MECHANISM OF RADIATION INDUCED DELAYED GENOTOXICITY

恶性肿瘤细胞的发展需要多种基因改变，许多环境诱导的癌症表现出暴露后20多年的延迟发病。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，并提出了一种亚群细胞形成突变子表型的观点。这种持续升高的遗传不稳定水平也是恶性疾病进行性、多阶段发展的主要原因。这种表型、电离辐射的作用机制还没有明确。在暴露于电离辐射的酵母中，我们观察到了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的持续下降，这些影响不是由于最初的损伤，因为它们持续了很多代。导致遗传不稳定性水平升高的单个基因突变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，基因表达的差异更可能是高频率缺失(HFD)表型的原因。这项建议旨在研究这些延迟的可遗传变化的机制。我们建议在对致癌物的敏感性、顺式和反式作用、DNA链断裂水平和氧化应激参与方面进一步表征呈现HFD表型的克隆的表型。此外，我们将确定在HFD克隆中是否存在非法DNA整合，如果是，我们将确定这种整合事件的靶点的序列特异性和基因组分布。我们还将为酵母菌HFD培养和对照培养建立完整的基因表达谱(6200个基因)，以确定可能涉及维持或破坏遗传完整性的基因。最后，我们将改变HFD克隆中上调或下调的基因的表达，并确定这种改变的基因表达对HFD表型启动和/或遗传的影响。这个项目应该描述持续升高的遗传不稳定现象的特征，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.