MECHANISMS OF MUTAGENESIS BY CYCLIC DNA ADDUCTS

这项建议的主要目的是研究 黄曲霉代谢物诱导的环DNA加合物致突变的研究 致癌物质氯乙烯。我们的初步数据表明 亚乙基胞嘧啶(Sigma C)优先结合A残基 在体内导致从C到T的转变。水化的皮损较少 具有突变性，但似乎具有相同的特异性。我们的结果 指出了乙烯加合物的两大类机理 诱变：a)乙烯损伤本质上是非指导性的 活着。与这些病变相反的突变是由于“非 聚合酶的特异性插入，可能由堆积辅助 这些平面病变需要相互作用。包括在此 机理的范畴是烯基衍生物的可能性 在活体内被转化为碱性位点。B)乙烯基衍生物具有 活体内与误编码不同的误编码特性 其他人在体外观察到的特性。为了测试以上内容， 和其他机制，我们提出了以下实验。 1.遗传参数对修复和诱变的影响： 具体地说，RecA、SOS和切除修复的影响。这 信息对于更完整地解释我们的 提供数据。 2.上述成因参数对水合乙烯的响应 损伤。 3.体内和体外定位实验。活体内 实验将通过更精确的方式确认目前的西格玛C数据 方法，专门测试Sigma A和Will的致突变性 检验文献中的预测，即Sigma A可能导致1个BP 删除。体外定位实验的目的是为了 更详细地了解与绕过 西格玛C和西格玛A。 除上述实验外，搜索E.ColiN- 能够作用于乙烯损伤的糖基酶将被安装 在以后的阶段。我们建议开发和使用敏感型 在上述研究过程中进行免疫分析。

The major aim of this proposal is to investigate the mechanisms of mutagenesis by cyclic DNA adducts induced by metabolites of the carcinogen vinyl chloride. Our preliminary data imply that etheno-cytosine (sigma C) preferentially incorporates A residues in vivo leading to C-to-T transitions. Hydrated lesions are less mutagenic, but appear to have the same specificity. Our results point to two major categories of mechanisms for etheno adduct mutagenesis: a) Etheno lesions are essentially non-instructional in vivo. Mutagenesis opposite these lesions arises due to 'non- specific' insertion by polymerase, possibly aided by stacking interactions expected of these planar lesions. Included in this category of mechanisms is the possibility that etheno derivatives are converted to abasic sites in vivo. b) Etheno derivatives have in vivo miscoding properties which are distinct from miscoding properties observed by others in vitro. In order to test the above and other mechanisms, we propose the following experiments. 1. The effect of genetic parameters on repair and mutagenesis: specifically, the effects of RecA, SOS and excision repair. This information is essential for a more complete interpretation of our present data. 2. Response of the above genetic parameters to hydrated etheno lesions. 3. Site-specific in vivo and in vitro experiments. The in vivo experiments will confirm present sigma C data by more exacting approaches, specifically test the mutagenicity of sigma A and will test a prediction in the literature that sigma A may cause 1 bp deletions. The in vitro site-specific experiments are aimed at a more detailed understanding of events associated with bypass of sigma C and Sigma A. In addition to the above experiments, a search for E. coli N- glycosylases capable of acting on etheno lesions will be mounted in later stages. We propose to develop and use sensitive immunoassays during the course of the above studies.