Frameshift mutation in PRKDC, the gene for DNA-PKcs, in the DNA repair-defective, human, glioma-derived cell line M059J

Radiat Res. 2001 Jul;156(1):2-9. doi: 10.1667/0033-7587(2001)156[0002:fmiptg]2.0.co;2.

Abstract

Anderson, C. W., Dunn, J. J., Freimuth, P. I., Galloway, A. M. and Allalunis-Turner, M. J. Frameshift Mutation in PRKDC, the Gene for DNA-PKcs, in the DNA Repair-Defective, Human, Glioma-Derived Cell Line M059J. Radiat. Res. 156, 2-9 (2001). The glioma-derived cell line M059J is hypersensitive to ionizing radiation, lacks DNA-PK activity, and fails to express protein for the catalytic subunit, DNA-PKcs, while a sister cell line, M059K, derived from the same tumor, has normal DNA-PK activity. Both cell lines are near pentaploid and have multiple copies of chromosome 8, the chromosome on which the DNA-PKcs gene, PRKDC, is located. Sequence analysis of PCR-amplified exons revealed the loss in M059J cells of a single "A" nucleotide in exon 32, corresponding to the first nucleotide of codon 1351 (ACC, Thr) of PRKDC. Loss of the "A" nucleotide would terminate the DNA-PKcs reading frame early in exon 33. DNA from M059K cells had only the wild-type sequence. An analysis of sequences surrounding PRKDC exon 32 from 87 unrelated individuals revealed no polymorphic nucleotides except for a triplet repeat near the 3' end of this exon; no individual had a frameshift mutation in exon 32. No other sequence differences in PRKDC between M059J and M059K cells were observed in approximately 15,000 bp of genomic sequence including the sequences of exons 5 through 38 and surrounding intron sequence, suggesting a possible reduction to homozygosity at this locus prior to acquisition of the mutation leading to the M059J cell line.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Catalytic Domain / genetics
  • Chromosomes, Human, Pair 8 / genetics
  • DNA Mutational Analysis
  • DNA Repair / genetics*
  • DNA-Activated Protein Kinase
  • DNA-Binding Proteins*
  • Exons / genetics
  • Frameshift Mutation / genetics*
  • Glioma / enzymology*
  • Humans
  • Male
  • Molecular Sequence Data
  • Nuclear Proteins
  • Polymerase Chain Reaction
  • Polyploidy
  • Protein Subunits*
  • Protein-Serine-Threonine Kinases / genetics*
  • Radiation Tolerance / genetics
  • Sequence Analysis, DNA
  • Tumor Cells, Cultured

Substances

  • DNA-Binding Proteins
  • Nuclear Proteins
  • Protein Subunits
  • DNA-Activated Protein Kinase
  • PRKDC protein, human
  • Protein-Serine-Threonine Kinases