transient 4-fold increase in the activity of cytosolic casein kinase II. Activity rose sharply between 20 and 30 min, peaked at approximately 50 min, and returned to basal levels by approximately 120 min. Similar results were obtained using the casein kinase II specific peptide substrate, Arg-Arg-Arg-Glu-Glu-Glu-Thr-Glu-Glu-Glu, or DNA topoisomerase II (which is specifically modified by the kinase in vivo and serves as a high affinity substrate in vitro) as the phosphate acceptor in assays. Identification of casein kinase II as the stimulated activity was confirmed by partial proteolytic mapping and phosphoamino acid analysis of modified topoisomerase II, by inhibition at nanomolar levels of heparin or micromolar levels of nonradioactive GTP, and by the ability to employ radioactive GTP as a direct phosphate donor. The EGF stimulation of casein kinase II was dependent on the availability of intracellular (but not extracellular) calcium. In addition, hormonal action was modulated by calcium/phospholipid-dependent protein kinase (protein kinase C). Casein kinase II stimulation did not require an increase in the concentration of the kinase, protein synthesis, the continual presence of a small effector molecule, or a direct interaction with the EGF receptor/tyrosine kinase. In contrast, hormonal activation of the kinase was dependent on the phosphorylation of casein kinase II or a terminal stimulatory factor.
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PMID:Regulation of casein kinase II activity by epidermal growth factor in human A-431 carcinoma cells. 247 67

The receptor for epidermal growth factor (EGF) is a single-chain transmembrane polypeptide of relative molecular mass (Mr) 170,000 (170K) which has been implicated in the regulation of both normal and abnormal cell proliferation. It has an externally facing EGF-binding domain and a cytoplasmically facing tyrosine-specific protein kinase site. Although the receptor has been well characterized, the mechanism by which it transmits the growth stimulatory signal from the plasma membrane to the nucleus is unclear. EGF binding to cells has been shown to enhance topoisomerase activity within the cells. Topoisomerases catalyse the interconversion of topological isomers of DNA and thus may influence replication and transcription. Mroczkowski et al. reported that purified EGF receptors of both human and murine origin can nick supercoiled double-stranded (ds) DNA in an ATP-dependent fashion, an activity related to those of topoisomerases. Another related tyrosine kinase, pp60src, has also been reported to have a similar DNA-nicking activity. We have now characterized the EGF receptor-associated DNA-nicking activity by sucrose gradient centrifugation. Our results, presented here, indicate that the DNA-nicking activity is not intrinsic to the EGF receptor, but is found in a distinct molecular species.
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PMID:EGF receptor-associated DNA-nicking activity is due to a Mr-100,000 dissociable protein. 299 1

Tyrosine protein kinase activity is associated with at least eight different retrovirus-encoded onc gene products and with cell receptors for epidermal growth factor, platelet-derived growth factor, tumour growth factor and insulin. Both the onc kinases and the growth factor receptors are membrane proteins whose enzymatic activity has been implicated in stimulation of growth. However, the mechanism by which a signal passes from the plasma membrane to the nucleus to initiate growth remains unknown. As DNA topoisomerases catalyse the interconversion of topological isomers of DNA and hence affect DNA replication, transcription and recombination, they may be involved also in stimulation of growth. Several DNA topoisomerases have been shown to form a covalent complex with DNA via a phosphotyrosine linkage. The DNA-protein complex is postulated to be an intermediate in breaking and rejoining of DNA. The aim of the present study was to determine whether tyrosine protein kinases modulate the activity of topoisomerases by phosphorylating the tyrosine residue involved in DNA binding. We report that incubation of Escherichia coli and calf thymus type I DNA topoisomerases with the Rous sarcoma virus transforming gene product, pp60src, and TPK75, a tyrosine protein kinase purified from normal rat liver, results in a 10-fold loss of topoisomerase activity.
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PMID:Virus- and cell-encoded tyrosine protein kinases inactivate DNA topoisomerases in vitro. 609 21

We have used epidermal growth factor (EGF) to investigate the relationship between eukaryotic topoisomerases and DNA synthesis. We found that EGF stimulates topoisomerase activity in human fibroblasts and Swiss/3T3 mouse fibroblasts. The first increase is seen in the cytoplasm, followed by increased activity in the nucleus. The nuclear increases correspond to increases in DNA synthesis. A type II topoisomerase is stimulated as indicated by the ATP dependence of the relaxing reaction and by the formation of catenanes. We have also found that the topoisomerase activity in the cytoplasm is sedimentable indicating that it is either membrane-associated or in a supramolecular complex. The stimulation of topoisomerase activity by EGF may represent a key step in the process by which EGF induces DNA synthesis and cell division.
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PMID:Epidermal growth factor-induced topoisomerase(s). Intracellular translocation and relation to DNA synthesis. 630 94

A rat pheochromocytoma cell line (PC12 cells) died within 24 h in the presence of etoposide (1-40 micrograms/ml), an inhibitor of topoisomerase II. This cytotoxic effect was prevented by either nerve growth (NGF) or epidermal growth factor (EGF). Cycloheximide and actinomycin D also suppressed the cell death as well. Furthermore, a difference among protective modes against etoposide-induced death by growth factors and a protein-synthesis inhibitor was observed: the protective effect of either NGF or EGF remained rather constant as a function of incubation time with etoposide whereas that of cycloheximide declined. These results indicate that etoposide induces programmed death in PC12 cells and that prevention of the programmed cell death by both NGF and EGF is mainly due to inactivation of molecules involved in the death processes rather than suppression of specific protein and/or mRNA synthesis.
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PMID:Nerve growth factor and epidermal growth factor rescue PC12 cells from programmed cell death induced by etoposide: distinct modes of protection against cell death by growth factors and a protein-synthesis inhibitor. 783 Sep 38

Mucosal healing requires enterocyte migration (restitution) supplemented by proliferation. Proliferation and migration may be studied independently by thymidine uptake and proliferation-blocked cell migration using human Caco-2 enterocyte monolayers in culture. Since epidermal growth factor (EGF) promotes mucosal healing and the EGF receptor is a tyrosine kinase, we hypothesized that tyrosine kinases might therefore modulate enterocyte migration and proliferation. The tyrosine kinase inhibitors genistein and 2,5-dihydroxymethylcinnamate, which block kinase ATP-binding and substrate-binding sites, respectively, were studied alone and with EGF. Proliferation was blocked with mitomycin. Although each inhibitor decreased basal and EGF-stimulated monolayer expansion when cell proliferation occurred, neither genistein nor 2,5-dihydroxymethylcinnamate decreased migration when proliferation was blocked. However, each inhibitor prevented EGF stimulation of proliferation-blocked migration and thymidine uptake. More substantial inhibition of basal proliferation by genistein correlated with increased protein-linked DNA breaks, which may reflect nonspecific inhibition of DNA topoisomerase activity by genistein. The more specific 2,5-dihydroxymethylcinnamate blocked changes in the alpha 2 integrin subunit organization which may modulate EGF-stimulated migration. Antiproliferative effects of tyrosine kinase inhibitors decrease basal monolayer expansion but true basal enterocyte migration appears independent of tyrosine kinase regulation. However, a specific tyrosine kinase-dependent modulation of cell-matrix interaction inhibits EGF-stimulated migration.
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PMID:Effect of tyrosine kinase inhibition on basal and epidermal growth factor-stimulated human Caco-2 enterocyte sheet migration and proliferation. 807 87

Because of its unique DNA-cleaving and strand-passing activities, topoisomerase II is involved in many aspects of DNA metabolism, including replication, transcription, recombination, and repair. The cytotoxic potential of topoisomerase II-targeted drugs, such as etoposide, is related to their ability to stabilize covalently linked enzyme-DNA complexes, which are intermediates in the enzyme's catalytic cycle. Epidermal growth factor receptor is expressed on the cell surface of the majority of squamous cell carcinomas, and epidermal growth factor binding is known to stimulate a number of cellular transduction pathways, including tyrosine kinase, protein kinase C, and phospholipase C. Because topoisomerase II is a proliferation-dependent protein and has been shown to be a high-affinity substrate for many of these cellular transduction pathways, the effects of epidermal growth factor on cellular regulation and sensitivity to etoposide were studied with the human oral cavity squamous cell line, KB. Topoisomerase II catalytic activity was rapidly and transiently inhibited after the addition of epidermal growth factor to the cellular growth media. Western blot on nuclear extracts did not demonstrate alterations in topoisomerase II polypeptide levels to account for changes in catalytic activity. Epidermal growth factor treatment also led to the formation of stabilized, covalently linked enzyme-DNA complexes. Furthermore, epidermal growth factor-induced, topoisomerase II-mediated DNA strand breaks were additive to those induced by etoposide. This study indicates that epidermal growth factor specifically regulates the catalytic and DNA-cleaving activities of topoisomerase II in KB cells. This may direct clinical strategies for circumventing the intrinsic cellular resistance to chemotherapy commonly observed in squamous cell carcinomas of the head and neck.
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PMID:Epidermal growth factor regulates topoisomerase II activity and drug sensitivity in human KB cells. 864 3

Breast cancer is the most common cancer in women. Because genetics is believed to account for only 10-15% of breast cancer cases, the environment, including nutrition, is thought to play a significant role in predisposing women to this cancer. Studies of Asian women suggest that those who consume a traditional diet high in soy products have a low incidence of breast cancer, but that among emigrants to the United States, the second generation, but not the first, loses this protection. These findings suggest a possible common mechanism of action for breast cancer protection from early, specific nutritional exposure. Genistein, an isoflavone found in soy, has been reported to have weak estrogenic and antiestrogenic properties, to be an antioxidant, to inhibit topoisomerase II and angiogenesis, and to induce cell differentiation. In studies of the mammary glands of immature rats, we showed that genistein up-regulates the expression of the epidermal growth factor receptor shortly after treatment, which may be responsible for the increased cell proliferation seen at that age. We hypothesize that the early genistein action promotes cell differentiation that results in a less active epidermal growth factor signaling pathway in adulthood that, in turn, suppresses the development of mammary cancer. We speculate that breast cancer protection in Asian women consuming a traditional soy-containing diet is derived from early exposure to soybean products containing genistein. We believe that early events are essential for the benefits of cancer protection.
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PMID:Protection against breast cancer with genistein: a component of soy. 1083 23

ErbB2 (HER-2) gene amplification and overexpression have been shown to predict a better outcome with doxorubicin-based chemotherapy as opposed to alkylator-based chemotherapy in early stage breast cancer. To understand the mechanism of differential response to these two regimens, we have evaluated the effect of signaling through the ErbB2 receptor on downstream enzymes that may affect drug response, using two different models. The first system employs breast cancer cells that have high levels of endogenous ErbB2 by gene amplification (BT-474 and SKBR3 cells). The second system allows us to isolate the effect of ErbB2 receptor-mediated intracellular signaling using an epidermal growth factor receptor-ErbB2 chimeric receptor activated by epidermal growth factor. Our experiments show that the cytotoxicity of doxorubicin is inhibited in ErbB2+ breast cancer cells by the anti-ErbB2 antibody, Herceptin. This is accompanied by a decrease in topoisomerase (topo) IIalpha protein and activity, suggesting that this is the mechanism of change in doxorubicin response. In addition, a 10-100-fold (1-2 log) decrease in the LD(50) of doxorubicin is seen after ErbB2 activation using the chimeric receptor model. Furthermore, we see a 100-fold decrease in the LD(50) of etoposide, another topo II inhibitor. This increase in doxorubicin sensitivity is associated with a 4.5-fold increase in the amount of topo IIalpha protein and an increase in topo II activity as measured by DNA decatenating and unknotting activities, as well as cleavable complex formation. In contradistinction to doxorubicin, we have observed an increased resistance to cyclophosphamide chemotherapy after chimeric receptor activation. We propose that the differential benefit seen with doxorubicin- versus alkylator-based chemotherapy in ErbB2+ breast cancer is due, in some cases, to ErbB2-mediated topo IIalpha activation. These data also suggest hypotheses for the optimal sequencing of Herceptin and chemotherapy agents in ErbB2+ breast cancer.
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PMID:Induction of topoisomerase II activity after ErbB2 activation is associated with a differential response to breast cancer chemotherapy. 1141 Apr 82

The relationship between expression and function of the epidermal growth factor (EGF) family of receptors and chemosensitivity remains controversial. We studied the chemosensitivity to various anticancer agents of human cervical squamous carcinoma ME180 cells, and two resistant subclones, ME180/TNF and ME180/Pt, which also differ in their EGF receptor (EGFR) expression. Compared with ME180 cells, EGFR is overexpressed sixfold in ME180/TNF cells and is barely detectable in ME 180/Pt cells. Cell cycle analysis by flow cytometry and BrdU incorporation into DNA showed a correlation between EGFR expression and percentage of cells in S phase and active DNA replication (35% in high EGFR-expressing ME180/TNF cells, 19% in non-EGFR-expressing ME180/Pt cells and 23% in parental, intermediate-level EGFR-expressing ME 180 cells). By MTT assay and compared with parental, intermediate-level EGFR-expressing ME180 cells, high EGFR-expressing ME180/TNF cells had a three- to fourfold increased sensitivity to cisplatin, camptothecin (CPT), and topotecan, and low EGFR-expressing ME180/Pt cells had a five- to ninefold reduced sensitivity to the same agents. In contrast, the degree of cross-resistance with the topoisomerase II inhibitors doxorubicin and etoposide was minimal and the pattern of sensitivity to the anti-microtubulin agents vinblastine and paclitaxel was different, with a two- to fourfold decreased sensitivity in the high EGFR-expressing ME180/TNF cells and only a 1.5-fold decreased sensitivity in the low EGFR-expressing ME180/Pt cells. Neither alterations in intracellular CPT levels nor changes in topoisomerase I expression or activity, measured as ability to form DNA-protein complexes, were found to explain the differences in sensitivity to CPT among the three cell lines. Co-treatment with CP358774, a specific EGFR tyrosine kinase inhibitor, reduced the enhanced sensitivity of high EGFR-expressing ME180/TNF cells to the values observed in intermediate EGFR-expressing ME180 cells, but only reduced modestly the sensitivity of intermediate expressing ME180 cells. As a result, the resistance index of low EGFR-expressing ME180/Pt cells compared with intermediate EGFR-expressing ME180 cells was reduced only from five- to fourfold for cisplatin and from seven- to fourfold for CPT when ME180 cells were exposed to CP358774. CP358774 did not affect the sensitivity to either agent in low EGFR-expressing ME180/Pt cells. These results provide evidence that changes in EGFR expression or function may play a role in determining chemosensitivity to platinum and topoisomerase I poisons in some human tumor systems, and that the EGFR-related changes in chemosensitivity may vary depending on the level of EGFR expression and/or function.
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PMID:Sensitivity to topoisomerase I inhibitors and cisplatin is associated with epidermal growth factor receptor expression in human cervical squamous carcinoma ME180 sublines. 1145 99


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