Radioprotection of acute and persistent DNA deletions

项目1的主要目标是找到能够抵消辐射诱导的DNA缺失的试剂，这些缺失导致细胞死亡和遗传不稳定，可能产生长期后果，如出生缺陷和癌症。我们以前已经表明，DNA缺失是诱导电离辐射在酵母和人类细胞，以及在小鼠。我们将在酵母和小鼠中使用报告基因测定来鉴定抵消辐射诱导的DNA缺失频率的试剂。我们已经开发了一种生物发光版本的酵母缺失试验，适用于自动化高通量筛选（HTS）格式。在目的1中，我们提出表征不同化合物文库组的化学品的作用，以确定其在电离辐射处理之前或之后添加时抵消细胞毒性的活性以及DNA缺失的频率（核心D）。酵母菌检测的能力在于它可以用作HTS 在目标3和目标4以及其他研究项目中，确定化学品的优先次序，以便在小鼠和人类细胞中进行进一步的试验。电离辐射也以延迟的方式诱导这些缺失，导致持续的过度重组表型，这是恶性疾病进行性、多阶段发展的主要原因。对于目标2，我们将重新格式化目标1的HTS酵母测定，并使用从目标1中选择的试剂的子集来特异性地对其对抗辐射诱导的过度重组的潜力进行评分。在目标3中，我们将使用目标1中最有效的化学品的子集，以确定它们是否降低小鼠体内辐射诱导的DNA缺失的频率（核心C）。最后，在目标4中，我们将确定来自所有三个项目的三种最有效的辐射防护剂中的任何一种组合是否会降低辐射诱导的小鼠肿瘤的频率。我们还将确定选自目标1的药剂对辐射诱导的氧化性8-氧代鸟嘌呤DNA损伤和链断裂水平的影响。此外，用这些化学物质处理的小鼠的淋巴细胞， 显示体内效应将用于蛋白质组学核心，以确定它们激活的信号通路。

The primary goal of Project 1 is to find agents that counteract radiation-induced DNA deletions leading to cell death and genetic instability that can have long-term consequences such as birth defects and cancer. We have previously shown that DNA deletions are inducible by ionizing radiation in yeast and human cells, as well as in mice. We will use reporter assays in yeast and mice to identify agents that counteract the radiation-induced frequency of DNA deletions. We have developed a bioluminescent version of the yeast deletion assay that is suitable for automation to a high throughput screening (HTS) format. In aim 1 we propose to characterize the effect of the chemicals of different compound library sets for their activity to counteract the cytotoxicity as well as the frequency of DNA deletions when added before or after treatment with ionizing radiation (Core D). The power of the yeast assay is that it can be used as an HTS assay to prioritize chemicals for further assays in mice and in human cells in aims 3 and 4 and in other Research Projects. Ionizing radiation induces these deletions also in a delayed fashion causing a persistent hyperrecombination phenotype, which is a major contributor to the progressive, multistage development of malignant disease. For aim 2 we will reformat the HTS yeast assay of aim 1 and use a subset of agents selected from Aim 1 to specifically score their potential to counteract radiation-induced hyperrecombination. In aim 3 we will use a subset of those chemicals that were most potent in aim 1 to determine whether they reduce the frequency of radiation-induced DNA deletions in vivo in mice (Core C). Finally, in aim 4 we will determine whether any of the three most potent radioprotectors from all three projects combined would reduce the frequency of radiation-induced tumors in mice. We will also determine effects of the agents selected from Aim 1 on levels of radiation-induced oxidative 8-oxoguanine DNA damage and strand breaks. Furthermore, lymphocytes of the mice treated with those chemicals that show an in vivo effect will be used in the Proteomics Core to determine the signaling pathways that they activate.