Kauh E A, Bjornsti M A
Department of Biochemistry and Molecular Biology, Thomas Jefferson University, Philadelphia, PA 19107, USA.
Proc Natl Acad Sci U S A. 1995 Jul 3;92(14):6299-303. doi: 10.1073/pnas.92.14.6299.
Camptothecin is a potent antineoplastic agent that interferes with the action of eukaryotic DNA topoisomerase I; the covalent enzyme-DNA intermediate is reversibly stabilized, leading to G2 arrest and cell death. We used a genetic screen to identify cellular factors, other than DNA topoisomerase I, that participate in the process of camptothecin-induced cell death. Following ethyl methanesulfonate mutagenesis of top1 delta yeast cells expressing plasmid-borne wild-type DNA topoisomerase I, six dominant suppressors of camptothecin toxicity were isolated that define a single genetic locus, sct1. Mutant SCT1 cells expressed DNA topoisomerase I protein of similar specific activity and camptothecin sensitivity to that of congenic, drug-sensitive sct1 cells, yet were resistant to camptothecin-mediated lethality. Moreover, camptothecin-treated SCT1 cells did not exhibit the G2-arrested, terminal phenotype characteristic of drug-treated wild-type cells. SCT1 cell sensitivity to other DNA-damaging agents suggests that alterations in SCT1 function suppress camptothecin-induced DNA damage produced in the presence of yeast DNA topoisomerase I.
喜树碱是一种强效抗肿瘤药物，它会干扰真核生物DNA拓扑异构酶I的作用；共价的酶-DNA中间体被可逆性稳定，导致G2期阻滞和细胞死亡。我们利用遗传筛选来鉴定除DNA拓扑异构酶I之外参与喜树碱诱导细胞死亡过程的细胞因子。在用甲磺酸乙酯对表达质粒携带的野生型DNA拓扑异构酶I的top1δ酵母细胞进行诱变后，分离出了六个喜树碱毒性的显性抑制子，它们定义了一个单一的基因座，即sct1。突变的SCT1细胞表达的DNA拓扑异构酶I蛋白具有与同基因的、对药物敏感的sct1细胞相似的比活性和喜树碱敏感性，但对喜树碱介导的致死作用具有抗性。此外，用喜树碱处理的SCT1细胞没有表现出药物处理的野生型细胞所特有的G2期阻滞的终末表型。SCT1细胞对其他DNA损伤剂的敏感性表明，SCT1功能的改变抑制了在酵母DNA拓扑异构酶I存在的情况下喜树碱诱导产生的DNA损伤。
