MECHANISMS OF METAL MUTAGENESIS: CR, NI, & BE

一些工业上重要的金属和致癌物。

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.