Role of DNA-dependent protein kinase in the process of radiation-induced aberration formation.
OBJECTIVE
To further investigate the role of DNA-dependent protein kinase (DNA-PK) and ataxia-telangiectasia mutation (ATM) in the formation of chromosome aberrations, we analysed radiation-induced aberrations in M059J cells, complemented with the PRKDC (DNA-PK) gene by introducing a fragment of human chromosome 8 containing a copy of the human PRKDC gene. One hybrid cell line (M059J Fus1) displayed kinase activity and was radioresistant; the other hybrid cell line (M059J Fus9) showed no kinase activity and was radiosensitive. Both Fus1 and Fus9 cells have only a low ATM activity. Wortmannin, an inhibitor of DNA-PKCS and ATM, was added before irradiation in order to study the effect of DNA-PKCS--and ATM--inhibition on the formation of chromosome aberrations. Furthermore, aberration formation was studied in a lymphoblastoid ATM-deficient cell line AT-1 and in an ATM-proficient control.


METHODS
Confluent cells were irradiated with 200 kV X-rays. Dicentrics, excess acentric fragments, chromatid/ isochromatid breaks and chromatid exchanges were scored in the absence and presence of wortmannin.


RESULTS
In M059J-Fus1 cells and normal lymphoblastoid cells, only chromosome-type aberrations were observed independently of the presence of wortmannin. In DNA-PK-deficient Fus9 cells and in ATM-deficient AT-1 cells, an increasing proportion (30-80%) of cells containing chromatid-type aberrations was observed. This proportion increased with irradiation dose and with wortmannin addition. The aberration yields observed in the complemented M059J-Fus1 cell line were much lower than the corresponding yields observed in the deficient M059J and AT-1 cell lines. However, the low yields observed in the DNA-PK-proficient 'wild-type' cell line M059K were not completely restored.


CONCLUSIONS
Since in M059J-Fus1 cells the radioresistant phenotype with respect to chromosome-type aberration formation was restored by the complementation of PRKDC, ATM expression determines the chromosomal radiosensitivity of M059J cells only to a minor extent. The increasing presence of chromatid-type aberrations in cells irradiated in G0/G1 phase as observed either in DNA-PK- or ATM-deficient cells definitely requires the lack of either kinase. Thus, the aberration spectrum observed is determined by the genetic profile of the respective cells and aberration class amplitudes can be modulated by the inhibition of either kinase.
Virsik-Köpp
P
P
Abteilung für Klinische Strahlenbiologie und Klinische Strahlenphysik, Georg-August-Universität Göttingen, D-37075 Göttingen, Germany. patricia.virsik@medizin.uni-goettingen.de
Rave-Fränk
M
M
Hofman-Hüther
H
H
Schmidberger
H
H
D000730:Androstadienes / Q000494:pharmacology; D064007:Ataxia Telangiectasia Mutated Proteins; D018797:Cell Cycle Proteins; D002460:Cell Line; D045744:Cell Line, Tumor; D002842:Chromatids / Q000648:ultrastructure; D002869:Chromosome Aberrations; D002875:Chromosomes / Q000528:radiation effects / Q000648:ultrastructure; D004247:DNA / Q000528:radiation effects*; D004249:DNA Damage; D051747:DNA-Activated Protein Kinase; D004268:DNA-Binding Proteins; D004307:Dose-Response Relationship, Radiation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D016193:G1 Phase; D005816:Genetic Complementation Test; D006801:Humans; D009687:Nuclear Proteins; D017346:Protein Serine-Threonine Kinases / Q000235:genetics* / Q000502:physiology*; D016192:Resting Phase, Cell Cycle; D025521:Tumor Suppressor Proteins; D000077191:Wortmannin; D014965:X-Rays
D000730:Androstadienes; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D004791:Enzyme Inhibitors; D009687:Nuclear Proteins; D025521:Tumor Suppressor Proteins; D004247:DNA; C576324:ATM protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; D051747:DNA-Activated Protein Kinase; C497263:PRKDC protein, human; D017346:Protein Serine-Threonine Kinases; D000077191:Wortmannin