Genetic complementation between UV-sensitive CHO mutants and xeroderma pigmentosum fibroblasts

Larry H. Thompson, Carolyn L. Mooney, Kerry W. Brookman

Research output: Contribution to journalArticle

27 Citations (Scopus)

Abstract

The purpose of this study was to determine the feasibility of doing complementation analysis between DNA-repair mutants of CHO cells and human fibroblasts based on the recovery of hybrid cells resistant to DNA damage. Two UV-sensitive CHO mutant lines, UV20 and UV41, which belong to different genetic complementation groups, were fused with fibroblasts of xeroderma pigmentosum in various complementation groups. Selection for complementing hybrids was performed using a combination of ouabain to kill the XP cells and mitomycin C to kill the CHO mutants. Because the frequency of viable hybrid clones was generally < 10-6 and the frequency of revertants of each CHO mutant was ∼ 2×10-7, putative hybrids required verification. The hybrid character of clones was established by testing for the presence of human DNA in a dot-blot procedure. Hybrid clones were obtained from 9 of the 10 different crosses involving 5 complementation groups of XP cells. The 4 attempted crosses with 2 other XP groups yielded no hybrid colonies. Thus, a definitive complementation analysis was not possible. Hybrids were evaluated for their UV resistance using a rapid assay that measures differential cytotoxicity (DC). All 9 hybrids were more resistant than the parental mutant CHO and XP cells, indicating that in each case complementation of the CHO repair defect by a human gene had occurred. 3 hybrids were analyzed for their UV-radiation survival curves and shown to be much more resistant that the CHO mutants but less resistant than normal CHO cells. With 2 of these hybrids, sensitive subclones, which had presumably lost the complementing gene, were found to have similar sensitivity to the parental CHO mutants. We conclude that the extremely low frequency of viable hybrids in this system limits the usefulness of the approach. The possibility remains that each of the nonhybridizing XP strains could be altered in the same locus as one of the CHO mutants.

Original languageEnglish (US)
Pages (from-to)423-429
Number of pages7
JournalMutation Research - Fundamental and Molecular Mechanisms of Mutagenesis
Volume150
Issue number1-2
DOIs
StatePublished - 1985
Externally publishedYes

Fingerprint

Xeroderma Pigmentosum
Fibroblasts
CHO Cells
Clone Cells
Hybrid Cells
Mitomycin
Ouabain
DNA Repair
Genes
DNA Damage
Radiation
Survival

ASJC Scopus subject areas

  • Molecular Biology
  • Health, Toxicology and Mutagenesis
  • Medicine(all)

Cite this

Genetic complementation between UV-sensitive CHO mutants and xeroderma pigmentosum fibroblasts. / Thompson, Larry H.; Mooney, Carolyn L.; Brookman, Kerry W.

In: Mutation Research - Fundamental and Molecular Mechanisms of Mutagenesis, Vol. 150, No. 1-2, 1985, p. 423-429.

Research output: Contribution to journalArticle

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