X rays induce interallelic homologous recombination at the human thymidine kinase gene.

Benjamin, Mark B.; Little, John B.

doi:10.1128/mcb.12.6.2730

Cited by 38 publications

(14 citation statements)

References 75 publications

(44 reference statements)

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“…The spontaneous recombination frequency in each of these lines was compared to that in a line carrying closely linked copies of the tk8 and tk26 alleles at the same genomic site. Although we did not recover any true spontaneous interchromosomal recombinants, given the number of cells we plated, our results are consistent with the low spontaneous interchromosomal recombination frequency at the human TK locus (14,15). We determined that the spontaneous frequency of interchromosomal recombination was <3.6 x 10-9, while the intrachromosomal recombination frequency at the identical genomic site was 6.7 x 10-5.…”

Section: Discussionsupporting

confidence: 63%

Spontaneous and restriction enzyme-induced chromosomal recombination in mammalian cells.

Godwin

Bollag

Christie

et al. 1994

Proc. Natl. Acad. Sci. U.S.A.

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We have derived Chinese hamster ovary (CHO) cell hybrids containing herpes simplex virus thymidine kinase (tk) heteroalleles for the study of spontaneous and restriction enzyme-induced interchromosomal recombination. These lines allowed us to make a direct comparison between spontaneous intrachromosomal and interchromosomal recombination using the same tk heteroalleles at the same genomic insertion site. We find that the frequency of interchromosomal recombination is less by a factor of at least 5000 than that of intrachromosomal recombination. Our results with mammalian cells differ markedly from results with Saccharomyces cerevisiae, with which similar studies typically give only a 10-to 30-fold difference. Next, to inquire into the fate of doublestrand breaks at either of the two different Xho I linker insertion mutations, we electroporated PaeR7I enzyme, an isoschizomer ofXho I, into these hybrids. A priori, these breaks can be repaired either by recombination from the homolog or by end-joining. Despite a predicted bias against recovering end-joining products in our system, all cells characterized by enzyme-induced resistance to hypoxanthine/aminopterin/thymidine were, in fact, due to nonhomologous recombination or end-joining. These results are in agreement with other studies that used extrachromosomal sequences to examine the relative efficiencies of end-joining and homologous recombination in mammalian cells, but are in sharp contrast to results of analogous studies in S. cerevisiae, wherein only products of homologous events are detected.Studies using introduced, extrachromosomal DNA substrates have shown that both end-joining (nonhomologous recombination) and homologous recombination occur in mammalian somatic cells; however, the process of endjoining occurs far more frequently than homologous recombination (1). When linear simian virus 40 (SV40) genomes containing terminal homology were examined for the relative rates of nonhomologous end-joining and homologous recombination in mammalian cells, end-joining was about 3-fold more efficient than homologous recombination (2). Similarly, repair oftransfected plasmids occurs primarily by end-joining in mammalian cells (1). Restriction enzyme-induced breaks in mammalian episomal vectors are also repaired by end-joining or by religation of cohesive ends (3). Random integration generally prevails over gene-targeting (4), again suggesting a bias against homologous recombination in mammalian cells.In this report, we present a study ofthe relative efficiencies of end-joining and homologous recombination at a defined in vivo double-strand break in a mammalian chromosome. Since differences exist between extrachromosomal recombination and intrachromosomal recombination (5) as well as between gene-targeting and intrachromosomal recombination (6) in mammalian cells, we sought to determine whether the fate of chromosomal breaks differed from the fate of extrachromosomal breaks.To examine directly the relative efficiencies of end-joining and homologous recombinat...

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Section: Discussionsupporting

confidence: 63%

Spontaneous and restriction enzyme-induced chromosomal recombination in mammalian cells.

Godwin

Bollag

Christie

et al. 1994

Proc. Natl. Acad. Sci. U.S.A.

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“…However, most studies to date have not succeeded in associating interchromosomal crossovers with specific conversion tracts (37,67,71,80,83); reports of crossovers are rare (7,72). Nevertheless, presumptive interchromosomal crossovers, in the form of chromosome-scale loss of heterozygosity (LOH), are commonly observed in mammalian cells.…”

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confidence: 99%

Interchromosomal Crossover in Human Cells Is Associated with Long Gene Conversion Tracts

Neuwirth

Honma

Grosovsky

2007

Molecular and Cellular Biology

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Crossovers have rarely been observed in specific association with interchromosomal gene conversion in mammalian cells. In this investigation two isogenic human B-lymphoblastoid cell lines, TI-112 and TSCER2, were used to select for I-SceI-induced gene conversions that restored function at the selectable thymidine kinase locus. Additionally, a haplotype linkage analysis methodology enabled the rigorous detection of all crossover-associated convertants, whether or not they exhibited loss of heterozygosity. This methodology also permitted characterization of conversion tract length and structure. In TI-112, gene conversion tracts were required to be complex in tract structure and at least 7.0 kb in order to be selectable. The results demonstrated that 85% (39/46) of TI-112 convertants extended more than 11.2 kb and 48% also exhibited a crossover, suggesting a mechanistic link between long tracts and crossover. In contrast, continuous tracts as short as 98 bp are selectable in TSCER2, although selectable gene conversion tracts could include a wide range of lengths. Indeed, only 16% (14/95) of TSCER2 convertants were crossover associated, further suggesting a link between long tracts and crossover. Overall, these results demonstrate that gene conversion tracts can be long in human cells and that crossovers are observable when long tracts are recoverable.In mammalian cells, gene conversion without associated crossover has been widely reported for both intrachromosomal and interchromosomal recombining substrates (13,51,62,80,87). However, most studies to date have not succeeded in associating interchromosomal crossovers with specific conversion tracts (37,67,71,80,83); reports of crossovers are rare (7,72). Nevertheless, presumptive interchromosomal crossovers, in the form of chromosome-scale loss of heterozygosity (LOH), are commonly observed in mammalian cells. LOH is the major contributor to the mutational spectrum in mice and humans (31,33,47,49,78), notably at tumor suppressor loci (3,11,15,25,34,35,40,68,88,89). Crossover products have also been associated with intrachromosomal recombination between repeat sequences in mammalian cells (12,30,52,75), and similar outcomes are observed during the integration of plasmids into host cell chromosomes (9,48,90). Additional evidence for crossovers in mammalian cells is provided by the relatively frequent occurrence of sister chromatid exchange (94), although these products may also result from other mechanisms (56). Since the occurrence of crossovers in mammalian cells has been clearly demonstrated, the absence of crossovers associated with specific interchromosomal gene conversion tracts requires further explanation.Several mechanisms have been proposed to explain the preference for gene conversion without associated crossover in eukaryotic organisms. Crossover is believed to require the formation of Holliday intermediates (17,36,44,76,77) and their resolution through structure-specific endonuclease scissions (79). Theoretically, half of Holliday junctions that are resol...

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“…Radiation directly induces mutations, chromosome aberrations, and recombination (2,6,10,18), reflecting repair or misrepair of DNA damage within or near the locus under study. The delayed effects of radiation, on the other hand, probably reflect dysregulation of genome-stabilizing factors and would therefore be expected to have more global effects.…”

Section: Discussionmentioning

confidence: 99%

“…Ionizing radiation can directly induce mutations (18), chromosome aberrations (10), and homologous recombination (HR) (2,6). In recent years, it has become evident that radiation also induces delayed genomic instability, defined as an increased rate of genetic alterations in the genome of progeny of irradiated cells multiple generations after the initial insult.…”

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confidence: 99%

Ionizing Radiation Induces Delayed Hyperrecombination in Mammalian Cells

Huang

Grim

Smith

et al. 2004

Molecular and Cellular Biology

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Exposure to ionizing radiation can result in delayed effects that can be detected in the progeny of an irradiated cell multiple generations after the initial exposure. These effects are described under the rubric of radiation-induced genomic instability and encompass multiple genotoxic endpoints. We have developed a green fluorescence protein (GFP)-based assay and demonstrated that ionizing radiation induces genomic instability in human RKO-derived cells and in human hamster hybrid GM10115 cells, manifested as increased homologous recombination (HR). Up to 10% of cells cultured after irradiation produce mixed GFP ؉/؊ colonies indicative of delayed HR or, in the case of RKO-derived cells, mutation and deletion. Consistent with prior studies, delayed chromosomal instability correlated with delayed reproductive cell death. In contrast, cells displaying delayed HR showed no evidence of delayed reproductive cell death, and there was no correlation between delayed chromosomal instability and delayed HR, indicating that these forms of genome instability arise by distinct mechanisms. Because delayed hyperrecombination can be induced at doses of ionizing radiation that are not associated with significantly reduced cell viability, these data may have important implications for assessment of radiation risk and understanding the mechanisms of radiation carcinogenesis.

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X rays induce interallelic homologous recombination at the human thymidine kinase gene.

Cited by 38 publications

References 75 publications

Spontaneous and restriction enzyme-induced chromosomal recombination in mammalian cells.

Spontaneous and restriction enzyme-induced chromosomal recombination in mammalian cells.

Interchromosomal Crossover in Human Cells Is Associated with Long Gene Conversion Tracts

Ionizing Radiation Induces Delayed Hyperrecombination in Mammalian Cells

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