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:2.7.1.21 (thymidine kinase)
7,561 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

From a population of wild type S49 cells, a clone, DTB6, was isolated in a single step from selective medium containing thymidine and dibutyryl cyclic AMP that exhibited a 60% deficiency in AMP deaminase (AMP-D) activity. The AMP-D deficiency conferred to the DTB6 cells a striking susceptibility to killing by low concentrations of either adenine or adenosine, the latter in the presence of an inhibitor of adenosine deaminase activity. This growth supersensitivity of DTB6 cells toward adenine could be ameliorated by the addition of hypoxanthine to the culture medium. Immunoprecipitation of AMP-D from wild type and mutant cells revealed that the DTB6 cell line contained markedly diminished amounts of the AMP-D isozyme which reacts with antisera to the predominant isoform expressed in adult kidney. The quantities of the AMP-D isozyme immunoprecipitated by antisera raised to the predominant isoform prepared from adult heart were equivalent in the two cell lines. Although Northern blot analyses revealed no alterations in mRNA sizes or levels encoded by either of the AMP-D genes, Southern blots of genomic DNA hybridized to a cDNA specific for the ampd2 gene revealed the presence of a new BamHI restriction fragment in the DNA of DTB6 cells. These data suggested that a point mutation has occurred in the ampd2 gene of DTB6 cells which encodes the AMP-D isozyme recognized by the kidney antisera. The DTB6 cells also possessed a virtual complete deficiency in thymidine kinase activity. The two enzyme deficiencies were distinguishable. The ability to isolate single step mutants with two seemingly independent biochemical abnormalities raises the speculation that there may be some link between cellular functions responsible for purine nucleotide and thymidine metabolism.
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PMID:Adenylate deaminase deficiency in a mutant murine T cell lymphoma cell line. 236 81

The mechanism of cyclic AMP (cAMP) induction of fibronectin (FN) in HT-1080 and JEG-3 cells differs (D. C. Dean, R. F. Newby, and S. Bourgeois, J. Cell Biol. 106:2159-2170, 1988). In the fibrosarcoma cell line HT-1080, induction requires both protein synthesis and a lag period of 12 to 24 h. In the choriocarcinoma cell line JEG-3, protein synthesis is not required and induction peaks before 24 h, declining thereafter. We show that the FN promoter is transcribed in vitro and that the transcripts initiate at the proper site. Based on transfection experiments with these cells and FN promoter constructions, a cAMP-responsive element (CRE) was identified between -157 and -188 base pairs upstream of the human FN gene. This sequence also conferred cAMP inducibility in both cell lines on the herpesvirus thymidine kinase promoter when it was placed upstream of a thymidine kinase-chloramphenicol acetyltransferase fusion gene. DNase I protection analysis and gel retardation experiments revealed that the CRE was bound by a protein(s) that was present in both HT-1080 and JEG-3 cells as well as in NIH 3T3 cells. Multiple protein-CRE complexes were resolved by gel retardation with extracts of both cell lines. Forskolin treatment of these cells did not alter qualitatively or quantitatively the pattern of CRE-binding proteins that was observed. The FN promoter was at least 10 times more active in HT-1080 than in JEG-3 cells, even though in JEG-3 cells both the rate of FN biosynthesis and the level of accumulated FN mRNA were greater than those in HT-1080 cells. The difference in promoter activity in HT-1080 and JEG-3 cell was mediated by sequences that were located between positions -510 and -56. Deletion of the FN promoter from positions -510 to -56 resulted in an ~30-fold decrease in promoter activity when this construction was transfected into HT-1080 cells, and similar results were observed in NIH 3T3 cells; however, less than a 2-fold effect was observed in JEG-3 cells. Results of these studies suggest that there is some degree of tissue specificity of FN gene expression and reveal that cAMP induction is mediated, in part, by the same element (CRE) in both HT-1080 and JEG-3 cells.
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PMID:Forskolin inducibility and tissue-specific expression of the fibronectin promoter. 254 72

The study was concerned with comparison of the activity of thymidine kinase, thymidine phosphorylase, adenosine desaminase and AMP 5'-nucleotidase in serum of cancer patients and healthy subjects. A significant change in 5'-nucleotidase level was registered in patients over 60 years of age and in those of adenosine desaminase and thymidine phosphorylase in patients older than 70 years. A comparative evaluation of the activity of those enzymes showed an increase in thymidine kinase activity matched by a sharp drop in that of thymidine phosphorylase in patients over 45 years. A sharp increase in the activity of adenosine metabolizing enzymes in cancer serum was accompanied by its decrease in lymphocytes. Among surgical patients, significant changes in the activity were observed in radically operated cases only.
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PMID:[The enzymatic activity of DNA metabolism of the blood in patients operated on in stomach cancer]. 254 45

The 5' flanking regions of the rat phosphoenolpyruvate carboxykinase gene were used to form chimeric gene constructs with the human growth hormone gene. These constructs were transfected into several renal and one liver cell line and the production of growth hormone (HGH) measured by immunoassay. Cyclic-AMP and glucocorticoid responsiveness of HGH production was observed in all cell lines. In two lines, the rat NRK52E renal epithelial line and the rat H4IIE hepatoma cell line, both capable of expressing PEPCK, lowering extracellular pH increased HGH production several fold. Comparison of hormone and pH effect on cells transfected with a thymidine kinase promoter-HGH chimera indicated that the PEPCK 5' flanking region effects were specific. Thus, part of the pH responsiveness of the PEPCK gene in vivo may be attributed to properties of the 5' flanking regions.
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PMID:The 5'region of the rat phosphoenolpyruvate carboxykinase gene confers pH sensitivity to chimeric genes expressed in renal and liver cell lines capable of expressing PEPCK. 255 24

From a mutagenized population of wild type S49 cells, a clone was isolated in a single step that possessed functional and biochemical deficiencies in both AMP deaminase and thymidine kinase activities. This mutant cell line, DTB6, was selected in semi-solid medium containing 1mM thymidine and 1mM dibutyryl cyclic AMP. In comparative growth rate experiments, DTB6 cells were considerably less sensitive than parental cells to the growth inhibitory effects of thymidine. In contrast, DTB6 cells were much more sensitive to the cytotoxic effects of adenine and adenosine. The supersensitivity of DTB6 cells toward adenine could be ameliorated by the addition of hypoxanthine to the culture medium. The growth phenotype of the mutant cells could be attributed to deficiencies in two enzyme activities. First, DTB6 cells possessed a 60-70% deficiency in AMP deaminase activity, although the residual activity appeared kinetically similar to the wild type enzyme. Second, DTB6 cells possessed a virtual complete deficiency in thymidine kinase activity. Both enzyme deficiencies behaved in a recessive fashion in intraspecies hybrids. Revertants of DTB6 cells possessed wild type levels of AMP deaminase activity but remained deficient in thymidine kinase activity, while another revertant of DTB6 cells expressed 11% of the wild type thymidine kinase level but did not perceptibly change its AMP deaminase activity. The ability to isolate single step mutants with two seemingly independent biochemical abnormalities raises the speculation that there may be some link between cellular functions responsible for purine nucleotide and thymidine metabolism.
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PMID:AMP deaminase and thymidine kinase deficiencies in a mutant mouse S49 cell clone. 261 Jan 48

Cleavage and polyadenylation of substrate RNAs containing the herpes simplex virus type 1 (HSV-1) thymidine kinase (tk) gene polyadenylation signal region were examined in HeLa cell nuclear extract. 3'-End RNA processing was accurate and efficient and required ATP and Mg2+. Cleavage, but not polyadenylation, occurred in the presence of EDTA or when ATP was replaced with 3' dATP (cordycepin) or AMP(CH2)PP, a nonhydrolyzable analog of ATP. Processing in vitro and in vivo showed the same signal element requirements: a series of substrates containing linker scanning, internal deletion, and small insertion mutations was processed with the same relative efficiencies and at the same sites in vitro and in vivo. A complex involved in 3'-end RNA processing was identified by gel mobility shift analysis. This complex formed rapidly, reached a maximum level after 20 to 30 min, and was much reduced after 2 h. Very little complex was formed at 0 degree C or with substrates lacking a polyadenylation signal. Entry of 32P-labeled tk substrate into the complex could be prevented by addition of excess 35S-labeled tk or adenovirus L3 precursor RNAs. Competition was not observed with tk RNAs lacking a complete polyadenylation signal.
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PMID:Identification of a complex associated with processing and polyadenylation in vitro of herpes simplex virus type 1 thymidine kinase precursor RNA. 282 24

DNA-mediated gene transformation of mouse Ltk-aprt-hprt-cells was used to obtain stable, doubly selected transformants simultaneously expressing herpes virus thymidine kinase (TK) and mammalian adenine phosphoribosyltransferase (APRT). Cotransformants occurred at a frequency of 5 X 10(-6), a similar frequency for the transfer of the aprt marker has been previously observed. Isozyme and Southern blot analysis show that the TK and APRT expressed in these transformants resulted from gene transfer. For one stable cotransformant, [3H]thymidine [( 3H]TdR) selection against TK activity resulted in the loss of APRT activity as well, suggesting that these genes had become genetically linked together. Similarly selection against APRT expression resulted in the loss of a subset of the transferred herpes simplex virus tk genes. 5-Bromodeoxyuridine (BUdR) selected TK- variants differed from [3H]TdR selected TK- variants, in that they retained tk genes. However, BUdR-selected variants expressed full levels of APRT. Therefore, even though the transferred tk and aprt genes had become genetically linked together, they were, in this case, independently expressed since these cells were phenotypically TK- and APRT+.
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PMID:Genetic linkage but independent expression of functional HSV-1 tk and mammalian aprt genes after cotransfer to L cells. 298 26

The mutagenic effect of cadmium chloride on somatic cells of F1 hybrid mice CBA X C57B1/6J in vivo and on an established line of CHO-ATZ-2 Chinese hamster cells in vitro has been studied. The induction of micronuclei has been demonstrated in mouse marrow cells as well as induction of point mutations at loci controlling the synthesis of hypoxanthine-phosphoribosyltransferase, thymidine kinase, adenine phosphoribosyltransferase and the resistance of Na+/K+ ATPase to ouabain in the cell line CHO-AT-2. A peak of mutagenic activity under the action of subtoxic doses of cadmium chloride has been revealed.
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PMID:[Mutagenic action of cadmium chloride in mammals]. 343 31

Mouse fibroblast cell lines which secrete bovine (b) GH have been generated. This was accomplished by cotransforming mouse L cells (thymidine kinase-negative [TK-] and adenine phosphoribosyl transferase-negative [APRT-]) with DNA molecules encoding the Rous sarcoma virus-long-terminal repeat and bGH genes along with the herpes viral TK gene and the hamster APRT gene. One stable cell line, Pd lambda-pbGH 4-13, was found to secrete approximately 75 micrograms bGH per 24 h/5.0 X 10(6) cells. Media from this cell line were collected for purification of recombinant bGH (rbGH). Purification involved (NH4)2SO4 fractionation, ion-exchange chromatography, and gel filtration on Sephacryl S-200. The rbGH was characterized by bioassay, RIA, radioreceptor assay, and sodium dodecyl sulfate gel electrophoresis. Results of these analyses were compared with those obtained with a highly purified pituitary bGH. In the rat tibia bioassay, rbGH was found to be as potent as pituitary bGH. Results from the RIA, radioreceptor assay, and sodium dodecyl sulfate gel electrophoresis and Western blot analysis also suggested that the rbGH was similar to that of pituitary origin. Amino acid composition, partial (amino-terminal) sequence, and tryptic peptide maps were also found to be similar between the rbGH and pituitary bGH preparations. The amino terminus of the rbGH showed similar heterogeneity to that of the bGH of pituitary origin. We conclude that rbGH which was synthesized, processed, and secreted from transformed mouse fibroblasts possessed almost exactly the same physiochemical properties as pituitary bGH.
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PMID:Purification and physiochemical properties of a recombinant bovine growth hormone produced by cultured murine fibroblasts. 375

The effect of DNA methylation on the transcriptional activity of the hamster adenine phosphoribosyltransferase (aprt) and the herpes thymidine kinase (tk) genes has been investigated. By using M13 constructs containing these gene sequences, specific segments of each gene were methylated in vitro by restriction fragment primer-directed second-strand synthesis using the substrate 2'-deoxy-5-methyl-cytidine triphosphate (dmCTP). These hybrid-methylated molecules were inserted into mouse Ltk- cells by DNA-mediated cotransfer. In all cases, the integrated sequences retained the in vitro-directed methylation pattern. The aprt gene was inhibited by CpG methylation in the 5' region but was unaffected by methylation at the 3' end or in adjacent M13 sequences. In contrast to this, DNA methylation in both the 5' promoter region and the 3' structural region of the tk gene had a strong inhibitory effect. This suggests that this modification may affect transcription by mechanisms that do not involve the direct alteration of recognition sequences for RNA polymerase.
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PMID:Effect of regional DNA methylation on gene expression. 385 99


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