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1.
Lenarczyk M Ueno A Vannais DB Kraemer S Kronenberg A Roberts JC Tatsumi K Hei TK Waldren CA 《Radiation research》2003,160(5):579-583
We are carrying out studies aimed at reducing the mutagenic effects of high-LET 56Fe ions and 12C ions (56Fe ions, 143 keV/microm; 12C ions, 100 keV/microm) with certain drugs, including RibCys [2-(R,S)-D-ribo-(1',2',3',4'-tetrahydroxybutyl)-thiazolidine-4(R)-carboxylic acid]. RibCys, formed by condensation of L-cysteine with D-ribose, is designed so that the sulfhydryl amino acid L-cysteine is released intracellularly through nonenzymatic ring opening and hydrolysis leading to increased levels of glutathione (GSH). RibCys (4 or 10 mM), which was present during irradiation and for a few hours after, significantly decreased the yield of CD59- mutants induced by radiation in AL human-hamster hybrid cells. RibCys did not affect the clonogenic survival of irradiated cells, nor was it mutagenic itself. These results, together with the minimal side effects reported in mice and pigs, indicate that RibCys may be useful, perhaps even when used prophylactically, in reducing the mutation load created by high-LET radiation in astronauts or other exposed individuals. 相似文献
2.
Wedemeyer N Greve B Uthe D Pötter T Denklau D Severin E Hacker-Klom U Köhnlein W Göhde W 《Mutation research》2001,473(1):73-84
Determination of the genotoxic effects of ionizing radiation, especially at low-doses, is of great importance for risk assessment, e.g. in radiological diagnostics. The human-hamster hybrid A(L) cell line has been shown previously to be a well-suited in vitro model for the study of mutations induced by various mutagens. The A(L) cells contain a standard set of hamster chromosomes and a single human chromosome 11, which confers the expression of the human cell surface protein CD59. Using CD59 specific antibodies, cells mutated in the CD59 gene can be detected and quantified by the loss of the cell surface marker. In contrast to previous studies, prior to irradiation we removed spontaneous mutants by magnetic cell separation (MACS) which allows analysis of radiation-induced mutation events only. We exposed A(L) cells to 100kV X-rays at 0.1 to 5Gy. The proportions of X-irradiation-induced CD59(-) mutants were quantified by flow cytometry after immunofluorescence labeling. Between 0.2 and 5Gy the yield of CD59 mutants was a linear function of dose. The molecular analysis of individual CD59-negative clones induced after exposure of 1, 3 and 5Gy of X-ray revealed a dose-dependent linear increase of large deletions (>6Mbp), whereas, point mutations could be seen only in spontaneous CD59 mutants or after low-dose exposure (< or =1Gy). We conclude that the modified A(L) assay presented here is appropriate for detection and quantification of non-lethal DNA lesions induced by low-dose ionizing radiation. 相似文献
3.
The flow cytometry mutation assay is based on detecting mutations in the CD59 gene on human chromosome 11 in CHO A(L) cells with flow cytometry, but the kinetics of mutant expression and the histogram region for mutant selection have not been studied in detail. CHO A(L) cells were analyzed by flow cytometry for CD59 expression at various times after irradiation. The mutant fraction increased to a maximum at day 6 but decreased to near background levels by day 20. Cells were sorted from six different regions on the CD59 histograms after irradiation. The growth rate was similar for cells from all regions, and the surviving fraction was 50% of that for control cells. By 14 days the CD59 expression of cells from regions 2-5 was reduced to that of region 1. Cells were also analyzed for simultaneous expression of CD59, CD44 and CD90 (all on chromosome 11) to roughly characterize the size of the mutations. Triple mutants from the sorted populations were reduced from 41% on day 6 to 8% on day 24. We conclude that the mutant region should be increased to include cells with intermediate CD59 expression; also, the loss of CD59 mutant expression over time could be explained in part by the loss of triple mutants from the population. 相似文献
4.
The effects of the radioprotector 2-[(aminopropyl)amino] ethanethiol (WR-1065) on radiation-induced cell killing and mutagenesis at the hypoxanthine-guanine phosphoribosyl transferase (HGPRT) locus in V79 Chinese hamster cells under hypoxic or aerobic conditions were examined. Conditions of acute hypoxia were attained by gassing 10(6) cells in 1-ml volumes in individual glass ampoules for 2 min with nitrogen. Ampoules were then sealed and incubated at 37 degrees C for 60 min. Following this treatment, cell survival after irradiation as expected was significantly enhanced. The effect of acute hypoxia on the formation of HGPRT mutants by irradiation was also investigated. Mutation frequencies were determined with a 6-day expression time and corrected for the number of spontaneous background mutants. Although mutation induction was approximately linear as a function of radiation dose under most conditions tested, it was significantly reduced in cell populations made acutely hypoxic prior to irradiation. Protection against mutation induction was apparent and similar when cells were irradiated in the presence of the radioprotector, regardless of whether they were also hypoxic or aerated. If cells were irradiated in air and then made hypoxic, no significant protection was still observed. These results suggest that the antimutagenic effect of WR-1065 is not due solely to its ability to scavenge radiation-induced oxygen-free radicals, but rather that it may also modulate these effects through the scavenging of metabolically induced free radicals and/or the chemical repair of radiation-induced DNA lesions. 相似文献
5.
Takahashi E Fujita K Kamataki T Arimoto-Kobayashi S Okamoto K Negishi T 《Mutation research》2002,500(1-2):147-156
Expanded simple tandem repeat (ESTR) loci include some of the most unstable DNA in the mouse genome and have been extensively used in pedigree studies of germline mutation. We now show that repeat DNA instability at the mouse ESTR locus Ms6-hm can also be monitored by single molecule PCR analysis of genomic DNA. Unlike unstable human minisatellites which mutate almost exclusively in the germline by a meiotic recombination-based process, mouse Ms6-hm shows repeat instability both in germinal (sperm) DNA and in somatic (spleen, brain) DNA. There is no significant variation in mutation frequency between mice of the same inbred strain. However, significant variation occurs between tissues, with mice showing the highest mutation frequency in sperm. The size spectra of somatic and sperm mutants are indistinguishable and heavily biased towards gains and losses of only a few repeat units, suggesting repeat turnover by a mitotic replication slippage process operating both in the soma and in the germline. Analysis of male mice following acute pre-meiotic exposure to X-rays showed a significant increase in sperm but not somatic mutation frequency, though no change in the size spectrum of mutants. The level of radiation-induced mutation at Ms6-hm was indistinguishable from that established by conventional pedigree analysis following paternal irradiation. This confirms that mouse ESTR loci are very sensitive to ionizing radiation and establishes that induced germline mutation results from radiation-induced mutant alleles being present in sperm, rather than from unrepaired sperm DNA lesions that subsequently lead to the appearance of mutants in the early embryo. This single molecule monitoring system has the potential to substantially reduce the number of mice needed for germline mutation monitoring, and can be used to study not only germline mutation but also somatic mutation in vivo and in cell culture. 相似文献
6.
Evidence that most radiation-induced HPRT mutants are generated directly by the initial radiation exposure. 总被引:1,自引:0,他引:1
Radiation-induced HPRT mutants are generally assumed to arise directly from DNA damage that is misrepaired within a few hours after X-irradiation. However, there is the possibility that mutations result indirectly from radiation-induced genomic instability that may occur several days after the initial radiation exposure. The protocols that commonly employ a 5-7 day expression period to allow for expression of the mutant phenotype prior to replating for selection of mutants would not be able to discriminate between mutants that occurred initially and those that arose during or after the expression period. To address this question, we performed a fluctuation analysis in which synchronous or asynchronous populations of human bladder carcinoma cells were treated with single doses of X-irradiation. For comparison, radiation was delivered during the expression period, either from an initial dose of 1.0 Gy followed by two 1.0 Gy doses separated by 24 h or from disintegrations resulting from I125dU incorporated into DNA. The mutation frequency observed at the time of replating was used to calculate the average number of mutants in the initial irradiated culture by assuming that the mutants were induced directly at the time of irradiation. Then, this average number was used to calculate the fraction of the irradiated cultures that would be predicted by a Poisson distribution to have zero mutants. There was reasonably good agreement between the predicted poisson distribution and the observed distribution for the cultures that received single doses. Moreover, as expected, when cultures were irradiated during the expression period, the fraction of the cultures having zero mutants was significantly less than that predicted by a Poisson distribution. These results indicate that most radiation-induced HPRT mutations are induced directly by the initial DNA damage, and are not the result of radiation-induced instability during the 5-7 day expression period. 相似文献
7.
pSV2gpt-Transformed and wild-type Chinese hamster ovary (CHO) cell lines have been used to study radiation-induced mutation at the molecular level. The transformant, designated AS52, was constructed from a hypoxanthine-guanine phosphoribosyl transferase (HPRT)-deficient CHO cell line and contains a single, functional copy of the Escherichia coli xanthine-guanine phosphoribosyl transferase (XPRT) gene (gpt) stably integrated into the Chinese hamster genome. AS52 and wild-type CHO-K1-BH4 cells exhibit similar cytotoxic responses to uv light and X rays; however, significant differences occur in mutation induction at the gpt and hprt loci. A number of HPRT and XPRT mutants which arose following irradiation were analyzed by Southern-blot hybridization. Most XPRT (21/26) and all HPRT (23/23) mutants induced by uv light exhibited hybridization patterns indistinguishable from their parental cell lines. In contrast, all XPRT (26/26) and most HPRT mutants (15/21) induced by X irradiation contained deletion mutations affecting some or all of the gpt and hprt loci, respectively. These results indicate that X rays induce predominantly deletion mutations, while uv light is likely to induce point mutations at both loci. 相似文献
8.
BACKGROUND: A sensitive mammalian cell mutation assay was developed previously using a Chinese hamster ovary cell line (CHO A(L)) that stably incorporates human chromosome 11. The assay measures mutations in the CD59 gene on chromosome 11 but it requires the use of rabbit complement and colony growth for mutant selection. We have developed a more rapid flow cytometry-based mutation assay with CHO A(L) cells that uses monoclonal antibodies against CD59 to detect mutants and does not require colony formation. METHODS: CHO A(L) cells were treated with gamma-radiation or N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) and then allowed to grow for various times for mutant expression. Cells were labeled with monoclonal antibodies against CD59 and analyzed by flow cytometry. RESULTS: Negative and positive populations were separated by over 100-fold. Mixing various proportions of CD59-positive and -negative cells demonstrated that the assay is highly linear (r2 = 0.9999) and sensitive (<0.05% background mutants). The yield of CD59-inducible mutants was linearly related to dose for a clastogen (gamma-radiation) and point mutagen (MNNG). The mutant yield was time and treatment specific. CONCLUSIONS: Mutations induced by genotoxic agents can be rapidly and sensitively measured in CHO A(L) cells using flow cytometry. 相似文献
9.
Kraemer, S. M., Vannais, D. B., Kronenberg, A., Ueno, A. and Waldren, C. A. Gamma-Ray Mutagenesis Studies in a New Human-Hamster Hybrid, A(L)CD59(+/-), which has Two Human Chromosomes 11 but is Hemizygous for the CD59 Gene. Radiat. Res. 156, 10-19 (2001).We have developed a human-CHO hybrid cell line, named A(L)CD59(+/-), which has two copies of human chromosome 11 but is hemizygous for the CD59 gene and the CD59 cell surface antigen that it encodes. Our previous studies used the A(L) and A(L)C hybrids that respectively contain one or two sets of CHO chromosomes plus a single copy of human chromosome 11. The CD59 gene at 11p13.5 and the CD59 antigen encoded by it are the principal markers used in our mutagenesis studies. The hybrid A(L)CD59(+/-) contains two copies of human chromosome 11, only one of which carries the CD59 gene. The incidence of CD59 (-) mutants (formerly called S1(-)) induced by (137)Cs gamma rays is about fivefold greater in A(L)CD59(+/-) cells than in A(L) cells. Evidence is presented that this increase in mutant yield is due to the increased induction of certain classes of large chromosomal mutations that are lethal to A(L) cells but are tolerated in the A(L)CD59(+/-) hybrid. In addition, significantly more of the CD59 (-) mutants induced by (137)Cs gamma rays in A(L)CD59(+/-) cells display chromosomal instability than in A(L) cells. On the other hand, the yield of gamma-ray-induced CD59 (-) mutants in A(L)CD59(+/-) cells is half that of the A(L)C hybrid, which also tolerates very large mutations but has only one copy of human chromosome 11. We interpret the difference in mutability as evidence that repair processes involving the homologous chromosomes 11 play a role in determining mutant yields. The A(L)CD59(+/-) hybrid provides a useful new tool for quantifying mutagenesis and shedding light on mechanisms of genetic instability and mutagenesis. 相似文献
10.
11.
Spontaneous Arg- mutants arose at high frequencies in Streptomyces lividans. Exposure to ethidium bromide increased the frequency of arg instability. In Pro+ strains the induced arg mutants were mainly argG, but in the proB mutants, a new mutation, argJ, prevailed which lacked ornithine acetyltransferase activity and required ornithine for growth. Introduction of the cloned proB gene of Streptomyces coelicolor A3(2) into the proB argJ mutants not only complemented the proB mutation but also suppressed the argJ mutation. The proB mutation was also suppressed by adding ornithine to the medium. These results indicated crossfeeding(s) between the arginine and proline pathways in S. lividans, which presumably circumvented the detection of argJ mutations in Pro+ strains. 相似文献
12.
Evidence is presented for the mutation of the tryptophan-requiring bacterial strain Escherichia coli WP2 uvrA from auxotrophy to prototrophy, and from streptomycin sensitivity to resistance, by Cerenkov emission associated with 137Cs gamma irradiation. Furthermore, the data strongly suggest a more than additive interaction between the gamma-induced damage and that induced by Cerenkov emission for both mutations scored. An additional observation is that mutant yields (expressed as mutants/10(7) survivors) show a dependence on the number of viable cells plated for both uv (254 nm) and Cerenkov-induced mutations, but not for those induced by gamma irradiation. This demonstrates another similarity between uv- and Cerenkov-induced damage. 相似文献
13.
Characterization of DNA polymerase I*, a form of DNA polymerase I found in Escherichia coli expressing SOS functions 总被引:8,自引:0,他引:8
DNA polymerase I* is a form of the DNA polymerase I isolated from Escherichia coli which are expressing recA/lexA (SOS) functions. Induction of recA or polA1 cells by nalidixic acid does not result in the appearance of pol I*, but lexA or recA mutants that are constitutive for SOS functions constitutively express pol I* and mutants which lack functional recA protein produce pol I* when they carry a lexA mutation which renders the lexA repressor inoperative. Pol I* has been induced by nalidixic acid in dinA, dinD, dinF, and umuC mutants. Polymerase I* has a lower affinity for single-stranded DNA-agarose than polymerase I and it sediments through sucrose gradients in a dispersed manner between 6.6-10.5 S, whereas polymerase I sediments at 5 S. Whereas pol I* migrates significantly faster than pol I in nondenaturing polyacrylamide gels, the active polypeptide of both forms migrates at the same rate in denaturing polyacrylamide gels. Compared with polymerase I, polymerase I* has an enhanced capacity to incorporate the adenine analog, 2-amino-purine, into activated salmon sperm DNA and a relatively low fidelity in replicating synthetic polydeoxyribonucleotides. Both the 3'----5' (proofreading) and 5'----3' (nick-translational) exonuclease activities of pol I* and pol I are indistinguishable. Estimates of processivity give a value of approximately 6 for both forms of the enzyme. 相似文献
14.
Akira Tashibana Tadaaki Ohbayashi Hiraku Takebe Kouichi Tatsumi 《Mutation research》1990,230(2):159-166
We have characterized the structural changes in the hypoxanthine-guanine phosphoribosyltransferase (HPRT) gene of 14 UV-induced, 15 γ-ray-induced and 17 spontaneous mutants of human lymphoblastoid cells selected for 6-thioguanine (6TG) resistance. Southern blot analysis using the full-length HPRT cDNA as a probe revealed that 29% (5/17) of the spontaneous mutants contained detectable alterations in their restriction fragment patterns. Among the 15 mutants induced by γ rays, 7 (47%) had such alterations indicative of large deletions in the HPRT gene. In contrast, all 14 UV-induced mutants exhibited hybridization patterns indistinguishable from those of the wild-type cells. These results suggest that UV is likely to induce point mutations at the HPRT locus on the human chromosome and that the molecular mechanism of UV-induced mutation is quite different from that of ionizing radiation-induced mutation or spontaneous mutation in human cells. 相似文献
15.
Hiroyuki Ichida Ryouhei Morita Yuki Shirakawa Yoriko Hayashi Tomoko Abe 《The Plant journal : for cell and molecular biology》2019,98(2):301-314
Heavy‐ion beams have been widely utilized as a novel and effective mutagen for mutation breeding in diverse plant species, but the induced mutation spectrum is not fully understood at the genome scale. We describe the development of a multiplexed and cost‐efficient whole‐exome sequencing procedure in rice, and its application to characterize an unselected population of heavy‐ion beam‐induced mutations. The bioinformatics pipeline identified single‐nucleotide mutations as well as small and large (>63 kb) insertions and deletions, and showed good agreement with the results obtained with conventional polymerase chain reaction (PCR) and sequencing analyses. We applied the procedure to analyze the mutation spectrum induced by heavy‐ion beams at the population level. In total, 165 individual M2 lines derived from six irradiation conditions as well as eight pools from non‐irradiated ‘Nipponbare’ controls were sequenced using the newly established target exome sequencing procedure. The characteristics and distribution of carbon‐ion beam‐induced mutations were analyzed in the absence of bias introduced by visual mutant selections. The average (±SE) number of mutations within the target exon regions was 9.06 ± 0.37 induced by 150 Gy irradiation of dry seeds. The mutation frequency changed in parallel to the irradiation dose when dry seeds were irradiated. The total number of mutations detected by sequencing unselected M2 lines was correlated with the conventional mutation frequency determined by the occurrence of morphological mutants. Therefore, mutation frequency may be a good indicator for sequencing‐based determination of the optimal irradiation condition for induction of mutations. 相似文献
16.
氟氏链霉菌离子束注入突变谱的分析 总被引:1,自引:0,他引:1
用低能N+离子束注入转谷氨酰胺酶产生菌氟氏链霉菌后,通过试验,初步确定了注入的效应曲线,获得了一系列突变菌株。提取原始菌株和突变菌株的DNA,采用PCR反应分段扩增出转谷氨酰胺酶基因进行单链构象多态性分析(SSCP),并将特异性条带克隆测序进行基因突变型的鉴定,分析离子束注入引起链霉菌基因的基因突变类型及特点。结果显示:碱基变异的类型包括转换、颠换和缺失。在检测到的24个碱基突变中,主要是碱基的置换(87.5%),碱基缺失的比例比较小(12.5%)。在碱基置换中,转换的频率(58.3%)高于颠换的频率(29.2%)。转换主要以C→T,A→G为主,颠换以G→T,C→G为主。此外构成DNA的4种碱基均可以被离子束辐照诱发变异,其中胞嘧啶发生突变的频率较高。 相似文献
17.
K Sankaranarayanan 《Mutation research》1991,258(1):51-73
This paper reviews data on the nature of spontaneous and radiation-induced mutations in the mouse. The data are from studies using a variety of endpoints scorable at the morphological or the biochemical level and include pre-selected as well as unselected loci at which mutations can lead to recessive or dominant phenotypes. The loci used in the morphological recessive specific-locus tests permit the recovery of a wide spectrum of induced changes. Important variables that affect the nature of radiation-induced mutations (assessed primarily using tests for viability of homozygotes) include: germ cell stage, type of irradiation and the locus. Most of the results pertain to irradiated stem cell spermatogonia. The data on morphological specific-locus mutations show that overall, more than two-thirds of the X- or gamma-ray-induced mutations are lethal when homozygous. This proportion may be lower for those that occur spontaneously, but the numbers of tested mutants are small. For spontaneous mutations, there is evidence for the occurrence of mosaics and for proviral insertions. Most or all tested induced enzyme activity variants, dominant visibles (recovered in specific-locus experiments) and dominant skeletal mutations are lethal when homozygous and this is true of 50% of dominant cataract mutations, but again, the numbers of tested mutants are small. Electrophoretic mobility variants, which are known to be due to base-pair changes, are seldom induced by irradiation. At the histocompatibility loci, no radiation-induced mutations have been recovered, presumably because deletions are incompatible with survival even in heterozygotes. All these findings are consistent with the view that in mouse germ cells, most radiation-induced mutations are DNA deletions. Some mutations (in the morphological specific-locus tests) which had previously been inferred to be deletions on the basis of genetic analyses have now been shown to be DNA deletions by molecular methods. However, the possibility cannot be excluded that at least a small proportion of induced mutations may be intragenic changes. The data on the rates of induction of recessive lethals and of dominant skeletal and dominant cataract mutations (and proportions of the latter two which are homozygous lethal) can be used to estimate the proportions of recessive lethals which are expressed as skeletal abnormalities or cataracts. These calculations show that about 10% of recessive lethals manifest themselves as skeletal and less than 0.2% as cataract mutations.(ABSTRACT TRUNCATED AT 400 WORDS) 相似文献
18.
J Thacker 《Mutation research》1986,160(3):267-275
DNA from 58 independent HPRT-deficient mutants of V79 hamster cells induced by ionising radiation was analysed by Southern blot hybridization to a full-length hamster hprt cDNA. About half of the gamma-ray-induced mutants (20/43) were apparently total gene deletions, because they lacked all functional hprt gene sequences hybridizing to the cDNA probe. Another 10 mutants showed various partial deletions and/or rearrangements of the hprt gene. The remaining 13 mutants showed no detectable change in comparison to the structure of the normal gene, which correlated well with previous characterization of these mutants indicating that most carry point mutations in the hprt gene. However, it is probable that some of these point mutations occurred spontaneously rather than being radiation-induced. A smaller number of alpha-particle induced mutants gave similar results: out of a total of 15 mutants, 6 appeared to be total gene deletions, 5 had partial deletions and/or rearrangements, and 4 had no detectable changes. Thus, 70% or more of radiation-induced HPRT-deficient mutants arise through large genetic changes, especially deletions of all or part of the hprt gene. This result is to be contrasted with data published previously by ourselves and others indicating that the majority of spontaneous and ethyl methanesulphonate-induced mutations of hprt and similar genes arise by point mutation. 相似文献
19.
Mapping of mutations associated with neurovirulence in monkeys infected with Sabin 1 poliovirus revertants selected at high temperature. 总被引:15,自引:14,他引:1
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C Christodoulou F Colbere-Garapin A Macadam L F Taffs S Marsden P Minor F Horaud 《Journal of virology》1990,64(10):4922-4929
Poliovirus type 1 neurovirulence is difficult to analyze because of the 56 mutations which differentiate the neurovirulent Mahoney strain from the attenuated Sabin strain. We have isolated four neurovirulent mutants which differ from the temperature-sensitive parental Sabin 1 strain by only a few mutations, using selection for temperature resistance: mutant S(1)37C1 was isolated at 37.5 degrees C, S(1)38C5 was isolated at 38.5 degrees C, and S(1)39C6 and S(1)39C10 were isolated at 39.5 degrees C. All four mutants had a positive reproductive capacity at supraoptimal temperature (Rct+ phenotype). Mutant S(1)37C1 induced paralysis in two of four cynomolgus monkeys, and the three other mutants induced paralysis in four of four monkeys. The lesion score increased from the S(1)37C1 mutant to the S(1)39 mutants. To map the mutations associated with thermoresistance and neurovirulence, we sequenced all regions in which the Sabin 1 genome differs from the Mahoney genome. The S(1)37C1 mutant had one mutation in the 5' noncoding region and another in the 3' noncoding region. Mutant S(1)38C5 had these mutations plus another mutation in the 3D polymerase gene. The S(1)39 mutants had three additional mutations in the capsid protein region. The mutations were located at positions at which the Sabin 1 and Mahoney genomes differ, except for the mutation in the 5' noncoding region. The noncoding-region mutations apparently confer a low degree of neurovirulence. The 3D polymerase mutation, which distinguishes S(1)38C5 and S(1)39 mutants from S(1)37C1, is probably responsible for the high neurovirulence of S(1)38C5 and S(1)39 mutants. The capsid region mutations may contribute to the neurovirulence of the S(1)39 mutants, which was the highest among the mutants. 相似文献
20.
Ultraviolet irradiation of cells can induce a state of genomic instability that can persist for several cell generations after irradiation. However, questions regarding the time course of formation, relative abundance for different types of ultraviolet radiation, and mechanism of induction of delayed mutations remain to be answered. In this paper, we have tried to address these questions using the hypoxanthine phosphoribosyl transferase (HPRT) mutation assay in V79 Chinese hamster cells irradiated with ultraviolet A or B radiation. Delayed HPRT(-) mutations, which are indications of genomic instability, were detected by incubating the cells in medium containing aminopterin, selectively killing HPRT(-) mutants, and then treating the cells with medium containing 6-thioguanine, which selectively killed non-mutant cells. Remarkably, the delayed mutation frequencies found here were much higher than reported previously using a cloning method. Cloning of cells immediately after irradiation prevents contact between individual cell clones. In contrast, with the present method, the cells are in contact and are mixed several times during the experiment. Thus the higher delayed mutation frequency measured by the present method may be explained by a bystander effect. This hypothesis is supported by an experiment with an inhibitor of gap junctional intercellular communication, which reduced the delayed mutation frequency. In conclusion, the results suggest that a bystander effect is involved in ultraviolet-radiation-induced genomic instability and that it may be mediated in part by gap junctional intercellular communication. 相似文献