Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: UNIPROT:P00492 (hypoxanthine-guanine phosphoribosyltransferase)
 2,385 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The Lesch-Nyhan syndrome is an X-linked disorder caused by a virtually complete absence of the key enzyme of purine recycling, hypoxanthine-guanine phosphoribosyltransferase (HPRT). It is characterized by uric acid overproduction and severe neurological dysfunction. No treatment is yet available for the latter symptoms. A possible long-term solution is gene therapy, and recombinant adenoviruses have been proposed as vectors for gene transfer into postmitotic neuronal cells. We have constructed an adenoviral vector expressing the human HPRT cDNA under the transcriptional control of a short human cytomegalovirus major immediate early promoter (RAd-HPRT). Here we show that infection of human 1306, HPRT-negative cells with RAd-HPRT, expressed high enough levels of HPRT enzyme activity, as to reverse their abnormal biochemical phenotype, thus enhancing hypoxanthine incorporation and restoring purine recycling, increasing GTP levels, decreasing adenine incorporation, and allowing cell survival in HAT medium in which only cells expressing high levels of HPRT can survive. Infection of murine STO cells, increased hypoxanthine incorporation and restored purine recycling, thus allowing cell survival in HAT medium, and reduced de novo purine synthesis. Although both cells were able to survive in HAT medium post infection with RAd-HPRT, some of the biochemical consequences differed. In summary, even though adenoviral vectors do not integrate into the genome of target HPRT-deficient human or murine cells, RAd-HPRT mediated enzyme replacement corrects abnormal purine metabolism, increases intracellular GTP levels, and allows cells to survive in a negative selection medium.
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PMID:Adenoviruses encoding HPRT correct biochemical abnormalities of HPRT-deficient cells and allow their survival in negative selection medium. 1085 May 48

Marked differences between the mutagenic efficiency of N-methyl-N-nitrosourea (MNU), a potent carcinogen, methyl methane sulphonate (MMS) and dimethyl sulphate (DMS), both weak carcinogens, have been reported at the hypoxanthine-guanine phosphoribosyltransferase (HGPRT) and ouabain loci in V79 cells. Differences in levels of O6-guanine methylation produced by these alkylating agents, has been interpreted as indicating that O6-methylguanine is the DNA lesion specifically responsible for their mutagenic and carcinogenic effects. Because of the heterogeneity of molecular events which can result in forward mutation this conclusion seems unjustified. The development and characterisation of a reverse assay from 6-thioguanine resistance and HAT medium sensitivity (TG(R) and HAT(S)), to 6-thioguanine sensitivity and HAT medium resistance (TG(S) and HAT(R)) HGPRT(-)-->HGPRT+ in V79 cells, has allowed us to test the above hypothesis in a more specific way. Ethyl methane sulphonate, a weak carcinogen and MNU, both of which produce significant levels of O atom alkylation, were similarly effective mutagens in the reverse direction. At equitoxic doses, DMS was 40-60 fold less efficient. There was however, no quantitative correlation between numbers of revertants induced and measured levels of O6-alkylguanine. From these and other observations it is concluded that O6-alkylguanine is not the only potentially mutagenic lesion in mammalian cells.
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PMID:Evidence for the involvement of lesions other than O6-alkylguanine in mammalian cell mutagenesis. 1121 71

Targeted correction of a single base in a gene of an eucaryotic cell by specific oligonucleotides is a yet controversial technique. Here, we introduce the correction of point mutations in the hypoxanthine-guanine-phosphoribosyl-transferase (HPRT) gene as an additional model system to test targeted gene correction. In human, Hprt mutations cause Lesch-Nyhan syndrome. Using hamster V79 cells, we generated three cell lines with one hprt point mutation each. These cell lines were treated with specific single-stranded 45 base phosphothioate modified oligonucleotides and selected by HAT medium. The surviving clones were investigated for the correction of the respective hprt mutation. Treatment with the oligonucleotides was successful in repairing all three hprt mutations (hprt cDNA position 74, C --> T; position 151, C --> T; and position 400, G --> A). The correction efficiency was very low but reproducible. We suggest that this system allows one to investigate targeted gene correction in dependence on the target sequence and the oligonucleotides used.
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PMID:Targeted gene correction of hprt mutations by 45 base single-stranded oligonucleotides. 1247 Jun 47

The repair of point mutations in hprt gene by single-stranded oligonucleotides represents a model to test targeted nucleotide exchange. We studied the concurrent nucleotide exchange of two or three nucleotides in the hprt deficient hamster cell line V79-151. The used oligonucleotides resulted in mismatches at two (151, 159) or three (151, 144, and 159) hprt positions. The hprt point mutation at position 151 was repaired in about 2/10(6) cells as shown by hprt sequencing in clones surviving HAT selection. The second nucleotide exchange at hprt position 159 was found in 7% of these HAT selected clones. Using oligonucleotides resulting in three mismatches, 29% of the clones showed nucleotide exchanges at the two hprt positions (151, 144) and about 4% at three positions (151, 144, and 159). These results indicate that single-stranded oligonucleotides can generate two or three nucleotide exchanges in a mammalian chromosomal gene.
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PMID:Concurrent targeted exchange of three bases in mammalian hprt by oligonucleotides. 1535 30


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