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Query: UNIPROT:P00492 (hypoxanthine-guanine phosphoribosyltransferase)
 2,385 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Recent molecular analysis of in vivo-derived hprt mutant T-lymphocytes cloned from human blood show that mutants occurring at the normal frequency (approximately 5 X 10(-6) in healthy young individuals generally represent independent hprt mutations. Here we report that in an individual with a high mutant frequency (86-620 X 10(-6],92% (61/66) of the mutant clones are descendents of an original mature T-cell precursor that has undergone in vivo clonal expansion. Therefore, these mutants could represent as few as one original hprt mutation. If so, correcting for the clonal expansion yields a revised calculated mutant frequency (Mf) value for this individual that is near the normal range. These hprt mutant clones all showed identically rearranged T-cell receptor (TCR) beta and gamma gene patterns by Southern blot analysis. All the clones were surface marker CD4+, showed no obvious chromosomal aberration, and had no detectable hprt gene structural alteration. This TCR-defined T-cell clone appears to have expanded in the blood of the individual over a 6-month period and persists at high levels after nearly 4 years. This finding illustrates the need to analyze mutants from individuals with high mutant frequencies at the molecular level in order to estimate hprt mutation frequency from the calculated hprt mutant frequency. The possibility that spontaneous hprt mutants might arise in vivo preferentially in dividing cells, and implications of this, are discussed.
Environ Mol Mutagen 1988
PMID:Molecular analyses of in vivo hypoxanthine-guanine phosphoribosyltransferase mutations in human T-lymphocytes: II. Demonstration of a clonal amplification of hprt mutant T-lymphocytes in vivo. 326 8

The localization of the A-raf cellular oncogene on the mouse X chromosome has been determined using Xbal-restricted DNAs prepared from progeny of an interspecies backcross between the B6.CBA.R1 and the Spe/Pas mouse strains. This localization to the proximal part of the mouse X chromosome has been confirmed by the use of somatic cell hybrids, carrying partially deleted X chromosomes and suggests that the A-raf oncogene localizes to a region lying some 10-17 centimorgans proximal to the hypoxanthine phosphoribosyltransferase (Hprt) gene between the locus DXPas4 and the locus DXPas7 defined by the cross-reacting human X chromosome-specific probe DXS32 (M2C). This localization on the mouse X chromosome is compatible with the presence of the A-raf oncogene on the short arm of the human X chromosome between the centromere and Xp21.
Somat Cell Mol Genet 1987 May
PMID:A-raf oncogene localizes on mouse X chromosome to region some 10-17 centimorgans proximal to hypoxanthine phosphoribosyltransferase gene. 329 48

Thioxanthine is toxic for mammalian cells transformed by the dominant selectable marker gpt. It allowed us to select, in the presence of the endogenous hypoxanthine-guanine phosphoribosyltransferase gene, mutants that did not express gpt any more and also hybrid cells that had lost the chromosome carrying it. The gpt marker is thus dominant in negative as well as in positive selection, which makes it potentially very useful for genetic studies of mammalian cells.
Mol Cell Biol 1987 Nov
PMID:Selection against expression of the Escherichia coli gene gpt in hprt+ mouse teratocarcinoma and hybrid cells. 332 88

Defective ecotropic and amphotropic retroviral vectors containing the cDNA for human hypoxanthine phosphoribosyltransferase (HPRT) were developed for efficient gene transfer and high-level cellular expression of HPRT. Helper cell clones which produced a high viral titer were generated by a simplified method which minimizes cell culture. We used the pZIP-NeoSV(X) vector containing a human hprt cDNA. Viral titers (1 X 10(3) to 5 X 10(4)/ml) of defective SVX HPRT B, a vector containing both the hprt and neo genes, were increased 3- to 10-fold by cocultivation of the ecotropic psi 2 and amphotropic PA-12 helper cells. Higher viral titers (8 X 10(5) to 7.5 X 10(6] were obtained when nonproducer NIH 3T3 cells or psi 2 cells carrying a single copy of SVX HPRT B were either transfected or infected by Moloney leukemia virus. The SVX HPRT B defective virus partially corrected the HPRT deficiency (4 to 56% of normal) of cultured rodent and human Lesch-Nyhan cells. However, instability of HPRT expression was detected in several infected clones. In these unstable variants, both retention and loss of the SVX HPRT B sequences were observed. In the former category, cells which became HPRT- (6-thioguanine resistant [6TGr]) also became G418s, indicative of a cis-acting down regulation of expression. Both hypoxanthine-aminopterin-thymidine resistance (HATr) and G418r could be regained by counterselection in hypoxanthine-aminopterin-thymidine. In vitro mouse bone marrow experiments indicated low-level expression of the neo gene in in vitro CFU assays. Individual CFU were isolated and pooled, and the human hprt gene was shown to be expressed. These studies demonstrated the applicability of vectors like SVX HPRT B for high-titer production of defective retroviruses required for hematopoietic gene transfer and expression.
Mol Cell Biol 1987 Feb
PMID:Construction of a defective retrovirus containing the human hypoxanthine phosphoribosyltransferase cDNA and its expression in cultured cells and mouse bone marrow. 346 9

Roberts syndrome (RS) is a rare human recessive disorder involving, in the chromosomes of some patients, a characteristic puffing or splitting apart of the constitutive heterochromatin (the RS effect). We carried out somatic cell hybridizations between an RS cell strain (R22) with the heterochromatin abnormality and a hypoxanthine phosphoribosyltransferase-deficient cell strain (GM1662) with normal chromosome structure to determine if the presence of the normal genome would correct the RS effect in the hybrid cells. In order to provide the fibroblast strains with dominant selection markers for the hybridizations, GM1662 was transfected with the plasmid pSV3neo which conferred resistance to the antibiotic G418, and R22 was transfected with the plasmid pSV3gpt which provided resistance to mycophenolic acid. Two somatic cell hybridizations were carried out: (1) R22 X GM1662 pSV3neo and (2) R22 pSV3gpt X GM1662 pSV3neo. The RS effect was found to be absent in 95% and 92%, respectively, of the 200 hybrid cells examined in each experiment. This indicated that the GM1662 genome was able to correct the RS effect. The presence of the RS effect in a few of the hybrid cells was attributed to the unstable karyotype resulting from pSV3 transfection which presumably caused the loss of the normal allele(s) of the RS gene in these hybrid cells.
Somat Cell Mol Genet 1987 May
PMID:Somatic cell hybridization of Roberts syndrome and normal human fibroblasts transfected with plasmids carrying dominant selection markers. 347 85

The feasibility of using retroviral gene therapy to overcome drug resistance was assessed by determining the efficiency by which a retrovirus containing the human HGPRT gene could sensitize hypoxanthine-guanine phosphoribosyltransferase (HGPRT) negative human promyelocytic leukemia cells to 6-thioguanine. A single three-hour exposure at a virus to cell ratio of 6 X 2:1 restored sensitivity to 70(+/- 18)% of the clonogenic cells. The efficacy varied as a function of virus concentration and duration of viral exposure; the time allowed for integration and expression between one and five days post-infection had little effect. Cells successfully sensitized contained a proviral insert and expressed HGPRT activity that ranged from 1 to 92% of that in the wild-type cells. The mutation rate of the inserted gene varied from the same as that of the endogenous HGPRT gene to 200-fold greater in different clones. Failure of sensitization following viral exposure was associated with absence of an integrated provirus, and clonogenic cells failing to be sensitized by one virus exposure were sensitized with approximately the same efficiency by a second viral exposure. These results demonstrate the feasibility of transferring a drug sensitivity gene to a human leukemia cell line.
Mol Biol Med 1987 Jun
PMID:Gene therapy for thioguanine-resistant human leukemia. 347 17

Treatment of hypoxanthine-guanine phosphoribosyltransferase (HGPRT)-deficient human promyelocytic leukemia (HL-60) cells with 6-thioguanine results in growth inhibition and cell differentiation. 6-Thioguanine is a substrate for the tRNA modification enzyme tRNA-guanine ribosyltransferase, which normally catalyzes the exchange of queuine for guanine in position 1 of the anticodon of tRNAs for asparagine, aspartic acid, histidine, and tyrosine. During the early stages of HGPRT-deficient HL-60 cell differentiation induced by 6-thioguanine, there was a transient decrease in the queuine content of tRNA, and changes in the isoacceptor profiles of tRNA(His) indicate that 6-thioguanine was incorporated into the tRNA in place of queuine. Reversing this structural change in the tRNA anticodon by addition of excess exogenous queuine reversed the 6-thioguanine-induced growth inhibition and differentiation. Similar results were obtained when 8-azaguanine (another inhibitor of queuine modification of tRNA that can be incorporated into the anticodon) replaced 6-thioguanine as the inducing agent. The data suggest a primary role for the change in queuine modification of tRNA in mediating the differentiation of HGPRT-deficient HL-60 cells induced by guanine analogs.
Mol Cell Biol 1987 Oct
PMID:Guanine analog-induced differentiation of human promyelocytic leukemia cells and changes in queuine modification of tRNA. 347 81

Hypoxanthine-guanine phosphoribosyltransferase (EC 2.4.2.8) was isolated from the malarial parasite, Plasmodium lophurae. The apparent pI, as determined by chromatofocusing, was 7.6. The native molecular weight was 79,000. The pH profile of HGPRT exhibited a broad pH optimum. With hypoxanthine as substrate maximal activity was achieved from pH 6.0-10.0, and with guanine as substrate maximal activity occurred from pH 7.5-9.5. The enzyme exhibited Michaelis-Menten kinetics with all substrates. The Km values were 3.8 microM (hypoxanthine), 2.4 microM (guanine), 6.2 microM (6-mercaptopurine), 7.6 microM (6-thioguanine), and 360 microM (8-azahypoxanthine). 6-Thioinosine, 9-beta-arabinofuranosylhypoxanthine, 6-chloropurine, xanthine and azaguanine were inhibitors of the P. lophurae enzyme. From the substrate and inhibitor data it appears that the sixth position on the purine ring plays a major role in enzyme activity.
Mol Biochem Parasitol 1987 Feb
PMID:Purification of hypoxanthine-guanine phosphoribosyltransferase of Plasmodium lophurae. 357 49

Somatic cell genetic analysis of purine base transporters in mouse S49 cells has demonstrated the existence of a unique high-affinity purine base transporter, which is mutationally expressed and is not found in wild-type S49 cells or any other cells of the animal kingdom (B. Aronow, et al. (1986) Mol. Cell. Biol. 6, 2957). In order to determine whether this nucleobase transport system is active and concentrative, a secondary mutation in hypoxanthine-guanine phosphoribosyltransferase (HGPRTase) was inserted into the cell line expressing this novel base transporter. The HGPRTase-deficient cells were capable of transporting hypoxanthine at increased rates but did not accumulate the base to concentrations in excess of that in the culture medium. Moreover, neither sodium azide nor ouabain had significant effects on hypoxanthine transport rates, indicating that energy metabolism and the maintenance of a sodium gradient were not required for transport function. These studies suggest that the novel mutationally expressed base transporter is independent of subsequent metabolism and does not require energy or a functioning Na+-K+-dependent ATPase activity.
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PMID:Genetic demonstration that the mutationally expressed nucleobase transporter of mouse S49 cells is nonconcentrative. 362 35

Material was introduced into cultures of cells by using the method of scrape loading, in which cells are simply rubbed from the surface of a plastic tissue culture dish by a rubber-tipped rod in the presence of a macromolecule of interest. The volume of solution introduced into cells was comparable to that generally injected in the direct microinjection method with glass capillaries, that is, about 50 to 100 fl per cell. Genetic defects (lack of hypoxanthine-guanine phosphoribosyltransferase and thymidine kinase) in several cell lines were transiently corrected by scraping the cells in the presence of crude cell extracts prepared from wild-type cells.
Mol Cell Biol 1987 Aug
PMID:Transient correction of genetic defects in cultured animal cells by introduction of functional proteins. 367 Mar 4


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