BIOLOGICALLY SIGNIFICANT DAMAGE IN DNA

一般认为，DNA损伤是细胞死亡的原因， 电离辐射 DNA中的一个损伤， 在这方面是双链断裂（DSB）。 从考虑到 DSB的生产机制，据推测， 损伤位点（LMDS），包括碱基损伤和链损伤 断裂是由类似的机制形成的。 这些LMDS也可以是 在存在有效的抗氧化剂的情况下， DSB修复。 类似的论点可以为这些重要性提出 辐射的其他细胞效应-突变和 转型 以前没有关于其结构的研究。 这些类型的病变无论是在细胞DNA或在模型系统。 这里 我们建议研究LMDS和DSB的结构， 会被关进牢房 一个模型系统，SV 40微型染色体照射 在模拟的细胞环境中，将用于研究， 讨论了这种方法的具体好处。 模型系统 已被表征和验证为代表的DNA中， 哺乳动物细胞和辐射参数的良好理解 已经实现了对这些分子的破坏。 的具体目标 建议的工作是描述结构的细节， 生物学意义的LMDS，以考虑细胞的能力 来修复它们。 该项目有三个主要的具体目标， 分别是：1.确定碱基损伤对链的相对产率 在LMDS中断开。2.确定LMDS的大小范围， 碱基对的数量。3.确定平均数 每个LMDS单独损坏的站点。 所有的信息都是从 这些研究将在考虑细胞对 电离辐射 一系列专门用于这些目的的新测定法 都是基于这些目标而设计的 几种不同的辐射 将采用以下来源完成研究： 伽马射线、软X射线和高LET粒子。 其他次要研究 该系统包括：确认的数学模型， 同步SSB法生产DSB; DNA-组蛋白测定 开发一种通用的碱基损伤分析方法; LMDS检测的一般方法。

It is generally accepted that damage to DNA is the cause of cell death by ionizing radiation. A lesion in the DNA which has particular importance in this respect is the double strand break (DSB). From considerations of the mechanisms of production of DSB it is reasoned that locally multiply damaged sites (LMDS), which include base damages as well as strand breaks, are formed by similar mechanisms. These LMDS could also be biologically significant for cell survival in the presence of efficient DSB repair. Similar arguments can be made for the significance of these lesions for other cellular effects of irradiation - mutation and transformation. There have been no previous studies of the structures of these types of lesion either in cellular DNA or in model systems. Here we propose to study the structures of the LMDS and DSB produced as it would be in a cell. A model system, the SV40 minichromosome irradiated in a simulated cellular environment, will be used for the studies, and the specific benefits of this approach are discussed. The model system has been characterized and validated as representative of the DNA in a mammalian cell and a good understanding of the parameters of radiation damage to these molecules has been attained. The Specific Aims of the proposed work are to describe the details of the structures of the biologically significant LMDS with a view to considering a cell's ability to repair them. There are three major Specific Aims in the project and they are: 1. Determine the relative yields of base damage to strand breaks within the LMDS. 2. Determine the spectrum of sizes of the LMDS in terms of numbers of base pairs. 3. Determine the average number of individually damaged sites per LMDS. All of the information gained from these studies will be significant in considering a cell's response to ionizing radiation. A series of novel assays specific for these purposes has been designed with these goals in mind. Several different radiation sources will be employed for the completion of the studies including gamma-rays, soft X-rays and high LET particles. Additional minor studies with this system include: Confirmation of the mathematical model for the production of DSB by coincident SSB; determination of DNA-histone cross-links; development of a general base damage assay; and, development of general method for assaying LMDS.