MECHANISM OF RADIATION INDUCED DELAYED GENOTOXICITY

恶性肿瘤细胞的发展需要多种遗传变化，许多环境诱发的癌症在暴露后20多年后才会发病。在癌细胞中发现的这种变化的频率高于通过随机突变可以解释的频率，并且提出了细胞亚群产生突变表型。这种持续升高的遗传不稳定性也是恶性疾病进行性、多阶段发展的主要因素。这种表型，但电离辐射的机制尚未明确。我们在暴露于电离辐射后的酵母中观察到类似的基因组不稳定性超过50个细胞分裂。我们观察到电镀效率持续下降，这些影响不可能是由于初始损伤，因为它们持续了许多代。单个基因的突变导致遗传不稳定性水平升高也不能解释这些影响，因为它们发生在高达70%的暴露细胞中。因此，基因表达的差异更有可能导致高频缺失（HFD）表型。本提案旨在研究这些延迟遗传变化的机制。我们建议从对致癌物的敏感性、顺式作用与反式作用的影响、DNA链断裂水平和氧化应激的参与等方面进一步表征显示HFD表型的克隆的表型。此外，我们将确定在HFD克隆中非法DNA整合是否升高，如果是，我们将定义序列特异性和这种整合事件的目标位点的基因组分布。我们还将开发酵母HFD培养物和对照培养物的完整基因表达谱（6200个基因），以确定可能参与维持或破坏遗传完整性的基因。最后，我们将改变HFD克隆中上调或下调基因的表达，并确定这种改变的基因表达对HFD表型的起始和/或遗传的影响。该项目将描述遗传不稳定性持续升高的现象，深入了解其机制，并可能为逆转表型的干预提供分子靶点。

Multiple genetic changes are required for the development of a malignant tumor cells and many environmentally induced cancers show a delayed onset of more than 20 years following exposure. The frequency of such changes found in cancer cells is higher than can be explained through random mutation and it was proposed that a sub-population of cells develop a mutator phenotype. Such a persistent elevated level of genetic instability is also a major contributor to the progressive, multistage development of malignant disease. This phenotype, ionizing radiation but the mechanism has not been defined. We have observed a similar genomic instability more than 50 cell divisions after exposure to ionizing radiation in the yeast Saccharomyces cerevisiae. We observed a persistently decreased plating efficiency These effects cannot be due to initial damage because of their persistence over many generations. Mutations in a single gene leading to an elevated level of genetic instability also cannot account for these effects because they occur in up to 70% of the exposed cells. It is thus more likely that a difference in gene expression accounts for the high frequency of deletions (HFD) phenotype. This proposal is designed to investigate the mechanism of these delayed inheritable changes. We propose to further characterize the phenotype of clones showing an HFD phenotype in terms of sensitivity to carcinogens, cis- versus trans-acting effects, levels of DNA strand breaks and the involvement of oxidative stress. Furthermore we will determine whether illegitimate DNA integration is elevated in HFD clones and if so, we will define the sequence specificity and the genomic distribution of the target sites of such integration events. We will also develop a complete gene expression profile (6200 genes) for yeast HFD cultures and control cultures to identify genes which may be involved in the maintenance or destabilization of genetic integrity. Finally, we will alter the expression of genes that are up or down regulated in HFD clones, and determine the effect of this altered gene expression on the initiation and/or inheritance of the HFD phenotype. This project should characterize the phenomenon of persistently elevated genetic instability, give insights into its mechanism and might also provide molecular targets for intervention to reverse the phenotype.