The overall goal of this project is to investigate the synthesis of the zinc binding protein, metallothionein, as it relates to mammalian zinc metabolism. Emphasis is placed on integrating the induction of the protein to zinc absorption by the small intestine, regulation of the p lasma zinc concentration and intracellular zinc metabolism in liver, kidney, intestine and prostate. Research supported by this project has demonstrated that metallothionein is induced by dietary and parenteral zinc as well as glucocorticoids. This is effected by a change in the translatable metallothionein mRNA pool, most likely as the result of altered metallothionein gene experssion. A partially purified rat liver metallothionein mRNA was used to prepare double stranded cDNA for insertion into a pBR325 plasmid and transfection of an E. coli host. A cDNA containing metallothionein sequencee was isolated. This cloned cDNA, when nick translated to incorporate 32P-dCTP, is used as a probe to quantitate metallothionein mRNA levels in liver kidney, intestine and prostate. The induction of metallothionein mRNA by altered dietary zinc supply, endotoxin and leukocytic endogenous mediator administration, fasting, acute infection, parenteral alimentation and streptozotocin-induced diabetes as well as glucagon and glucocorticoids administration are variables that will be examined. The metallothionein mRNA levels during gestation and lactation will be examined in both the fetus, neonate and dam. The effect of maternal zinc deficiency, alcohol consumption and endotoxin exposure on fetal metallothionein mRNA will be evaluated. For all experiments metallothionein-bound zinc and copper and total metallothionein will be measured as variables and related to mRNA levels. The induction of renal metallothionein synthesis on the renal handling of zinc and copper by the isolated, perfused kidney will be examined. The absorption of zinc by the isolated vascularly perfused intestine will be related to metallothionein synthesis under a variety of conditions that will alter absorption. The relationship of liver zinc metabolism and metallothionein induction to fructose 1,6-bisphosphatase activity will be examined.

Agency
National Institute of Health (NIH)
Institute
National Institute of Arthritis, Diabetes, Digestive and Kidney Diseases (NIADDK)
Type
Research Project (R01)
Project #
5R01AM031127-04
Application #
3152209
Study Section
Metallobiochemistry Study Section (BMT)
Project Start
1982-07-01
Project End
1987-06-30
Budget Start
1985-07-01
Budget End
1986-06-30
Support Year
4
Fiscal Year
1985
Total Cost
Indirect Cost
Name
University of Florida
Department
Type
Earth Sciences/Resources
DUNS #
073130411
City
Gainesville
State
FL
Country
United States
Zip Code
32611
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Cousins, R J; Coppen, D E (1987) Regulation of liver zinc metabolism and metallothionein by cAMP, glucagon and glucocorticoids and suppression of free radicals by zinc. Experientia Suppl 52:545-53
Cousins, R J; Dunn, M A; Leinart, A S et al. (1986) Coordinate regulation of zinc metabolism and metallothionein gene expression in rats. Am J Physiol 251:E688-94
Cowen, L A; Bell, D E; Hoadley, J E et al. (1986) Influence of dietary zinc deficiency and parenteral zinc on rat liver fructose 1,6-bisphosphatase activity. Biochem Biophys Res Commun 134:944-50
Pattison, S E; Cousins, R J (1986) Zinc uptake and metabolism by hepatocytes. Fed Proc 45:2805-9
Cousins, R J; Swerdel, M R (1985) Ceruloplasmin and metallothionein induction by zinc and 13-cis-retinoic acid in rats with adjuvant inflammation. Proc Soc Exp Biol Med 179:168-72
Steel, L; Cousins, R J (1985) Kinetics of zinc absorption by luminally and vascularly perfused rat intestine. Am J Physiol 248:G46-53
Hoadley, J E; Cousins, R J (1985) Effects of dietary zinc depletion and food restriction on intestinal transport of cadmium in the rat. Proc Soc Exp Biol Med 180:296-302
Cousins, R J (1985) Absorption, transport, and hepatic metabolism of copper and zinc: special reference to metallothionein and ceruloplasmin. Physiol Rev 65:238-309