Differential human nucleotide excision repair of paired and mispaired cisplatin-DNA adducts.
In order to understand the action of the chemotherapeutic drug cisplatin, it is necessary to determine why some types of cisplatin-DNA intrastrand crosslinks are repaired better than others. Using cell extracts and circular duplex DNA, we compared nucleotide excision repair of uniquely placed 1,2-GG, 1,2-AG, and 1,3-GTG cisplatin-crosslinks, and a 2-acetylaminofluorene lesion. The 1,3 crosslink and the acetylaminofluorene lesion were repaired by normal cell extracts approximately 15-20 fold better than the 1,2 crosslinks. No evidence was found for selective shielding of 1,2 cisplatin crosslinks from repair by cellular proteins. Fractionation of cell extracts to remove putative shielding proteins did not improve repair of the 1,2-GG crosslink, and cell extracts did not selectively inhibit access of UvrABC incision nuclease to 1,2-GG crosslinks. The poorer repair of 1,2 crosslinks in comparison to the 1,3 crosslink is more likely a consequence of different structural alterations of the DNA helix. In support of this, a 1,2-GG-cisplatin crosslink was much better repaired when it was opposite one or two non-complementary thymines. Extracts from cells defective in the hMutSalpha mismatch binding activity also showed preferential repair of the 1,3 crosslink over the 1,2 crosslink, and increased repair of the 1,2 adduct when opposite thymines, showing that hMutSalphais not involved in the differential NER of these substrates in vitro. Mismatched cisplatin adducts could arise by translesion DNA synthesis, and improved repair of such adducts could promote cisplatin-induced mutagenesis in some cases.
Moggs
J G
JG
Imperial Cancer Research Fund, Clare Hall Laboratories, South Mimms, Hertfordshire EN6 3LD, UK.
Szymkowski
D E
DE
Yamada
M
M
Karran
P
P
Wood
R D
RD
D000818:Animals; D016466:CHO Cells; D002945:Cisplatin; D006224:Cricetinae; D003432:Cross-Linking Reagents; D018736:DNA Adducts; D004260:DNA Repair; D004706:Endodeoxyribonucleases / Q000378:metabolism; D029968:Escherichia coli Proteins; D006367:HeLa Cells; D006801:Humans; D009711:Nucleotides
D003432:Cross-Linking Reagents; D018736:DNA Adducts; D029968:Escherichia coli Proteins; D009711:Nucleotides; C057270:cisplatin-DNA adduct; D004706:Endodeoxyribonucleases; C036385:endodeoxyribonuclease uvrABC; D002945:Cisplatin