Reduced X-ray Resistance and Homologous Recombination Frequencies in a RAD54-/- Mutant of the Chicken DT40 Cell Line

Cell. 1997 Apr 18;89(2):185-93. doi: 10.1016/s0092-8674(00)80198-1.

Abstract

rad54 mutants of the yeast Saccharomyces cerevisiae are extremely X-ray sensitive and have decreased mitotic recombination frequencies because of a defect in double-strand break repair. A RAD54 homolog was disrupted in the chicken B cell line DT40, which undergoes immunoglobulin gene conversion and exhibits unusually high ratios of targeted to random integration after DNA transfection. Homozygous RAD54-/- mutant clones were highly X-ray sensitive compared to wildtype cells. The rate of immunoglobulin gene conversion was 6- to 8-fold reduced, and the frequency of targeted integration was at least two orders of magnitude decreased in the mutant clones. Reexpression of the RAD54 cDNA restored radiation resistance and targeted integration activity. The reported phenotype provides the first genetic evidence of a link between double-strand break repair and homologous recombination in vertebrate cells.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alkylating Agents / pharmacology
  • Amino Acid Sequence
  • Animals
  • B-Lymphocytes / drug effects
  • B-Lymphocytes / physiology*
  • B-Lymphocytes / radiation effects
  • Base Sequence
  • Cell Line
  • Chickens
  • Cloning, Molecular
  • DNA Helicases
  • DNA Repair Enzymes
  • DNA, Complementary / genetics
  • DNA, Recombinant
  • Fungal Proteins / genetics
  • Fungal Proteins / physiology*
  • Gamma Rays
  • Gene Conversion
  • Gene Targeting
  • Genes, Immunoglobulin / genetics*
  • Immunoglobulin Light Chains / genetics
  • Immunoglobulin M / genetics
  • Methyl Methanesulfonate / pharmacology
  • Molecular Sequence Data
  • Mutation
  • Radiation Tolerance*
  • Recombination, Genetic / genetics*
  • Saccharomyces cerevisiae / genetics
  • Saccharomyces cerevisiae Proteins*
  • Sequence Homology, Amino Acid

Substances

  • Alkylating Agents
  • DNA, Complementary
  • DNA, Recombinant
  • Fungal Proteins
  • Immunoglobulin Light Chains
  • Immunoglobulin M
  • Saccharomyces cerevisiae Proteins
  • Methyl Methanesulfonate
  • RAD54 protein, S cerevisiae
  • DNA Helicases
  • DNA Repair Enzymes

Associated data

  • GENBANK/U92461