Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.4.2.8 (hypoxanthine-guanine phosphoribosyltransferase)
 2,527 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Polyethylene glycol-1000 (PEG-1000) induced fusion of HPRT (E.C. 2.4.2.8) deficient Chinese hamster cells with alpha-galactosidase A (E.C. 2.3.1.22) deficient cells from a patient with Fabry's disease yielded hybrids which contained both human and hamster HPRT, G6PD (E.C. 1.1.1.49), and APRT (E.C. 2.4.2.7) and Chinese hamster alpha-galactosidase B. Thus PEG-1000 mediated somatic cell fusion led to reexpression of Chinese hamster HPRT. It did not restore the expression of human alpha-galactosidase. Since PEG-1000 treatment of HPRT- Chinese hamster cells in the absence of human cells yielded no HPRT+ cells, it is concluded that the element responsible for the restoration of rodent HPRT was contributed by the human cells and not by the agent employed to promote fusion.
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PMID:Reexpression of HPRT activity following cell fusion with polyethylene glycol. 20 82

In the present study we tested the feasibility of utilizing a structurally modified polyene antibiotic, amphotericin B methyl ester (AME), as a half-selection agent for isolating somatic cell hybrids. By using HAT medium supplemented with AME we have isolated interspecific mouse-hamster hybrids from mixed cultures of mouse (TK-C1 ID or HPRT-A9) and hamster (BHK/C 13) cells fused with Sendai virus, lysolecithin or polyethylene glycol. Hybrid cells proliferated and clones were isolated after 2 to 3 weeks growth in three changes of HAT-AME medium and subsequent growth in HAT medium alone. In contrast, genetically deficient parental C1 1D or A9 cells and AME-sensitive BHK/C 13 cells were killed using a similar growth protocol. The described technique is simple, efficient and permits one to use a cell line without a genetic defect in combination with a genetically deficient cell type in hybrid formation.
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PMID:Selection of mouse X hamster hybrids using HAT medium and a polyene antibiotic. 35 13

Male New Zealand White rabbits were immunized with human adenine phosphoribosyltransferase (APRT) and hypoxanthine-guanine phosphoribosyltransferase (HGPRT), which were purified about 2000-fold and 800-fold, respectively, from erythrocytes by DEAE-cellulose chromatography, ammonium sulfate precipitation and preparative polyacrylamide gel electrophoresis. Specific immunoprecipitations of APRT and HGPRT were achieved with the antisera that were obtained and by using polyethylene glycol as a substitute for goat anti-(rabbit) gamma globulin. The activities of the human forms of these enzymes, whether from red blood cells or from cultured cells, were almost completely eliminated under the conditions of immunoprecipitation used. Little or no reduction of APRT and HGPRT activities from mouse and Chinese hamster cells was observed. This discriminatory capacity of the antisera was successfully used for the identification of human APRT and HGPRT in human-mouse and human-hamster cell hybrids using the immunoprecipitation reaction.
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PMID:Adenine phosphoribosyltransferase and hypoxanthine-guanine phosphoribosyltransferase immunoprecipitation reactions in human-mouse and human-hamster cell hybrids. 118 4

The clonal rat pituitary tumor cell line GH4C1 secretes PRL but does not respond to dopamine, a physiological inhibitor of PRL. In an attempt to generate a dopamine-responsive cell line, GH4C1 cells, which lack the enzyme hypoxanthine-guanine phosphoribosyltransferase, were fused with cells from the pituitary glands of lactating rats to generate cell hybrids. The GH4C1 cells were fused with dispersed normal pituitary cells by either chemical fusion in 40% polyethylene glycol or electrofusion. The fused cells were grown in medium with hypoxanthine, aminopterin, and thymidine (HAT) for 4 weeks to select for hybrid cells. Control fusions between GH4C1 cells only or normal cells only did not produce viable colonies. Of 36 HAT-selected colonies, 3 responded to 10 nM bromocryptine (BCR) with inhibition of TRH-stimulated PRL release. These hybrid colonies had an inhibitory response similar to that of normal pituitary cells in culture. Both TRH- and vasoactive intestinal peptide-stimulated PRL release were inhibited to basal levels by 10 nM BCR, with an IC50 for BCR of approximately 0.25 nM. Basal hormone release was not inhibited by BCR. The BCR-sensitive hybrid cells grew more slowly than the parental GH4C1 line both in culture and when passaged in female Wistar-Furth rats. The response of the hybrid cells to the dopamine agonist and the characteristic of slow growth were lost after 9 months of continuous culture and after freezing cells. The parental GH4C1 cells were grown in female Wistar-Furth rats, the resulting tumors were dissociated, and the cells were grown in culture. This resulted in a brief establishment of the dopamine response. Stimulated PRL and GH release from freshly dispersed GH4C1 tumor cells was inhibited by BCR at concentrations from 0.1-10 nM, and spiroperidol reversed the inhibition. The inhibitory response to the dopaminergic agonist was lost quickly as the cells were carried in culture. These results demonstrate that GH4C1 cells may have the genetic information necessary for dopaminergic inhibition of PRL synthesis, but that the dopamine response is not observed under standard tissue culture conditions.
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PMID:Transient dopaminergic inhibition of prolactin release from hybrid cells derived by fusion of normal rat pituitary and GH4C1 tumor cells. 312 81

In studies of gene regulation using somatic cell fusion techniques, the analysis of heterokaryons circumvents several problematic aspects of the more traditional approach utilizing proliferating hybrid cells. We have analyzed the expression of muscle specific properties in heterokaryons between muscle and nonmuscle cells in order to investigate whether differentiating cells contain regulatory factors that repress the expression of alternative developmental pathways. Heterokaryons and cybrids were derived from polyethylene glycol-mediated fusion of differentiated mononucleate chicken myocytes with mouse melanoma cells, mouse melanoma cytoplasts, chicken fibroblasts, or other chicken myocytes. Our results demonstrate that fusion of a myocyte with a nonmyogenic cell generally results in extinction of muscle-specific properties in the immediate fusion product. Myocyte X melanoma heterokaryons ceased to express the skeletal muscle forms of myosin, desmin and creatine kinase, reinitiated DNA synthesis, and showed a loss of spontaneous fusion competence within 96 hr after their formation. Although chicken myocyte X mouse melanoma heterokaryons showed extinction of muscle specific properties, they continued to synthesize protein and to incorporate [3H]hypoxanthine, presumably due to the continued production of constitutive chicken HPRT. That presence of the melanoma nucleus was required for extinction to be observed was demonstrated by the continued expression of muscle proteins in cybrids between chicken myocytes and melanoma cytoplasts. Significantly, heterokaryons between chicken myocytes and chicken fibroblasts also exhibited extinction of muscle proteins, demonstrating for the first time that extinction is not restricted to fusions in which at least one parental cell type was derived from an established cell line. Our results strongly support the notion that extinction reflects cell-type specific gene regulatory mechanisms operative during development.
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PMID:Extinction of muscle-specific properties in somatic cell heterokaryons. 669 89

After polyethylene glycol treatment of GPRT- mutant cells, 12 clones were isolated on the ATG medium showing intragenic complementation. Karyological analysis confirmed the hybrid nature of the clones isolated. The GPRT activity in the hybrid clones, as assessed in vitro, exceeded the sum of parental activities. In vivo incorporation of 14C-hypoxanthine showed the GPRT activity in the hybrids to be an order of magnitude higher than in the mutant parental cells. Moreover, the GPRT activity in the hybrid clones was found to increase considerably during cultivation on the ATG medium; hence, their ability to multiply on this selective medium. All hybrid cells surviving and multiplying on the ATG medium were shown to maintain a high enough resistance to 8-AG, 6-MP and, to somewhat less extent, to 6-TG. The frequency of complementation was determined for the cells of mutant clones selected on media with different purine base analogues. The complementation map for the GPRT locus was constructed and proved to be linear. Five groups of complementation were specified.
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PMID:[Chinese hamster cells mutant for the hypoxanthine-guanine phosphoribosyltransferase locus. II. Characteristics of the hybrids revealing intragenic complementation]. 719 1

P388 mouse leukemia lines, one sensitive (P388/S) and the other resistance (P388/R) to vincristine (VCR), cultured in vitro, were hybridized with polyethylene glycol (PEG). A thymidine kinase-deficient mutant (TK-) was isolated from the sensitive line, and a hypoxanthine-guanine phosphoribosyltransferase-deficient mutant (HPRT-) from the resistant line. The hybrid line grows slower than the mutants. The modal chromosome numbers are: TK- = 38, HPRT- = 40, hybrid = 69 (72). The TK- cells contain a large metacentric marker which is missing from the HPRT- cells. Hybrid cells are as resistant to VCR as the P388/R and HPRT- cells.
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PMID:Drug resistance studies on intraspecific hybridomas. 725 34

2-Methoxyethanol (ethylene glycol monomethyl ether) (EGME), is one of the most commonly used solvents for industrial and consumer products. Although the solvent has been shown to be a reproductive toxin the genotoxic activities of EGME especially its metabolites, have not been adequately investigated. The mutagenicity and cytotoxicity of EGME and its major metabolites, methoxyacetaldehyde (MALD) and methoxyacetic acid (MAA) in Chinese hamster ovary (CHO) cells were therefore examined by us. We have determined the mutagenicity of these compounds at the hypoxanthine-guanine phosphoribosyltransferase (hprt) locus in CHO-K1-BH4 cells (CHO/HPRT assay) and the xanthine-guanine phosphoribosyl transferase (gpt) locus in CHO AS52 cells (AS52/GPT assay). The results show that these chemicals are not mutagenic to the hprt locus in CHO-K1-BH4 cells either with or without rat liver S9 mix as the metabolic activating system. With AS52 cells, only MALD is mutagenic in the absence of S9. It induced a dose-dependent mutagenic response. A dose-dependent cytotoxicity was induced by all compounds in both cell lines. MALD is the most and EGME is the least cytotoxic compounds. Our study shows that a metabolite of EGME, MALD, is highly cytotoxic and likely induces deletion-type mutations in AS52 cells. The genotoxic effect of EGME is, therefore, dependent upon its metabolism and its detection is dependent upon the assays used.
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PMID:Mutagenicity and cytotoxicity of 2-methoxyethanol and its metabolites in Chinese hamster cells (the CHO/HPRT and AS52/GPT assays). 767 57

Native and His-tagged mutant (L160I) hypoxanthine-guanine phosphoribosyltransferase (HGPRT) from Thermoanaerobacter tengcongensis were cloned, expressed in Escherichia coli and purified. Both proteins were crystallized with polyethylene glycol as the main precipitant at 293 K using the hanging-drop vapour-diffusion method. The crystal of native HGPRT belongs to space group C222(1), with unit-cell parameters a = 65.77, b = 137.73, c = 95.27 A, and diffracted to 2.2 A resolution on an in-house X-ray generator. The crystal of the His-tagged mutant (L160I) HGPRT belongs to the space group I222, with unit-cell parameters a = 52.21, b = 88.36, c = 93.03 A, and diffracted to 1.7 A resolution in-house.
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PMID:Protein preparation, crystallization and preliminary X-ray crystallographic studies of a thermostable hypoxanthine-guanine phosphoribosyltransferase from Thermoanaerobacter tengcongensis. 1450 Nov 39