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J Biol Chem, Vol. 273, Issue 21, 13058-13064, May 22, 1998
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From the We established the radiosensitive cell line SX9
from mammary carcinoma cell line FM3A. In SX9 cells a defect of
DNA-dependent protein kinase (DNA-PK) activity was
suggested. Additionally, a complementation test suggested that the SX9
cell line belongs to a x-ray cross-complementing group (XRCC) 7. Isolation and sequence analyses of DNA-dependent protein
kinase catalytic subunit (dna-pkcs) cDNA in SX9 cells
disclosed nucleotide "T" (9572) to "C" transition causing
substitution of amino acid residue leucine (3191) to proline. Interestingly, the mutation occurs in one allele, and transcripts of
the dna-pkcs expressed exclusively from mutated allele.
V(D)J recombination assay using extrachromosomal vector revealed the defects of not only coding but also signal joint formation. The frequency of the signal joint decreased to approximately one-tenth and
the fidelity drastically decreased to 12.2% as compared with the
normal cell line. To confirm the responsibility of the
dna-pkcs gene for abnormal V(D)J recombination in SX9, the
full-length dna-pkcs gene was introduced into SX9. As a
result, restoration of V(D)J recombination by wild type
dna-pkcs cDNA was observed. SX9 is a novel
dna-pkcs-deficient cell line.
National Institute of Radiological Sciences,
Anagawa 4-9-1, Inage-ku, Chiba-shi, Chiba 263-8555, Japan, the
§ Graduate School of Science and Technology, Chiba
University, Yayoi-cho 1-33, Inage-ku, Chiba-shi, Chiba 263, Japan,
Perkin-Elmer Japan Co., Ltd., Applied Biosystems Division,
Mihama 1-9-2, Urayasu-shi, Chiba 279, Japan, and the ** Faculty of
Biology-Oriented Science and Technology, Kinki University, Uchita-cho,
Naga-gun, Wakayama 649-64, Japan
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