BIOCHEMISTRY OF RADIATION-INDUCED DNA STRAND BREAKS

这项研究的目的是了解辐射诱导的DNA 从生物化学的角度来看， DNA双螺旋的中断。 新的生物化学和 分子生物学技术，如快速蛋白液 色谱法、DNA电泳端基分析，以及 将使用逆转录病毒表达载体。 实验 将决定生产的生化基础， 处理，并最终修复这些病变， 人体细胞 这些信息将增加我们的基本知识， 细胞辐射效应 它也应该有助于我们的 了解辐射致癌作用，并可能允许更好地使用 放射治疗在癌症治疗中的应用。 有待检验的假设是，DNA链断裂是由 对修复受辐射损害的 DNA. 它们不仅是DNA损伤本身， 代表其他损害修复的中间状态，以及 可能对人类系统中的DNA修复具有调节功能。 因此，对链断裂的研究应该提供基本的见解， 细胞如何保持其遗传稳定性。 本研究 将扩大知识链断裂获得通过在 体外和细菌研究，通过直接研究这些病变， 人体细胞 人类系统中最显著的区别是 染色质的参与，这可以影响两个 DNA损伤的诱导和修复。 为此该 DNA链断裂和染色质之间的关系将是 研究，特别是关于染色质相关的 酶聚（ADP-核糖）聚合酶。 研究计划分为三个部分。 首先，不同的DNA链 人类细胞中产生的断裂将被生物化学鉴定， 定，并确定修复率。 二是 负责修复这些不同损伤的人类酶 将被净化和鉴定。 第三，能力 不同的DNA链断裂以刺激聚（ADP-核糖基化） 将进行研究和评估，以确定可能的机制 聚（ADP-核糖）聚合酶激活：以及 聚（ADP-核糖）聚合酶对链断裂修复的活性将 使用表达载体进行评估以改变细胞内水平 的酶。 这项工作应该有助于我们更好地了解 辐射在人体系统中作用的分子机制。

The goal of this research is to understand radiation-induced DNA strand breaks in terms of their biochemistry and not simply as interruptions in the DNA double helix. New biochemical and molecular biological techniques, such as Fast Protein Liquid Chromatography, end-group analysis by DNA electrophoresis, and retroviral expression vectors, will be employed. Experimentation will determine the biochemical basis for the production, processing, and ultimate repair of these lesions in irradiated human cells. This information will add to our basic knowledge of cellular radiation effects. It should also contribute to our understanding of radiation carcinogenesis, and may allow better use of radiotherapy in cancer treatment. The hypothesis, to be tested, is that DNA strand breaks are of fundamental importance to repair mechanisms for radiation-damaged DNA. Not only are they DNA lesions themselves, but they also represent intermediate states in the repair of other damage, and may serve a regulatory function for DNA repair in the human system. The study of strand breaks, therefore, should provide basic insight into how the cell maintains its genetic stability. This research will expand upon knowledge about strand breaks gained through in vitro and bacterial studies, by directly studying these lesions in human cells. The most notable difference in the human system is the added involvement of chromatin, which can affect both the induction and repair of DNA lesions. For this reason, the relationship between DNA strand breaks and chromatin will be studied, particularly with respect to the chromatin-associated enzyme poly(ADP-ribose) polymerase. The research plan has three parts. First, the different DNA strand breaks produced in human cells will be biochemically identified and quantitated, and repair rates will be determined. Second, the human enzyme(s) responsible for repairing these different lesions will be purified and characterized. Third, the ability of different DNA strand breaks to stimulate poly(ADP-ribosylation) will be studied and evaluated to determined the possible mechanism of poly(ADP-ribose) polymerase activation: and the effect of poly(ADP-ribose) polymerase activity on strand-break repair will be assessed using expression vectors to vary intracellular levels of the enzyme. This work should help us to better understand the molecular mechanisms of radiation action in the human system.