BIOLOGICALLY SIGNIFICANT DAMAGE IN DNA

人们普遍认为，DNA损伤是细胞死亡的原因 电离辐射。DNA中具有特殊重要性的损伤 在这方面是双链断裂(DSB)。从考虑到 DSB的产生机制据推测，局部相乘 受损部位(LMD)，包括碱基损坏和链损坏 中断，是由类似的机制形成的。这些LMD也可能是 在高效细胞存在的情况下对细胞存活具有生物学意义 DSB修复。对于这些问题的重要性，也可以提出类似的论点 辐射引起的其他细胞效应的损伤-突变和 转型。在此之前，还没有对其结构的研究。 这些类型的损伤要么存在于细胞DNA中，要么存在于模型系统中。这里 我们建议研究由此产生的LMDS和DSB的结构 会在牢房里。一个模型系统，SV40微染色体辐照 在模拟的蜂窝环境中，将用于研究，以及 讨论了这种方法的具体好处。模型系统 已经被表征和验证为DNA的代表 哺乳动物细胞和对辐射参数的良好理解 对这些分子的损害已经达到。《公约》的具体目标 拟议的工作是描述建筑结构的细节 具有生物学意义的LMDS，以期考虑细胞的能力 去修理它们。该项目有三个主要的具体目标和 它们是：1.确定碱基损伤对链的相对产率 LMDS内的中断。2.确定LMD的大小谱 碱基对数目的术语。3.确定平均数量 每个LMD的单个损坏站点。所有从网上获得的信息 这些研究将对考虑细胞对 电离辐射。专门针对这些目的的一系列新的分析方法 在设计时考虑到了这些目标。几种不同的辐射 完成研究的资料来源包括 伽马射线、软X射线和高LET粒子。额外的辅修课程 该系统的主要内容包括：确定系统的数学模型； 双链霉菌生产DSB.DNA-组蛋白的测定 交叉连接；一般碱基损伤分析的发展；和发展 LMDS分析的一般方法。

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.