title: Repair of intermediate structures produced at DNA interstrand cross-links in Saccharomyces cerevisiae.
abstract: Bifunctional alkylating agents and other drugs which produce DNA interstrand cross-links (ICLs) are among the most effective antitumor agents in clinical use. In contrast to agents which produce bulky adducts on only one strand of the DNA, the cellular mechanisms which act to eliminate DNA ICLs are still poorly understood, although nucleotide excision repair is known to play a crucial role in an early repair step. Using haploid Saccharomyces cerevisiae strains disrupted for genes central to the recombination, nonhomologous end-joining (NHEJ), and mutagenesis pathways, all these activities were found to be involved in the repair of nitrogen mustard (mechlorethamine)- and cisplatin-induced DNA ICLs, but the particular pathway employed is cell cycle dependent. Examination of whole chromosomes from treated cells using contour-clamped homogenous electric field electrophoresis revealed the intermediate in the repair of ICLs in dividing cells, which are mostly in S phase, to be double-strand breaks (DSBs). The origin of these breaks is not clear since they were still efficiently induced in nucleotide excision and base excision repair-deficient, mismatch repair-defective, rad27 and mre11 disruptant strains. In replicating cells, RAD52-dependent recombination and NHEJ both act to repair the DSBs. In contrast, few DSBs were observed in quiescent cells, and recombination therefore seems dispensable for repair. The activity of the Rev3 protein (DNA polymerase zeta) is apparently more important for the processing of intermediates in stationary-phase cells, since rev3 disruptants were more sensitive in this phase than in the exponential growth phase.
authors.affiliation: CRC Drug-DNA Interactions Research Group, Department of Oncology, Royal Free and University College Medical School, University College London, London W1P 8BT, United Kingdom. p.mchugh@ucl.ac.uk
authors.forename: P J
authors.initials: PJ
authors.lastname: McHugh
authors.forename: W R
authors.initials: WR
authors.lastname: Sones
authors.forename: J A
authors.initials: JA
authors.lastname: Hartley
mesh_terms: D018906:Antineoplastic Agents, Alkylating / Q000494:pharmacology*; D002945:Cisplatin / Q000494:pharmacology; D003432:Cross-Linking Reagents / Q000494:pharmacology*; D004249:DNA Damage; D004260:DNA Repair; D004268:DNA-Binding Proteins / Q000235:genetics; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism; D004706:Endodeoxyribonucleases / Q000235:genetics; D005090:Exodeoxyribonucleases; D045585:Flap Endonucleases; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism; D006238:Haploidy; D008466:Mechlorethamine / Q000494:pharmacology; D009154:Mutation; D051721:Rad52 DNA Repair and Recombination Protein; D011995:Recombination, Genetic / Q000187:drug effects*; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins
chemical_list: D018906:Antineoplastic Agents, Alkylating; D003432:Cross-Linking Reagents; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C497197:RAD52 protein, S cerevisiae; D051721:Rad52 DNA Repair and Recombination Protein; D029701:Saccharomyces cerevisiae Proteins; D008466:Mechlorethamine; D004259:DNA-Directed DNA Polymerase; C092602:REV3 protein, S cerevisiae; D004706:Endodeoxyribonucleases; D005090:Exodeoxyribonucleases; D045585:Flap Endonucleases; C094191:MRE11 protein, S cerevisiae; C468323:RAD27 protein, S cerevisiae; D002945:Cisplatin