MECHANISMS OF METAL MUTAGENESIS: CR, NI, & BE

一些工业上重要的金属和致癌物。 镍和铬之所以脱颖而出，是因为它们是人类 动物致癌物质，短期毒性试验也呈阳性。 铍，一种动物致癌物和诱变剂，在 短期测试。这些基因的分子机制 三种金属行为尚不为人所知。 这些金属与DNA的相互作用将在一系列的 体外和体内实验旨在测量 金属在诱变中的作用机制和重要性 损伤、聚合酶功能和DNA-蛋白质交联。 金属诱变，将在体外用M13mp2进行检测 真核DNA聚合酶-α正向突变试验。 铍、镍或铬的相互作用在体外产生的突变 将被量化、测序和比较。金属诱导的 突变将与金属诱导的DNA类型进行比较 损害；将特别评估碱基和/或序列。 金属诱导的非特异性DNA损伤也将被识别为 DNA聚合酶暂停位置。因为DNA聚合酶需要 二价金属阳离子的活性，可能会受到不利影响 在错误的阳离子作用下，DNA聚合酶活性的动力学 将在金属离子存在的情况下用动力学方法测定 引子-延伸技术。因为镍和铬都能产生DNA- 蛋白质体外交联剂的研究进展 聚合酶活性和突变的研究将使用 非序列特异性DNA结合蛋白。每种金属的 突变谱也将在体内使用一种 综合穿梭载体。金属诱变作用的比较研究 体内具有损伤和突变谱的体外将提供 机制是否相同的证据。

A number of industrially important metals and carcinogens. Nickel and chromium stand out because they are human and animal carcinogens and also positive in short-term toxicity tests. Beryllium, an animal carcinogen and mutagen, is also positive in short-term tests. The molecular mechanisms by which these three metals act are not known. These metal-DNA interactions will be examined in a series of in vitro and in vivo experiments designed to measure the mechanisms and importance of metals in mutagenesis, DNA damage, polymerase function, and DNA-protein crosslinking. Metal mutagenesis, will be assayed in vitro with the M13mp2 forward-mutation assay using eukaryotic DNA polymerase-alpha. Mutations produced in vitro by the interactions of Be, Ni, or Cr will be quantitated, sequenced, and compared. The metal-induced mutations will be compared with the types of metal-induced DNA damage; base- and/or sequence-specifically will be assessed. Metal-induced nonspecific DNA damage also will be identified as DNA polymerase pause sites. Because DNA polymerase require divalent metal cations for activity and can be adversely affected by the wrong cations, the kinetics of DNA polymerase activity will be determined in the presence of the metal ions using kinetic primer-extension techniques. Since both Ni and Cr produce DNA- protein crosslinking in vitro, the effects of crosslinking on polymerase activity and mutagenesis will be studied using nonsequence-specific DNA binding proteins. Each metal's mutation spectrum will also be analyzed in vivo using an integrated shuttle-vector. Comparison of metal mutagenesis in vivo with the damage and mutation spectrum in vitro will provide evidence of whether the mechanisms are the same or not.