Dr. Dixon has moved to the University of Arizona.  Project One will continue until March, 2006.

Overview: Both chromium and arsenic are important contaminants at Superfund sites, and in many cases they exist in complex mixtures with PAHs. Exposure by inhalation to chromate compounds has been associated with the development of lung cancer, particularly in cigarette smokers. Exposure to arsenite in drinking water has been associated with the development of skin cancer, as well as internal cancers of the lung, liver, and bladder. Chromate has been shown to be mutagenic in a variety of test systems; in contrast, arsenite and arsenate have generally tested negative for mutagenesis. However, there is evidence that arsenic is clastogenic and it appears to enhance the mutagenic activity of other agents in co-exposures. Previously, project investigators demonstrated glutathione (GSH)-mediated reduction of chromate (CrVI). Furthermore, the mutagenic specificity of chromate is consistent with oxidative DNA damage in yeast, mammalian cells, and the lungs of transgenic mice. The researchers are further characterizing the mutagenic potential and mutagenic specificity of chromate, particularly with respect to the induction of deletion mutations. In addition, the scientists are investigating the mechanisms of metal-induced mutagenesis to include the analysis of the mutagenic potential of arsenic. These metals are being investigated as co-mutagens in combination with PAHs, because environmental exposures often involve complex mixtures of the two classes of carcinogenic compounds. The following two hypotheses are being investigated: 1) Arsenic and chromium function as mutagens by a mechanism involving interaction with intracellular GSH and generation reactive oxygen species, and 2) arsenic and chromium act as co-mutagens by potentiating the mutagenic activity of PAHs. Project investigators are addressing these hypotheses by investigating the mutagenic, and co-mutagenic potential with PAHs, of arsenic and chromium in yeast, mammalian cells, and transgenic mice.

Progress to Date: Recent studies focused on uncovering the mechanism by which arsenic enhances the mutagenic activity of other carcinogenic agents. Project investigators have used DNA-damage signaling as a means of assessing the effects of arsenic on the persistence of DNA damage after UV irradiation. Phosphorylation of the single stranded DNA binding protein (RPA) is a sensitive indicator of DNA damage and arsenite dramatically enhances UV-induced RPA phosphorylation. The magnitude of this effect is equivalent to that observed when nucleotide excision repair is blocked in an XPA cell mutant. Experiments are underway to determine directly if arsenic interferes with removal of DNA damage after UV irradiation.

Proteomics approaches are being used to examine cellular responses to arsenic on a more global level. The patterns of protein expression in arsenic-treated and untreated cells have been compared using 2-D polyacrylamide gel electrophoresis and silver staining. Analysis of gel patterns reveals that there are only a few changes in protein expression in response to arsenite treatment. Further analysis should reveal the identity of these proteins.

Application of Knowledge: The purpose of this project is to provide better understanding of molecular mechanisms by which environmental contaminants such as arsenic and PAHs synergistically cause DNA damage. Such knowledge may be further used for development of chemopreventive treatments and recommendations for minimizing the risk of developing cancer due to arsenic toxicity.

Future Directions: Upcoming experiments will help to determine if arsenic interferes with the removal of DNA damage after UV irradiation, as well as if protein expression patterns are correlated with co-exposures of arsenic and UV or PAHs. Future research will also investigate the effects of nutrient deficiencies on arsenic mutagenicity and carcinogenicity in vivo and in vitro.

Techniques Incorporated:
Cell culture
Transgenic mouse assays
DNA analysis

Keywords:
Chromium
Arsenic
DNA damage
Mixtures
Co-mutagen
Mutations