Mechanistic insights into cellular metal detoxification

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DESCRIPTION (provided by applicant): Detoxification of non-physiological metals and homeostatic acquisition of nutritional yet toxic metals are fundamental biological processes. A number of health issues linked to heavy metal toxicity underscore the physiological significance of metal metabolism. For example, cadmium is a highly toxic environmental contaminant and implicated in disorders, including kidney failure, cancer, reproductive defects, and endocrine disruption. While exposure to cadmium is unavoidable and widespread, the cellular mechanisms of cadmium metabolism, 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 intake. During the search for genes involved in metal resistance in yeast Saccharomyces cerevisiae, a model eukaryote, the PI identified a novel cadmium extruding P-type ATPase that is non- functional in cadmium sensitive yeast strains. Virtually all organisms rely on this family of transporters for maintaining a transmembrane gradient of various ions, which is vital for nutrient uptake, neurotransmission, signaling, and/or prevention of toxic accumulation of ions. Mutations in copper transporting P-type ATPases lead to lethal genetic diseases in humans, which highlights the essential role for P-type ATPases in metal metabolism. However, mechanistic details of metal-transporting P-type ATPases remain to be elucidated. This application focuses on the characterization of the function, mechanisms of action, and regulation of a cadmium transporting P-type ATPase. 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 a multi-disciplinary approach. First, cadmium specificity of the P-type ATPase will be elucidated, and structural determinants of the specificity will be identified. This study will largely focus on ATPase assays and structure-function analysis of metal-binding domains and residues. Second, yeast genetics, cell biology, and biophysical approaches will identify trans-acting regulatory factors and cis-acting elements involved in the unique mode of cadmium-dependent expression control of this cadmium efflux pump. The proposed studies are expected to reveal a novel cadmium detoxification system in a eukaryote, shed light on the mechanism of P-type ATPase-mediated metal transport, and ultimately advance the ability to combat metal-related disorders in humans. PUBLIC HEALTH RELEVANCE: Cadmium is a highly toxic metal that is implicated in kidney disease, cancer, reproductive defects, and endocrine disruption. Given that cadmium exposure to humans is unavoidable and widespread, mechanistic insights into cellular cadmium absorption, sequestration, and extrusion would facilitate prevention and treatment of cadmium-related disorders and help scientists develop methods for the reduction of cadmium intake in humans. This application addresses this problem through the identification and characterization of a cellular cadmium efflux mechanism that plays a critical role in preventing excess accumulation of cadmium.