BIOCHEMISTRY OF RADIATION-INDUCED DNA STRAND BREAKS

这项研究的目标是了解辐射诱导的DNA 在生物化学方面链断裂，而不是简单地 DNA双螺旋的中断。新的生化和 分子生物学技术，如快速蛋白液 层析，DNA电泳法进行端基分析，以及 逆转录病毒表达载体，将被使用。实验 将决定生产的生化基础， 照射后这些损伤的处理和最终修复 人类细胞。这些信息将增加我们对 细胞辐射效应。它也应该为我们的 对辐射致癌的了解，并可能允许更好地使用 放射治疗在癌症治疗中的重要性。 有待检验的假设是，DNA链断裂是 辐射损伤修复机制的根本重要性 DNA它们不仅是DNA损伤本身，而且还 表示修复其他损害的中间状态，以及 可能对人类系统中的DNA修复起到调节作用。 因此，对链断裂的研究应该提供基本的见解 了解细胞如何维持其遗传稳定性。这项研究 将扩展有关链断裂的知识 体外和细菌研究，通过直接研究这些损伤在 人类细胞。人类系统中最显著的区别是 染色质的额外参与，可以影响到 DNA损伤的诱导和修复。出于这个原因， DNA链断裂和染色质之间的关系将是 研究，特别是关于与染色质相关的 酶聚(ADP-核糖)聚合酶。 研究计划包括三个部分。首先，不同的DNA链 在人类细胞中产生的断裂将被生化鉴定并 量化，并确定修复率。第二， 负责修复这些不同损伤的人体酶(S) 将被提纯和鉴定。第三，有能力 不同的DNA链断裂刺激聚(ADP-核糖化) 将被研究和评估以确定可能的机制 聚腺苷二磷酸核糖聚合酶激活的影响 聚(ADP-核糖)聚合酶活性对链断裂修复的影响 使用表达载体改变细胞内水平进行评估 酶的活性。这项工作应该有助于我们更好地理解 人体系统中辐射作用的分子机制。

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.