Chromosome painting analysis of spontaneous and methyl methanesulfonate-induced trifluorothymidine-resistant L5178Y cell colonies

Zhang, Lishi; Honma, Masamitsu; Matsuoka, Atsuko; Suzuki, Takayoshi; Sofuni, Toshio; Hayashi, Makoto

doi:10.1016/s0165-1218(96)00069-9

Cited by 15 publications

(7 citation statements)

References 31 publications

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“…We observed a similar phenomenon in a study of the tk gene mutation in L5178Y mouse lymphoma cells (51). Almost all TK-deficient L5178Y cells showed unbalanced translocations and other complicated rearrangements involving chromosome 11 (which harbors the mouse tk gene).…”

Section: Discussionsupporting

confidence: 69%

Illegitimate Recombination Leading to Allelic Loss and Unbalanced Translocation in p53-Mutated Human Lymphoblastoid Cells

Honma¹,

Zhang²,

Hayashi³

et al. 1997

Molecular and Cellular Biology

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Allelic loss and translocation are critical mutational events in human tumorigenesis. Allelic loss, which is usually identified as loss of heterozygosity (LOH), is frequently observed at tumor suppressor loci in various kinds of human tumors. It is generally thought to result from deletion or mitotic recombination between homologous chromosomes. In this report, we demonstrate that illegitimate (nonhomologous) recombination strongly contributes to the generation of allelic loss in p53-mutated cells. Spontaneous and X-ray-induced LOH mutations at the heterozygous thymidine kinase (tk) gene, which is located on the long arm of chromosome 17, from normal (TK6) and p53-mutated (WTK-1) human lymphoblastoid cells were cytogenetically analyzed by chromosome 17 painting. We observed unbalanced translocations in 53% of LOH mutants spontaneously arising from WTK-1 cells but none spontaneously arising from TK6 cells. We postulate that illegitimate recombination was occurring between nonhomologous chromosomes after DNA replication, leading to allelic loss and unbalanced translocations in p53-mutated WTK-1 cells. X-ray irradiation, which induces DNA double-strand breaks (DSBs), enhanced the generation of unbalanced translocation more efficiently in WTK-1 than in TK6 cells. This observation implicates the wild-type p53 protein in the regulation of homologous recombination and recombinational DNA repair of DSBs and suggests a possible mechanism by which loss of p53 function may cause genomic instability.

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Section: Discussionsupporting

confidence: 69%

Illegitimate Recombination Leading to Allelic Loss and Unbalanced Translocation in p53-Mutated Human Lymphoblastoid Cells

Honma¹,

Zhang²,

Hayashi³

et al. 1997

Molecular and Cellular Biology

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“…This assay was developed by Drs D.Clive and M.Moore (Clive and Spector, 1975;Clive et al, 1979;Moore-Brown et al, 1981) and the protocol has been optimized Moore and Howard, 1982;Cole et al, 1983;Turner et al, 1984;Majeska and Matheson, 1990). Some TK-deficient mutants in the MLA exhibit not only point mutations but also gross structural and numerical changes at the chromosomal level (Hozier et al, 1982;Moore et al, 1985;Blazak et al, 1989;Clive et al, 1990;Combes et al, 1995;Zhang et al, 1996), thus the MLA can detect a wide range of genetic damage, including gene mutations, larger scale chromosomal changes, recombination, aneuploidy and others. Most of these changes occur in tumors and are presumably relevant for carcinogenesis.…”

Section: Introductionmentioning

confidence: 99%

Evaluation of the mouse lymphoma tk assay (microwell method) as an alternative to the in vitro chromosomal aberration test

et al. 1999

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In order to evaluate the utility of the mouse lymphoma assay (MLA) for detecting in vitro clastogens and spindle poisons and to compare it with the in vitro chromosomal aberration test (CA), we conducted an international collaborative study of the MLA that included 45 Japanese laboratories and seven overseas laboratories under the cooperation of the Ministry of Health and Welfare of Japan and the Japanese Pharmaceutical Manufacturer's Association. We examined 40 chemicals; 33 were reportedly positive in the CA but negative in the bacterial reverse mutation assay, six were negative in both assays and one was positive in both. We assayed mutations of the thymidine kinase (TK) locus (tk) of L5178Y tk ϩ/-mouse lymphoma cells using the microwell method. According to our standard protocol, cells were exposed to the chemical for 3 h, cultured for 2 days and TK-deficient mutants were expressed in 96-well plates under trifluorothymidine. Each chemical was coded and tested by two or three laboratories. Among the 34 CA-positive chemicals, positive MLA results were obtained for 20 and negative results were obtained for nine. The remaining five chemicals were inconclusive or equivocal because of discrepant inter-laboratory results or reproduced discrepant results, respectively. Among the six CA-negative chemicals, one was negative in the MLA, two were positive and three were inconclusive. Thus, the MLA could detect only 59% (20/34) of CA-positive chemicals. We concluded that the MLA was not as sensitive as the CA. Some MLA-negative chemicals evoked positive responses in the CA only after long continuous treatment. These might also be genotoxic in the MLA with long continuous treatment. Improvement of the MLA protocol, including alteration of the duration of the treatment, might render the MLA as sensitive as the CA. IntroductionThe genotoxicity tests that have been developed and validated over the years differ in the biological system used (prokaryotic,

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“…Initially, detailed banded chromosome analysis of colonies selected from agar plates showed that a high percentage of small colony mutants have a wide variety of visible chromosome 11b aberrations, while large colonies do not, leading to the conclusion that large colonies are the result of single gene mutations, while small colonies represent chromosomal mutations (Hozier et al, 1981). However, more recently chromosome painting analysis of colonies selected using the microwell protocol (Zhang et al, 1996) demonstrated that 25% of large colonies did show rearranged chromosome 11, compared with 75% of small colonies, and that the majority of the chromosomal abnormalities involved complex rearrangements often involving an increase in chromosome 11 material. The NcoI restriction fragment length polymorphism (RFLP) analysis.…”

Section: Overview Of Mla Development and Assay Characterizationmentioning

confidence: 99%

“…Each chemical was tested blind by two or three laboratories. The results were compared with the historical database of in vitro CA tests, including the Chinese hamster lung fibroblast (CHL/IU) tests undertaken over a period of years by the Japanese (Matsuoka et al, 1996;Honma et al, 1999). Initial reports suggested that using the 'standard' MLA protocol of 3-4 h treatment time, while many clastogens were deemed 'positive', nine of 34 tested were negative.…”

Section: Ich4 and The Japanese Trial: Is A New Protocol Justified?mentioning

confidence: 99%

“…The mouse lymphoma assay (MLA) has been widely used for many years for determining the potential for chemicals to cause the full broad spectrum of mutational damage Mitchell et al, 1997). Substantial research has been conducted to understand the capabilities (and limitations) of the assay, its proper conduct and the appropriate interpretation of test data (Clive et al, 1979(Clive et al, , 1990Hozier et al, 1981Hozier et al, , 1982Hozier et al, , 1989Moore-Brown, 1981;Cole et al, 1983Cole et al, , 1990Cole et al, , 1991Turner et al, 1984;Moore et al, 1985aMoore et al, ,b, 1989Blazak et al, 1986;Doerr et al, 1989;Applegate et al, 1990;Moore and Doerr, 1990;Liechty et al, 1993Liechty et al, , 1994Liechty et al, , 1996Zhang et al, 1996;Mitchell et al, 1997;Clark et al, 1998).…”

Section: Introductionmentioning

confidence: 99%

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The mouse lymphoma assay in the wake of ICH4where are we now?

1999

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Chromosome painting analysis of spontaneous and methyl methanesulfonate-induced trifluorothymidine-resistant L5178Y cell colonies

Cited by 15 publications

References 31 publications

Illegitimate Recombination Leading to Allelic Loss and Unbalanced Translocation in p53-Mutated Human Lymphoblastoid Cells

Illegitimate Recombination Leading to Allelic Loss and Unbalanced Translocation in p53-Mutated Human Lymphoblastoid Cells

Evaluation of the mouse lymphoma tk assay (microwell method) as an alternative to the in vitro chromosomal aberration test

The mouse lymphoma assay in the wake of ICH4where are we now?

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