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 double-strand break (DSB) repair and V(D)J recombination. Using a genetic approach we have isolated cell mutants sensitive to ionizing radiation (IR) in the hope of elucidating the mechanism and components required for these pathways. We describe here, an X-ray-sensitive and DSB repair defective Chinese hamster ovary (CHO) cell line, XR-C2, which was assigned to the X-Ray Cross Complementation (XRCC) group 7. This group of mutants is defective in the XRCC7/SCID/Prkdc gene, which encodes the catalytic subunit of DNA-PK (DNA-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, confirming the genetic analyses that these cells harbor a dysfunctional gene for DNA-PKcs. In contrast to other IR group 7 mutants, which contain undetectable 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 suggests that a normal level of kinase activity is critical for radiation resistance but not for V(D)J recombination, whereas the overall structure of the DNA-PKcs protein appears to be of great importance for this process.
Errami
A
A
Department of Radiation Genetics and Chemical Mutagenesis, MGC, Leiden University-Medical Center, Wassenaarseweg 72, 2333 AL Leiden, The Netherlands.
Overkamp
W J
WJ
He
D M
DM
Friedl
A A
AA
Gell
D A
DA
Eckardt-Schupp
F
F
Jackson
S P
SP
Hendrickson
E A
EA
Lohman
P H
PH
Zdzienicka
M Z
MZ
D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D004260:DNA Repair / Q000235:genetics*; D051747:DNA-Activated Protein Kinase; D004268:DNA-Binding Proteins; D004307:Dose-Response Relationship, Radiation; D005816:Genetic Complementation Test; D009153:Mutagens / Q000494:pharmacology; D009154:Mutation; D017346:Protein Serine-Threonine Kinases / Q000235:genetics*; D011836:Radiation Tolerance / Q000235:genetics*; D011995:Recombination, Genetic / Q000235:genetics*; D014965:X-Rays
D004268:DNA-Binding Proteins; D009153:Mutagens; D051747:DNA-Activated Protein Kinase; D017346:Protein Serine-Threonine Kinases