SPONTANEOUS AND GENOTOXICANT INDUCED MUTATION MECHANISMS

描述：自发突变和诱导突变 环境致癌物可导致人类遗传疾病， 癌在本建议书中，申请人将使用E。大肠杆菌继续 来验证DNA序列可以形成 二级结构底物，促进特异性突变 高频率的事件，或者在某些情况下，防止特定的 突变用于诱变的二级结构底物可以包括 由反向重复DNA序列形成的十字形结构; 滑动的错配结构，在DNA复制过程中形成， 直接重复序列;和其它非沃森-克里克双链体， 三链体或四链体结构。DNA可以接受这样的 在人类基因组中广泛存在替代构象。博士 Sinden的实验室已经开发出一种灵敏的体内检测方法， 十字形，并将继续确定之间的关系， 十字形在体内存在及其相关致突变性 潜力反向重复似乎因未对准而被删除 通过形成为十字形或单链DNA的发夹来稳定 在牢房里他的实验室还开发了氯霉素耐药性 回复突变试验，用于测量缺失和重复的频率 在17-23 bp“突变插入”DNA序列的正向重复之间 其可以形成限定的二级结构基底。使用此 系统，他们将继续确定分子中间体， 参与直接重复之间的缺失和复制的机制。 Sinden博士的实验室将继续验证这一假设， 通过三通连接稳定未对准，导致缺失 优先出现在滞后链中。通过合成和 特别设计的准回文素的突变分析，它们 将继续研究导致 将准回文序列校正为完美的反向重复序列， 似乎涉及到一种分子间的链转换， 在DNA复制过程中优先在前导链中。他们将 还研究了参与DNA复制的基因的作用， 在自发DNA定向诱变中， 确定负责引入 基因组中的突变环境诱变剂对 DNA定向突变的频率不仅应该用于 确定促进此类事件的代理，但还应 提供对分子机制的深入了解。

DESCRIPTION: Spontaneous mutation and mutations induced by environmental carcinogens can lead to human genetic disease and cancer. In this proposal, the applicant will use E. coli to continue to test the hypothesis that certain sequences of DNA can form secondary structure substrates, which promote specific mutation events at a high frequency or, in some cases, prevent a specific mutation. Secondary structure substrates for mutagenesis can include cruciform structures, formed from inverted repeated DNA sequences; slipped mis-paired structures, formed during DNA replication in direct repeated sequences; and other non Watson-Crick duplex, triplex, or quadruplex structures. DNA that can adopt such alternative conformations occurs widely in the human genome. Dr. Sinden's laboratory has developed a sensitive in vivo assay for cruciforms and will continue to determine the relationship between the in vivo existence of cruciforms and their relative mutagenic potential. Inverted repeats appear to be deleted by misalignment stabilized by a hairpin formed as a cruciform or in single-stranded DNA in the cell. His lab has also developed a chloramphenicol resistance reversion assay to measure the frequency of deletion and duplication between direct repeats of 17-23 bp "mutation insert" DNA sequences that can form defined secondary structure substrates. Using this system, they will continue to identify the molecular intermediates and mechanisms involved in deletion and duplication between direct repeats. Dr. Sinden's laboratory will continue to test out the hypothesis that misalignment stabilized by a three-way junction leading to deletion occurs preferentially in the lagging strand. By the synthesis and mutational analysis of specifically designed quasipalindromes, they will continue to investigate the molecular mechanism responsible for the correction of quasipalindromes to perfect inverted repeats, which appears to involve an intermolecular strand switch that happens preferentially in the leading strand during DNA replication. They will also investigate the role of genes involved in DNA replication, recombination, and repair in spontaneous DNA directed mutagenesis to identify proteins and enzymes responsible for the introduction of mutations into the genome. The influence of environmental mutagens on the frequency of DNA directed mutation should not only serve to identify agents that facilitate this type of event, but should also provide insight into molecular mechanisms.