| The emergence of genomic techniques and their recent introduction into ecotoxicology provides new opportunities for revealing toxic mechanisms and uncovering novel biomarkers of exposure. The goal of our research is to illustrate the potential of gene expression profiling in ecotoxicology. This project focuses on three well-characterized metal pollutants: copper, cadmium, and zinc, and the invertebrate indicator species, Daphnia magna. Our first objective was to establish that each metal had a discernable gene expression pattern. Using a custom D. magna cDNA microarray containing approximately 5000 cDNA clones, we identified distinct expression profiles in response to sublethal copper, cadmium, and zinc exposures and discovered specific biomarkers of exposure including two probable metallothioneins. The gene expression patterns support known mechanisms of metal toxicity and reveal novel modes of action including zinc inhibition of chitinase activity. Our second objective was to determine how the gene expression profiles change at different effective concentrations. We found that at the lowest concentrations, which do not result in chronic toxicity, few genes were differentially expressed and there was no overlap with the expression profiles of the higher, toxic concentrations, suggesting that expression profiles can distinguish between toxic and non-toxic concentrations of pollutants. Our final goal was to use the gene expression signatures of Cu, Cd, and Zn to predict the presence of metals in blinded field samples collected from copper mines in Eastern California. Water samples containing toxic levels of Cu resulted the differential expression of many of the Cu specific biomarkers discovered in the laboratory exposures and clustering analysis successfully predicted Cu as the primary pollutant responsible for toxicity. Our study suggests that gene expression analysis using cDNA microarrays is capable of producing distinct gene expression profiles for metals, which can be validated in field studies. By integrating gene expression profiling into an environmentally important organism and challenging our microarray to identify the presence of metal contamination in blinded field samples, this study provides experimental support for the utility of Ecotoxicogenomics. |
