McHugh P J, Gill R D, Waters R, Hartley J A
CRC Drug-DNA Interactions Research Group, Department of Oncology, Royal Free and University College Medical School, 91 Riding House Street, London W1P 8BT, UK and.
Nucleic Acids Res. 1999 Aug 15;27(16):3259-66. doi: 10.1093/nar/27.16.3259.
The bifunctional alkylating anticancer drug nitrogen mustard forms a variety of DNA lesions, including monoadducts and intrastrand and interstrand crosslinks. Although it is known that nucleotide excision repair (NER) is important in processing these adducts, the role of the other principal excision repair pathway, base excision repair (BER) is less well defined. Using isogenic Saccharomyces cerevisiae strains disrupted for a variety of NER and BER genes we have examined the relative importance of the two pathways in the repair of nitrogen mustard adducts. As expected, NER defective cells (rad4 and rad14 strains) are extremely sensitive to the drug. One of the BER mutants, a 3-methyladenine glycosylase defective (mag1) strain also shows significant hypersensitivity. Using a rad4/mag1 double mutant it is shown that the two excision repair pathways are epistatic to each other for nitrogen mustard sensitivity. Furthermore, both rad14 and mag1 disruptants show elevated levels of nitrogen mustard-induced forward mutation. Measurements of repair rates of nitrogen mustard N-alkylpurine adducts in the highly transcribed RPB2 gene demonstrate defects in the processing of mono-adducts in rad4, rad14 and mag1 strains. However, there are differences in the kinetics of adduct removal in the NER mutants compared to the mag1 strain. In the mag1 strain significant repair occurs within 1 h with evidence of enhanced repair on the transcribed strand. Adducts however accumulate at later times in this strain. In contrast, in the NER mutants repair is only evident at times greater than 1 h. In a mag1/rad4 double mutant damage accumulates with no evidence of repair. Comparison of the rates of repair in this gene with those in a different genomic region indicate that the contributions of NER and BER to the repair of nitrogen mustard adducts may not be the same genome wide.
双功能烷基化抗癌药物氮芥会形成多种DNA损伤，包括单加合物以及链内和链间交联。虽然已知核苷酸切除修复（NER）在处理这些加合物中很重要，但另一个主要的切除修复途径——碱基切除修复（BER）的作用却不太明确。我们使用了因各种NER和BER基因而被破坏的同基因酿酒酵母菌株，来研究这两种途径在氮芥加合物修复中的相对重要性。正如预期的那样，NER缺陷细胞（rad4和rad14菌株）对该药物极其敏感。其中一个BER突变体，即3-甲基腺嘌呤糖基化酶缺陷（mag1）菌株也表现出显著的超敏反应。使用rad4/mag1双突变体表明，对于氮芥敏感性，这两种切除修复途径相互上位。此外，rad14和mag1破坏菌株都显示出氮芥诱导的正向突变水平升高。对高度转录的RPB2基因中氮芥N-烷基嘌呤加合物修复率的测量表明，rad4、rad14和mag1菌株在单加合物处理方面存在缺陷。然而，与mag1菌株相比，NER突变体中加合物去除的动力学存在差异。在mag1菌株中，显著的修复在1小时内发生，转录链上有增强修复的证据。然而，加合物在该菌株的后期会积累。相比之下，在NER突变体中，修复仅在大于1小时时才明显。在mag1/rad4双突变体中，损伤积累且没有修复的迹象。将该基因的修复率与不同基因组区域的修复率进行比较表明，NER和BER对氮芥加合物修复的贡献在全基因组范围内可能并不相同。
