Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Recombinant DNA techniques have been used to develop Chinese hamster ovary cell lines for studying chemically induced genotoxicity. These cell lines express a specific cytochrome P450 isozyme responsible for the metabolism of polycyclic aromatic hydrocarbons and exhibit defined differences in DNA repair capacity. A bacterial gene (neo) conferring resistance to gentamicin was inserted into the pcD expression vector containing the mouse cytochrome P1450 (P450IA1) cDNA to facilitate the selection of transformed cells. This plasmid was introduced into the nucleotide-excision-repair-deficient UV5 cell line by electroporation. Transformed clonal isolates expressing the P1450 cDNA were identified by differential cytotoxicity assays using benzo[a]pyrene (B[a]P). One such clone, termed UV5P1, was mutagenized with ethyl methanesulfonate and selected for resistance to killing by UV radiation to derive a repair-competent clone that expresses similar P1450 activity to that of the parental cell line. Two repair-competent clones were selected and called 5P1R1 and 5P1R3. The resulting cell lines (UV5P1, 5P1R1, and 5P1R3) expressed arylhydrocarbon hydroxylase activity. UV5P1 and 5P1R3 were compared in terms of cytotoxicity and mutagenicity after exposure to B[a]P. Induced mutations were measured at the adenine phosphoribosyltransferase (aprt) and hypoxanthine guanine phosphoribosyltransferase (hprt) loci. Repair-deficient UV5P1 cells were killed by B[a]P at concentrations below 0.1 microM, while the repair-proficient 5P1R3 cells showed no toxicity up to 60 microM. Mutation induction at both loci was also much more efficient in UV5P1 cells. These new cell lines provide a more sensitive system that can be used in a battery of assays to evaluate the genotoxicity of chemicals requiring P450IA1 metabolic activation and to assess the role of DNA repair in modulating the biological effects of DNA damage.
Mol Carcinog 1991
PMID:Expression of mouse cytochrome P450IA1 cDNA in repair-deficient and repair-proficient CHO cells. 179 88

The genotoxicity of the terpene beta-myrcene was evaluated in mammalian cells in vitro. Myrcene is the major constituent of oil of bay and hop which are used in the manufacture of alcoholic beverages. Myrcene is also present in lemon grass (Cymbopogon citratus), a plant widely used in Brazilian folk medicine. Recently, it was shown that myrcene is a very potent analgesic substance and might be an alternative to the already available analgesic drugs. Myrcene was tested up to 1,000 micrograms/ml (limit of solubility) in the presence and absence of S9-mix and did not induce chromosome aberrations and sister chromatid exchanges (SCEs) in human lymphocytes in vitro. Neither the mitotic index nor the proliferation index was influenced by the myrcene treatment. Myrcene did not cause increased mutation frequencies at the hprt-locus in V79-cells. Tests with and without S9-mix revealed negative results. There was no indication for induced cytotoxicity. However, myrcene reduced the SCE-inducing effect of cyclophosphamide in human lymphocytes in a dose dependent manner and also reduced the toxic and mutagenic effect of cyclophosphamide in V79-cells. Under the same test conditions, SCE induction by ethyl methanesulfonate (EMS) and benzo [a]pyrene (BP) was not significantly influenced by simultaneous myrcene treatment. The in vitro results show that myrcene is not mutagenic in mammalian cells, but has antimutagenic properties. The possibility that myrcene exerts its antimutagenic activity by inhibiting certain forms of the cytochrome P-450 isoenzymes required for activation of premutagens and precarcinogenes is discussed.
Environ Mol Mutagen 1991
PMID:Evaluation of the mutagenicity of beta-myrcene in mammalian cells in vitro. 186 66

In order to determine the relationships among the reduction in relative cloning efficiency (RCE), sister-chromatid exchange (SCE) formation, and interference with progression through the cell-cycle, human teratocarcinoma-derived (P3) cells were exposed to either ethyl methanesulfonate or to methyl methanesulfonate. The relationship between SCE and toxicity was quantified, the progression through the cell-cycle was evaluated with flow cytometric methods, and the effects of these chemicals on cell growth and average generation time (AGT) were determined. A strong correlation existed between RCE and SCE (r = -0.978, p less than .001) which was accompanied by an inhibition of growth as evidenced by a significant (p less than .0001) negative linear effect of concentration on the relative cell count from 24 to 72 hours after exposure and by a concentration-dependent increase (p less than .0001) in the AGT. Delays in the transit through S-phase were evident 4 hours after exposure to toxic concentrations of either carcinogen and by 8 to 12 hours post-exposure at the lower concentrations. Increases in the percentage of nuclei in G2 + M, indicative of G2 arrest, occurred from 12 to 24 hours after exposure. One interpretation of these results is that those effects of EMS and MMS exposure which result in S-phase delay and G2 arrest may be those elements common to the induction of SCE and cellular toxicity.
Environ Mol Mutagen 1991
PMID:Flow cytometric evaluation of cell-cycle progression in ethyl methanesulfonate and methyl methanesulfonate-exposed P3 cells: relationship to the induction of sister-chromatid exchanges and cellular toxicity. 187 6

The vermilion gene was used as a target to determine the mutational specificity of ethyl methanesulfonate (EMS) in germ cells of Drosophila melanogaster. To study the impact of DNA repair on the type of mutations induced, both excision-repair-proficient (exr+) and excision-repair-deficient (exr-) strains were used for the isolation of mutant flies. In all, 28 mutants from the exr+ strain and 24 from the exr- strain, were characterized by sequence analysis. In two mutants obtained from the exr+ strain, small deletions were observed. All other mutations were caused by single base-pair changes. In two mutants double base-pair substitutions had occurred. Of the mutations induced in the exr+ strain, 22 (76%) were GC----AT transitions, 3 (10%) AT----TA transversions, 2 (6%) GC----TA transversions and 2 (6%) were deletions. As in other systems, the mutation spectrum of EMS in Drosophila is dominated by GC----AT transitions. Of the mutations in an exr- background, 12 (48%) were GC----TA transitions, 7 (28%) AT----TA transversions, 5 (20%) GC----TA transversions and 1 (4%) was a AT----GC transition. The significant increase in the contribution of transversion mutations obtained in the absence of an active maternal excision-repair mechanism, clearly indicates efficient repair of N-alkyl adducts (7-ethyl guanine and 3-ethyl adenine) by the excision-repair system in Drosophila germ cells.
Mol Gen Genet 1991 Oct
PMID:Mutational specificity of ethyl methanesulfonate in excision-repair-proficient and -deficient strains of Drosophila melanogaster. 192 71

We have developed a plasmid-based system for isolating gene rearrangements in mammalian cells by selection for reversion of a promoterless drug resistance gene. pNH4 contains the selectable marker gene neo under the control of the herpes simplex virus, thymidine kinase (tk) promoter and, upstream and in the opposite orientation, a dormant promoterless hygromycin B resistance gene (hph) that can be expressed following rearrangement events. An NIH 3T3 cell line stably transfected with pNH4 that has a spontaneous frequency of generation of Hphr colonies of approximately 10(-8) was isolated. Treatment of this line with ethyl methanesulfonate raised the frequency of Hphr colony formation approximately 100-fold. Approximately 60% (21 of 35) of ethyl methanesulfonate-induced Hphr clones showed rearrangements detectable by Southern blot analysis within a 40-kb region surrounding the integrated construct, including a nonhomologous recombination event and, possibly, a large insertion. Additionally, three Hphr clones showed evidence of gene amplification. Northern (RNA) blot analysis of hph mRNA suggests that the rearrangements may provide a function that allows the tk promoter to initiate transcription off the opposite strand, thus yielding hph transcripts. Cell lines harboring pNH4, or modifications of it, may be valuable for studying recombination mechanisms responsible for the various types of genetic rearrangements found in cancer cells.
Mol Cell Biol 1991 Aug
PMID:A novel, plasmid-based system for studying gene rearrangements in mammalian cells. 207 98

The HPR1 gene has been cloned by complementation of the hyperrecombination phenotype of hpr1-1 strains by using a color assay system. HPR1 is a gene that is in single copy on chromosome IV of Saccharomyces cerevisiae, closely linked to ARO1, and it codes for a putative protein of 752 amino acids (molecular mass, 88 kilodaltons). Computer searches revealed homology (48.8% conserved homology; 24.8% identity) with the S. cerevisiae TOP1 gene in an alpha-helical stretch of 129 amino acids near the carboxy-terminal region of both proteins. The ethyl methanesulfonate-induced hpr1-1 mutation is a single-base change that produces a stop codon at amino acid 559 coding for a protein that lacks the carboxy-terminal TOP1 homologous region. Haploid strains carrying deletions of the HPR1 gene show a slightly reduced mitotic growth rate and extremely high rates of intrachromosomal excision recombination (frequency, 10 to 15%) but have a undetectable effect on rDNA recombination. Double-null mutants hpr1 top1 grow very poorly. We conclude that Hpr1 is a novel eucaryotic protein, mutation of which causes an increase in mitotic intrachromosomal excision recombination, and that it may be functionally related to an activity of the topoisomerase I protein.
Mol Cell Biol 1990 Apr
PMID:HPR1, a novel yeast gene that prevents intrachromosomal excision recombination, shows carboxy-terminal homology to the Saccharomyces cerevisiae TOP1 gene. 218 Dec 75

Colony probe oligonucleotide hybridisation was used for the unambiguous identification of DNA alterations and the determination of distributions and frequencies of forward mutation at the molecular level. To demonstrate the reliability and versatility of this technique, distributions of spontaneous and ethyl methanesulfonate (EMS)-induced mutations have been reproduced using a battery of oligonucleotide probes complementary to specific sites and classes of mutation. These studies are indicative of the diagnostic potential of oligonucleotide colony hybridisation to the characterisation of mutation; oligonucleotide hybridisation used in conjunction with a well studied mutational target provides a rapid and reliable alternative to DNA sequencing for the characterisation of all classes of mutations.
Environ Mol Mutagen 1990
PMID:Colony hybridisation in Escherichia coli: a rapid procedure for determining the distribution of specific classes of mutations among a number of preselected sites. 220 71

We describe the cloning and function of the human XRCC1 gene, which is the first mammalian gene isolated that affects cellular sensitivity to ionizing radiation. The CHO mutant EM9 has 10-fold-higher sensitivity to ethyl methanesulfonate, 1.8-fold-higher sensitivity to ionizing radiation, a reduced capacity to rejoin single-strand DNA breaks, and a 10-fold-elevated level of sister chromatid exchange compared with the CHO parental cells. The complementing human gene was cloned from a cosmid library of a tertiary transformant. Two cosmid clones produced transformants that showed approximately 100% correction of the repair defect in EM9 cells, as determined by the kinetics of strand break repair, cell survival, and the level of sister chromatid exchange. A nearly full-length clone obtained from the pcD2 human cDNA expression library gave approximately 80% correction of EM9, as determined by the level of sister chromatid exchange. Based on an analysis of the nucleotide sequence of the cDNA insert compared with that of the 5' end of the gene from a cosmid clone, the cDNA clone appeared to be missing approximately 100 bp of transcribed sequence, including 26 nucleotides of coding sequence. The cDNA probe detected a single transcript of approximately 2.2 kb in HeLa polyadenylated RNA by Northern (RNA) blot hybridization. From the open reading frame and the positions of likely start sites for transcription and translation, the size of the putative XRCC1 protein is 633 amino acids (69.5 kDa). The size of the XRCC1 gene is 33 kb, as determined by localizing the endpoints on a restriction endonuclease site map of one cosmid clone. The deduced amino acid sequence did not show significant homology with any protein in the protein sequence data bases examined.
Mol Cell Biol 1990 Dec
PMID:Molecular cloning of the human XRCC1 gene, which corrects defective DNA strand break repair and sister chromatid exchange. 224 54

It has been shown by genetic complementation analysis that a mitomycin C-sensitive mutant (V-H4) of Chinese hamster V79 cells is the first rodent equivalent of Fanconi anemia (FA) group A. The V-H4 mutant shows many typical characteristics of cells derived from FA patients. V-H4 cells exhibit increased sensitivity towards cross-linking agents as MMC (approximately 30-fold), cis-DDP (approximately 10-fold), DEB (approximately 10-fold), and PUVA (approximately 1.6-fold), but an only slightly increased sensitivity to monofunctional alkylating agents (EMS and MMS) and actinomycin D. V-H4 cells are also moderately sensitive to adriamycin (1.6-fold), and not sensitive to H2O2. The levels of chromosomal aberrations induced by MMC and cis-DDP treatment are higher (4- to 6-fold) in V-H4 cells than in the wild-type V79 cells. Genetic complementation analysis with other Chinese hamster mutants hypersensitive to MMC (irs1, irs1SF, UV20 and UV41) indicates clearly that V-H4 belongs to a different, new complementation group. This unique mutant is very stable and can serve as a vehicle to isolate the complementing FA-A gene from normal human DNA.
Somat Cell Mol Genet 1990 Nov
PMID:The Chinese hamster V79 cell mutant V-H4 is phenotypically like Fanconi anemia cells. 226 31

Data presented in this paper deal with a further molecular characterization of 2 out of 32 EMS-induced Arabidopsis ADH null mutants that we isolated previously. In order to localize and characterize each mutation at the molecular level, we have cloned and completely sequenced the R002 and R006 null mutant alleles. For mutant R002, which does not contain any detectable levels of ADH protein and mRNA, we have found that the mutation is due to a single C to T base pair substitution in the reading frame; this leads to the incorporation of a TAG stop codon (amber nonsense mutation). For mutant R006, which contains normal levels of inactive protein and mRNA levels, we found a G to A base pair transition. This gives rise to a Cys to Tyr amino acid substitution in the active site of the ADH enzyme.
Mol Gen Genet 1990 Nov
PMID:Sequence analysis of two null-mutant alleles of the single Arabidopsis Adh locus. 227 48


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