title: Repair of cisplatin--DNA adducts by the mammalian excision nuclease.
abstract: Nucleotide excision repair is one of the many cellular defense mechanisms against the toxic effects of cisplatin. An in vitro excision repair assay employing mammalian cell-free extracts was used to determine that the 1,2-d(ApG) intrastrand cross-link, a prevalent cisplatin-DNA adduct, is excised by the excinuclease from a site-specifically modified oligonucleotide 156 base pairs in length. Repair of the minor interstrand d(G)/d(G) cross-link was not detected by using this system. Proteins containing the high mobility group (HMG) domain DNA-binding motif, in particular, rat HMG1 and a murine testis-specific HMG-domain protein, specifically inhibit excision repair of the intrastrand 1,2-d(GpG) and -d(ApG) cross-links. This effect was also exhibited by a single HMG domain from HMG1. Similar inhibition of repair of a site-specific 1,2-d(GpG) intrastrand cross-link by an HMG-domain protein also occurred in a reconstituted system containing highly purified repair factors. These results indicate that HMG-domain proteins can block excision repair of the major cisplatin-DNA adducts and suggest that such an activity could contribute to the unique sensitivity of certain tumors to the drug. The reconstituted excinuclease was more efficient at excising the 1,3-d(GpTpG) intrastrand adduct than either the 1,2-d(GpG) or d(ApG) intrastrand adducts, in agreement with previous experiments using whole cell extracts [Huang, J. -C., Zamble, D. B., Reardon, J. T., Lippard, S. J., Sancar, A. (1994) Proc. Natl. Acad. Sci. U.S.A. 91, 10394-10398]. This result suggests that structural differences among the platinated DNA substrates, and not the presence of unidentified cellular factors, determine the relative excision repair rates of cisplatin-DNA intrastrand cross-links in the whole cell extracts.
authors: Department of Chemistry, Massachusetts Institute of Technology, Cambridge 02139, USA.
authors: D B
authors: DB
authors: Zamble
authors: D
authors: D
authors: Mu
authors: J T
authors: JT
authors: Reardon
authors: A
authors: A
authors: Sancar
authors: S J
authors: SJ
authors: Lippard
mesh_terms: D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002784:Cholesterol / Q000378:metabolism; D002945:Cisplatin; D018736:DNA Adducts; D004260:DNA Repair; D004706:Endodeoxyribonucleases / Q000378:metabolism*; D006367:HeLa Cells; D006609:High Mobility Group Proteins / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D006801:Humans; D007700:Kinetics; D008297:Male; D008322:Mammals; D051379:Mice; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D013379:Substrate Specificity; D013737:Testis
chemical_list: D018736:DNA Adducts; D006609:High Mobility Group Proteins; D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins; C057270:cisplatin-DNA adduct; D002784:Cholesterol; D004706:Endodeoxyribonucleases; D002945:Cisplatin