This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Detoxification of non-physiological metals and homeostatic acquisition of nutritional yet toxic metals are fundamental biological processes. Cadmium is a highly toxic environmental contaminant, which causes a number of human disorders, including kidney disease, cancer, and endocrine disruption. Oxidative cellular damage, perturbation of nutritional metal homeostasis, inhibition of DNA repair, and estrogenic activities are implicated with cadmium toxicity. However, the mechanisms of cadmium detoxification in eukaryotes, especially cadmium excretion systems, are largely unknown. The long-term goals of this project are the characterization of molecular mechanisms of cadmium detoxification and employing this knowledge to reduce cadmium exposure to humans. During the search for genes involved in heavy metal resistance in yeast Saccharomyces cerevisiae, we have identified a P-type ATPase. All organisms ranging from bacteria to humans rely on this family of transporters for maintaining a trans-membrane gradient of various ions. Our data strongly suggest that this P-type ATPase is a cadmium selective exporter. Moreover, when cells grow in cadmium-containing media, the expression levels of this transporter are rapidly up regulated through cadmium-mediated inhibition of active turnover. This application focuses on characterization of the function, mechanisms of action and regulation of this cadmium transporter. The central hypothesis is that this P-type ATPase is the first cadmium-specific efflux pump that is unique in structure, substrate specificity and mode of regulation. This hypothesis will be tested using biochemical, cell biological and genetic approaches. First, metal specificity of the P-type ATPase will be elucidated. This study will largely focus on in vivo metal resistant and accumulation assays and in vitro ATPase assays. Second, a multi-disciplinary approach combining yeast genetics, cell biology and chemistry will identify regulatory elements involved in the unique mode of cadmium-dependent post-translational control of this P-type ATPase. The proposed studies will reveal a novel cadmium detoxification mechanism mediated by a P-type ATPase in yeast, a model eukaryote, and ultimately advance our ability to combat cadmium related disorders in humans.
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