BLEOMYCINS CONTROL OF OXIDATIVE DNA DAMAGE

说明(改编自申请)：的作用机制 氧化DNA损伤剂和细胞对这些试剂的反应是 具有相当重要医学意义的问题。然而，所有的 细胞用来防御和修复损伤的机制并不是 为人所知。该项目的长期目标是确定机制 细胞用来保护免受氧化损伤。具体地说，长期- 学期目标是了解DNA修复途径、恢复 过程和蛋白质在细胞氧化反应中的重要作用 破坏DNA的毒剂。三类氧化性DNA损伤剂是 研究包括：电离辐射、抗癌博莱霉素和 结构上相关的金霉素类和过氧化氢。博莱霉素 一组抗生素是一种重要的治疗剂，可用作 单一药物治疗几种人类癌症并广泛应用于 联合化疗和放射治疗。突变的blm5菌株将是 在拟议的工作中用于研究遗传和生化 控制氧化损伤。酵母菌的BLm突变体 酿酒酵母的分离是基于它们对 氧化剂的致死作用。具体目标包括研究 细胞暴露于辐射后DNA损伤的处理和修复 DNA氧化损伤剂和BLM5的作用 基因在维持真核细胞正常DNA功能和修复中的作用 氧化损伤。这些研究将增加对一般知识的新了解 重视DNA修复和新陈代谢以及遗传学领域。 了解氧化后损伤的引入和修复 此外，直到所有主要基因都被发现后，破坏才能完成。 已经确定并确定了涉案人员的特征。将获取数据 来自细胞和分子水平的相互关联的研究，使用 基因特征、分子生物学和生物化学。这些 包括DNA序列分析、研究DNA损伤的方法和 修复，以及透射和扫描电子显微镜。DNA修复 将使用脉冲场电泳法，重组 利用酵母质粒和有丝分裂修复双DNA断裂 重组分析。实验还包括体外诱变，即 零突变的产生和特征，鉴定 双杂交鉴定系统与蛋白质相互作用的研究 利用双杂交系统鉴定相互作用的蛋白质 相互作用的蛋白质，以及从基因组文库中克隆人类同源物。 基因的复杂性、分子的灵活性和可获得性 酿酒酵母的突变菌株允许进行这些研究和技术 接近了。在这个项目中获得的独特知识有 对细胞研究的意义 氧化剂损伤修复机制的确定及临床应用 癌症管理。

Description (Adapted from Application): Mechanisms of action of oxidative DNA-damaging agents and cellular responses to these agents are problems of considerable medical importance. However, all of the mechanisms cells use to defend against and repair the damage are not known. The long-term goal of this project is to determine mechanisms cells use to protect against oxidative damage. Specifically, the long- term objectives are to understand DNA repair pathways, recovery processes, and proteins important in cellular responses to oxidative DNA-damaging agents. Three classes of oxidative DNA-damaging agents are included in the study: ionizing radiation, anti-cancer bleomycins and structurally related phleomycins, and hydrogen peroxide. The bleomycin group of antibiotics is an important therapeutic agent useful as a single agent in treating several human cancers and widely used in combination chemotherapy and radiotherapy. Mutant blm5 strains will be used in the proposed work to investigate the genetic and biochemical controls of oxidative damage. The blm mutants of Saccharomyces cerevisiae were isolated on the basis of their hypersensitivities to lethal effects of oxidants. Specific aims include studies of the processing and repair of DNA lesions produced after exposure of cells to oxidative DNA-damaging agents, and establishing the role of the BLM5 gene in maintaining normal eukaryotic DNA function and repair after oxidative damage. The studies will add new knowledge of general importance to fields of DNA repair and metabolism, and genetics. Understanding the introduction and repair of lesions after oxidative damage, moreover, will not be completed until all the major genes involved have been identified and characterized. Data will be acquired from interrelated studies at the cellular and molecular levels, using genetic characterization, molecular biology, and biochemistry. These include DNA sequence analyses, methodologies for studying DNA damage and repair, and transmission and scanning electron microscopy. DNA repair will be studied using pulsed-field electrophoresis, recombinational repair of double DNA breaks using yeast plasmids, and mitototic recombination assays. Experiments also include in vitro mutagenesis, the production and characterization of null mutation, identifying interactions with proteins using the two-hybrid system of identifying interacting proteins using the two-hybrid system of identifying interacting proteins, and cloning human homologs from genomic libraries. The genetic sophistication, molecular flexibility, and availability of mutant strains of S. cerevisiae permit these studies and technical approaches. The unique knowledge gained in this project has signification implications for cellular studies directed toward determining mechanisms of oxidant injury and repair as well as clinical cancer management.