Application of the biotin-dUTP chromosome labelling technique to study the role of 5-bromo-2'-deoxyuridine in the formation of UV-induced sister chromatid exchanges in CHO cells.
The role of 5-bromo-2'-deoxyuridine (BrdU) in the formation of sister chromatid exchanges (SCEs) in cells exposed to UV radiation was studied. Cells were unifilarily labelled (labelling of one strand of chromosomal DNA) with BrdU or biotin-16-2'-deoxyuridine (biotin-dU) and irradiated in G(1) phase of the cell cycle either with 254 nm, which is absorbed by all nucleobases including bromouracil (BrU) or with 313 nm radiation, which is predominantly absorbed by the BrU moiety. Elevated SCE frequencies were observed in cells irradiated at 254 nm (1.2 and 3.0 J m(-2)) which were pre-labelled with BrdU or biotin-dU. Following irradiation at 313 nm (38 and 96 J m(-2)) a statistically elevated SCE frequency was observed in cells pre-labelled with BrdU but not with biotin-dU. In cells pre-labelled with BrdU, UV-radiation at 254 nm was 50-80 times more effective in inducing SCEs than that at 313 nm. This result can be accounted for by the fact that in BrdU-DNA the cross-section for uracilyl radical and bromine atom formation is approximately 100-fold higher at 254 nm than that at 313 nm. Upon irradiation at 254 nm, BrdU had a strong sensitising effect on SCE induction: the SCE frequencies observed in cells pre-labelled with BrdU are approximately 6 times higher than in cells pre-labelled with biotin-dU. From this it follows that BrdU-induced damage is responsible for more than 80% of the SCEs formed in UV irradiated cells unifilarily labelled with BrdU. Based on photochemical considerations and the fact that chemical agents which form DNA interstrand cross-links are among the most potent inducers of SCEs, we propose that an interstrand cross-link may be the major lesion leading to SCEs in BrdU-labelled cells.
Wojcik
Andrzej
A
Institute of Nuclear Chemistry and Technology, 03-195 Warszawa, Poland. guenter.obe@uni-essen.de
von Sonntag
Clemens
C
Obe
Günter
G
D000818:Animals; D001710:Biotin / Q000031:analogs & derivatives*; D001973:Bromodeoxyuridine; D016466:CHO Cells / Q000378:metabolism* / Q000473:pathology / Q000528:radiation effects*; D006224:Cricetinae; D003856:Deoxyuracil Nucleotides; D015224:Dideoxynucleosides / Q000378:metabolism*; D004307:Dose-Response Relationship, Radiation; D008856:Microscopy, Fluorescence / Q000379:methods; D017319:Photosensitizing Agents / Q000378:metabolism; D012854:Sister Chromatid Exchange / Q000528:radiation effects*; D013194:Staining and Labeling / Q000379:methods; D014466:Ultraviolet Rays; D014544:Uridine Triphosphate / Q000031:analogs & derivatives
D001710:Biotin; D001973:Bromodeoxyuridine; D003856:Deoxyuracil Nucleotides; D015224:Dideoxynucleosides; D017319:Photosensitizing Agents; C087624:biotin-16-dUTP; D014544:Uridine Triphosphate; C073006:5-bromo-2',3'-dideoxyuridine