Cadmium exposure is an important problem in human toxicology. A major site of toxicity is the kidney where cadmium causes moderate renal failure including an inability to resorb nutrients such as glucose, phosphate, calcium, and amino acids. In mouse kidney cortical cells, which resemble proximal tubule cells, concentrations of cadmium, which do not affect cell viability, ATP levels, or the activity of the (Na+, K+)-ATPase, inhibit (Na+)-dependent glucose and phosphate co-transport. The cadmium ion apparently acts directly on transporter biochemistry not indirectly through disruption of energy conservation mechanisms. Indeed, cadmium ion down regulates (Na+)-glucose (SGLT1) and (Na+)-phosphate co-transporter mRNA. It has also been found that metallothionein (MT), the principal site of binding of cadmium ion, does not reverse inhibition of SGLT1 by cadmium ion even though it is an effective scavenger for protein-bound cadmium ion. To understand the mechanisms of down regulation of (Na+)-nutrient dependent co transporters involved in cadmium ion nephrotoxicity and their relationship to metallothionein induction and activity as a potent metal binding agent, the following specific aims will be pursued: 1) Define the effects of cadmium ion on SGLT1 hnRNA and mRNA. 2) Determine the rate constants for degradation of SGLT1 mRNA from control and cadmium treated cells. 3) Measure the impact of cadmium ion on SGLT1 protein degradation and synthesis. 4) Establish the relationship between cadmium ion-dependent inhibition of SGLT1 activity and reduction in SGLT1 mRNA. 5) Investigate the mechanism of down-regulation of SGLT1 mRNA concentration-inhibition of SGLT1 promoter-driven luciferase expression. 6) Develop an in vivo model that can be used to assess the significance of the in vitro mechanistic studies. 7) Carry out studies on the effects of cadmium ion on (Na+)-phosphate co transport that parallel aims 1-6. 8) Establish the mechanism of induction of MT mRNA synthesis by cadmium ion and zinc ion. 9) Examine the mechanism of down-regulation of SGLT1 mRNA concentration from a chemical perspective. 10) Probe the metal ion exchange competition between MT and other key molecules interacting with cadmium ion.
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