Correction of a nucleotide-excision-repair mutation by human chromosome 19 in hamster-human hybrid cells

Thompson, Larry H.; Mooney, Carolyn L.; Burkhart-Schultz, Karolyn; Carrano, A.V.; Siciliano, Michael J.

doi:10.1007/bf01534738

Cited by 78 publications

(33 citation statements)

References 13 publications

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“…Moreover, we have shown here that other radiation-sensitive strains were not reverted by AC, and the frequency at which they were isolated suggests that a single gene copy is present (44). Preliminary mapping studies locate some of these genes to chromosomes which appear to be associated with structural rearrangements (43). Finally, there is evidence that large parts of the CHO-Kl genome remain diploid with two active alleles, since enzyme polymorphisms persist in these strains (34).…”

Section: Resultsmentioning

confidence: 59%

Azacytidine-induced reactivation of a DNA repair gene in Chinese hamster ovary cells.

Jeggo¹,

Holliday²

1986

Mol. Cell. Biol.

104

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Six X-ray-sensitive (xrs) strains of the CHO-Kl cell line were shown to revert at a very high frequency after treatment with 5-azacytidine. This suggested that there was a methylated xrs+ gene in these strains which was structurally intact, but not expressed. The xrs strains did not complement one another, and the locus was autosomally located. In view of the frequency of their isolation and their somewhat different phenotypes, we propose that the xrs strains are mutants derived from an active wild-type gene. However, there is in addition a methylated silent gene present in the genome. Azacytidine treatment reactivated this gene. We present a model for the functional hemizygosity of mammalian cell lines, which is based on the inactivation of genes by de novo hypermethylation. In contrast to results with xrs strains, other repair-defective lines were found not to be reverted by azacytidine.A large number of mammalian cell variants were isolated after mutagenic treatments, and there is convincing evidence that many of these are structural gene mutations, comparable to those isolated in various microorganisms (for reviews, see references 2, 7, and 36). Although the cells used were either diploids or originally derived from diploids, recessive autosomal mutants were often isolated at frequencies of 10' or 10-4, suggesting that only a single gene copy is present.This has led to Siminovitch's (36) concept of "functional hemizygosity," in which a substantial part of the genome is effectively haploid. One way this could arise is by chromosome rearrangement and the loss of part of the diploid genome (36).Although there is strong evidence for a genetic basis for stable cell variants, there remains the possibility that heritable variation might be caused by epigenetic alterations in gene activity (15). In support of this theory, it has recently been shown that several enzyme-deficient strains of rodent cells are reverted at very high frequency after treatment with 5-azacytidine (AC) (16)(17)(18)27). This is only a weak mutagen, but is known to be highly effective in reducing DNA methylation, probably through the inhibition of a maintenance methylase (23,40). The results make it likely that these enzyme-deficient strains are epigenetic variants, in which transcription has been shut off by methylation. Considerable independent evidence exists that transcription is associated with hypomethylation, and that certain silent genes, for instance those on the inactive X chromosome or in latent retroviruses, are hypermethylated and can be reactivated by AC (for reviews, see references 8, 11, 22, 32, and 33).We previously described the isolation and partial characterization of six CHO strains which are sensitive to X rays (xrs) and defective in the repair of double strand breaks (21, 25). All of these are recessive traits and fall into one complementation group. We show here that all are reverted to X-ray resistance at very high frequency after AC treatment, indicating that the xrs strains have a copy of the xrs gene inactivated thro...

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Section: Resultsmentioning

confidence: 59%

Azacytidine-induced reactivation of a DNA repair gene in Chinese hamster ovary cells.

Jeggo¹,

Holliday²

1986

Mol. Cell. Biol.

104

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“…CHO UV20s were derived from an EMS mutagenesis screen for UV-sensitive mutants and is a member of complementation group 1 for UV sensitivity [38]. The human gene demonstrating cross complementation in these hamster cell lines was termed ERCC1 [39].…”

Section: Resultsmentioning

confidence: 99%

ERCC1/XPF limits L1 retrotransposition

Gasior¹,

Roy-Engel²,

Deininger³

2008

DNA Repair

101

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Retrotransposons are currently active in the human and mouse genomes contributing to novel disease mutations and genomic variation via de novo insertions. However, little is known about the interactions of non-Long Terminal Repeat (non-LTR) retrotransposons with the host DNA repair machinery. Based on the model of retrotransposition for the human and mouse LINE-1 element, one likely intermediate is an extension of cDNA that is heterologous to the genomic target, a flap intermediate. To determine whether a human flap endonuclease could recognize and process this potential intermediate, the genetic requirement for the ERCC1/XPF heterodimer during LINE-1 retrotransposition was characterized. Reduction of XPF in human cells increased retrotransposition whereas complementation of ERCC1-deficiency in hamster cells reduced retrotransposition. These results demonstrate for the first time that DNA repair enzymes act to limit non-LTR retrotransposition and may provide insight into the genetic instability phenotypes of ercc1 and xpf individuals.

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“…The rodentihuman hybrid cells were selected to contain one or a few human chromosome types (14) as indicated in Table 1. These cells were grown as monolayer cultures in minimum essential medium-alpha (MEM-a) containing 15% fetal calf serum (FCS) as described previously (15).…”

Section: Methodsmentioning

confidence: 99%

“…the genetic defects leading to cancer. For example, t(8; 14) This approach allows all human chromosomes, except and t(9;22) are diagnostic of Burkitt's lymphoma and 9-12 and sometimes 14 and 15, to be resolved when the chronic myelogeneous leukemia, respectively. In ad&-analyses are conducted with precision approaching 2%.…”

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confidence: 99%

Centromeric index versus DNA content flow karyotypes of human chromosomes measured by means of slit‐scan flow cytometry

Lucas

Gray

1987

Cytometry

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We have applied slit-scan flow cytometry (SSFCM) to classify human chromosomes according to their centromeric index (CI) and relative DNA content. The resulting bivariate-CI vs. DNA content-distributions shows 14 peaks for normal human chromosomes. Distinct peaks are produced by chromosomes 1, 2, 3, 4+5, 6+7+X, 8, 13+14+15, 16, 17+18, 19+20, and 21+22+Y. In addition, chromosomes 9 through 12 are resolved into three peaks. The identity of the chromosomes comprising each peak was determined by comparing CT vs. DNA content distributions measured for normal human chromosomes by means of SSFCM with CI and DNA content values measured for human chromosomes with image analysis. The accuracy of CI measurement by SSFCM was verified by measuring CIS for human chromosomes isolated from humadrodent hybrid cell lines containing only a few known human chromosomes. These studies showed CIS measured for human chromosomes 1-19 and 21 to be in close agreement with the CIS calculated by means of image analysis. We further confirmed the chromosome assignments for each peak by showing that the relative volumes of the peaks in the CI vs. DNA content distributions for chromosomes from normal cells are similar to the relative frequencies of chromosomes expected for these peaks based on the peak assignments.

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Correction of a nucleotide-excision-repair mutation by human chromosome 19 in hamster-human hybrid cells

Cited by 78 publications

References 13 publications

Azacytidine-induced reactivation of a DNA repair gene in Chinese hamster ovary cells.

Azacytidine-induced reactivation of a DNA repair gene in Chinese hamster ovary cells.

ERCC1/XPF limits L1 retrotransposition

Centromeric index versus DNA content flow karyotypes of human chromosomes measured by means of slit‐scan flow cytometry

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