Iron is an essential nutrient to living systems. As a result, organisms have evolved specific mechanisms for the acquisition and storage of iron for use in growth and development. Ferritin is the major iron storage protein in eukaryotes. The transferrin receptor (TfR) is the major route of iron transport into the animal cells. These proteins play a central role in iron metabolism in animal cells. Synthesis of both ferritin and the TfR is regulated in coordination with the iron balance of the cell. when iron is limiting, synthesis of ferritin is repressed, whereas that of the TfR is elevated. When iron is in excess, ferritin synthesis is elevated while synthesis of the TfR is depressed. Regulation of synthesis of these proteins is mediated by a cytosolic protein called the Iron Responsive Element-Binding Protein (IRE-BP). The IRE-BP binds to a conserved 28 nucleotide element, the IRE, located in the 5' untranslated region of ferritin mRNAs and in the 3' untranslated region of the TfR mRNA. In binding to the IRE, the IRE-BP represses translation of ferritin mRNA while it prevents rapid degradation of the TfR mRNA, resulting in reciprocal but coordinated synthesis of ferritin and TfR. Our long term objectives are to understand the mechanisms of iron regulated gene expression, the molecular basis underlying protein/RNA interaction and the mechanism of translational control via trans-acting repressors. During the next 5 years, we will use yeast as a system for expression of the rabbit IRE-BP to investigate structure/function relationships in the IRE-BP with a particular focus on determining the basis for its specific interaction with the IRE and how this is integrated with iron sensing domains. We will also investigate the mechanism of translational repression by the lRE-BP through the identification of genes in yeast that disrupt (or aid) IRE-BP function. Information gained in these areas will shed light on the molecular basis underlying a variety of nutritional and genetic iron metabolism disorders, including iron deficiency anemia and hemochromatosis. Towards these goals we will: 1) Determine structure/function relationships in the IRE-BP; 2) Investigate the mechanism(s) of regulating IRE-BP activity by iron in vivo, determine the significance of its aconitase activity to iron metabolism; Identify extragenic suppressors of IRE-BP function. These studies will be performed using standard molecular biology, biochemical and yeast genetic techniques. Successful completion of these studies will provide significant insight towards our understanding of nutrient regulated gene expression.
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