Fellowship for Megan Sampley: PDX-1 Regulates Insulin Biosynthesis by Interaction with Transpriction and Translation

The United States spends over 132 billion dollars annually to combat the increasing diabetes crisis, a leading cause of cardiovascular disease and stroke. Type II diabetes is characterized by insulin resistance and improper insulin production and secretion from the pancreatic beta-cell. Detailed studies of molecular mechanisms of insulin production and secretion are essential in developing new clinical treatment strategies to fight diabetes and associated complicating factors. Insulin biosynthesis is regulated at the levels of transcription, translation, and secretion by glucose. Glucose regulation of insulin gene transcription is mainly mediated by the pancreas-specific homeodomain transcription factor PDX-1 Our previous data suggest that PDX-1 stimulates insulin gene transcription in response to glucose by causing changes in histone H4 acetylation levels. On high glucose PDX-1 recruits the histone acetyltransferase p300 to the insulin promoter to induce histone H4 aceylation and insulin gene transcription. On low glucose, PDX-1 associates with histone deacetylases (HDACs), resulting in deacetylation of histone H4 and down-regulation of insulin gene transcription. Treatment with okadaic acid, a PP1 and PP2A protein phosphatase inhibitor, causes a constitutive interaction of PDX-1 with p300, suggesting that the phosphorylation status of PDX-1 is important for its interaction with various factors. Using a yeast-based screen, we have determined that PDX-1 also interacts with the eukaryotic translation initiation factor eIF4E, which is involved in translation initiation in the cytosol and in mRNA export in the nucleus. Based on these findings, we hypothesize that PDX-1 mediates mulitiple protein-protein interactions in the nucleus to coordinate insulin gene transcription and insulin mRNA export in the pancreatic beta-cell in response to glucose. Furthermore, the phosphorylation status of PDX-1 is important in determining its interaction with various proteins. This hypothesis will be tested by the following specific aims: 1. Identification of PDX-1 interacting partners and phosphorylated residues utilizing a proteomics approach; 2.To characterize the interaction of PDX-1 with elF4E and its role in insulin mRNA export.