Tupl and Ssn6 transcriptionally repress a wide variety of genes in yeast but do not appear to bind DNA. We provide genetic and biochemical evidence that the DNA-binding protein a2, a regulator of cell-type-specific genes, recruits the Tupl/Ssn6 repressor by directly interacting with Tupl. This interaction is mediated by a region of Tupl containing seven copies of the WD repeat, a 40 amino acid motif of unknown function found in many other proteins. We have found that a single WD repeat will interact with a2, indicating that the WD repeat is a protein-protein interaction domain. Furthermore, a fragment of Tupl containing primarily WD repeats provides at least partial repression in the absence of Ssn6, suggesting that the repeats also mediate interaction between Tupl and other components of the repression machinery.

Toxicogenomics has provided innovative approaches to chemical screening, risk assessment, and predictive toxicology. If applied to ecotoxicology, genomics tools could greatly enhance the ability to understand the modes of toxicity in environmentally relevant organisms. Daphnia magna, a small aquatic crustacean, is considered a "keystone" species in ecological food webs and is an indicator species for toxicant exposure. Our objective was to demonstrate the potential utility of gene expression profiling in ecotoxicology by identifying novel biomarkers and uncovering potential modes of action in D. magna. Using a custom D. magna cDNA microarray, we identified distinct expression profiles in response to sublethal copper, cadmium, and zinc exposures and discovered specific biomarkers of exposure including two probable metallothioneins, and a ferritin mRNA with a functional IRE. The gene expression patterns support known mechanisms of metal toxicity and reveal novel modes of action including zinc inhibition of chitinase activity. By integrating gene expression profiling into an environmentally important organism, this study provides experimental support for the utility of ecotoxicogenomics.