S-adenosylmethionine decarboxylase (AdoMetDC) is a key regulated enzyme in the pathway of synthesis of the polyamines, spermidine and spermine. The polyamines are necessary for cell growth; deregulation of polyamine biosynthesis can lead to neoplastic transformation of normal cells. Expression of the AdoMetDC gene is controlled at the translational level by exogenous regulatory signals from growth factors and hormones in certain cell types, as well as by the endogenous concentrations of the polyamine end products of the pathway in all cells. Translational control is mediated by an upstream open reading frame (uORF) located in the 5' leader, 14 nucleotides from the cap. The goal of this project is to define the mechanism of translational control by the AdoMetDC uORF by testing the following model. We hypothesize that the nascent peptide product from the uORF induces a blockage to translation of the AdoMetDC mRNA by interacting with a target in the translational apparatus. Normal intracellular levels of spermidine and/or spermine are required for this interactions and, therefore, reduced polyamine levels alleviate translational suppression and stimulate synthesis of AdoMetDC protein. In addition, because of the proximity of the initiation codon of the uORF to the 5' end of the mRNA, initiation at the uORF, and therefore suppression of downstream translation, seems to be sensitive to, and therefore possibly regulated by, the intracellular activity of eIF-4E. To test this model, we will: 1.) Define the mechanisms of inhibition of translation by the peptide encoded by the uORF, including identification of its target, 2.) test the involvement of eIF-4E in the cell type-specific regulation of AdoMetDC translation, and 3.) test the hypothesis that polyamines modulate the interaction of the peptide product of the uORF with its target. These studies will not only define the mechanism of regulation of a key step in polyamine biosynthesis, but will also extend our fundamental knowledge of the role uORFs in translational control generally.
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