A new X-ray sensitive CHO cell mutant of ionizing radiation group 7, XR-C2, that is defective in DSB repair but has only a mild defect in V(D)J recombination

The DNA-dependent protein kinase (DNA-PK) complex plays a key role in DNA d ouble-strand break (DSB) repair and V(D)J recombination. Using a genetic ap proach we have isolated cell mutants sensitive to ionizing radiation (IR) i n the hope of elucidating the mechanism and components required for these p athways. We describe here, an X-ray-sensitive and DSB repair defective Chin ese hamster ovary (CHO) cell Line, XR-C2, which was assigned to the X-Ray C ross Complementation (XRCC) group 7. This group of mutants is defective in the XRCC7/SCID/Prkde gene, which encodes the catalytic subunit of DNA-PK (D NA-PKcs), Despite the fact that XR-C2 cells expressed normal levels of DNA- PKcs protein, no DNA-PK catalytic activity could be observed in XR-C2, conf irming the genetic analyses that these cells harbor a dysfunctional gene fo r DNA-PKcs, In contrast to other IR group 7 mutants, which contain undetect able or low levels of DNA-PKcs protein and which show a severe defect in V( D)J recombination, XR-C2 cells manifested only a mild defect in both coding and signal junction formation. The unique phenotype of the XR-C2 mutant su ggests that a normal level of kinase activity is critical for radiation res istance but not for V(D)J recombination, whereas the overall structure of t he DNA-PKcs protein appears to be of great importance for this process. (C) 2000 Published by Elsevier Science B.V.