Hormonal and Nutritional Regulation of Protein Synthesis in Eukaryotic Cells

The overall goal of the research program is to understand the regulation of the initiation phase of protein synthesis in eukaryotic cells. Of the many steps in initiation, only two have been shown to be subject to regulation in vivo. These steps are the binding of initiator methionyl-tRNAi to the 40S ribosomal subunit and the binding of mRNA to the 43S preinitiation complex. The former step is mediated by a protein termed eukaryotic initiation factor (eIF)-2 and is regulated by changes in the activity of a guanine nucleotide exchange protein, eIF2B, that catalyzes GDP exchange on eIF2. The binding of mRNA to the 43S preinitiation complex involves the interaction among a group of proteins collectively referred to as eIF4F and the binding of the eIF4F complex to the 5'-end of mRNA.

Our previous studies have shown that in rat liver and skeletal muscle, the initiation of protein synthesis is responsive to a variety of stimuli including insulin, essential amino acids, and calcium-mobilizing hormones. In each of these cases, initiation is regulated through changes in the interaction of specific initiation factors with each other and/or with various regulatory proteins. The binding of the initiation factors to each other and to the regulatory proteins is controlled through changes in phosphorylation state. For example, eIF4E is an mRNA cap binding protein important in controlling not only the overall rate of protein synthesis, but also in selecting which mRNAs will be translated into protein. The ability of eIF4E to bind to the mRNA cap structure is regulated by its interaction with a second initiation factor, eIF4G, and the translational regulator, 4E-BP1. In skeletal muscle of diabetic rats, the amount of eIF4E present in the active eIF4F complex is decreased relative to controls whereas the amount in the inactive 4E-BP1·eIF4E complex is increased. The binding of eIF4E to these other proteins is correlated with changes in phosphorylation of each of the proteins. Current efforts are directed at delineating the mechanism of interaction of eIF4E with 4E-BP1 and eIF4G as well as the interaction between eIF2 and eIF2B using biochemical and molecular biological approaches. In addition, the effect of phosphorylation on the activity and interaction of the proteins is being investigated. Finally, identification and characterization of the kinases and phosphatases involved in mediating the changes in phosphorylation state is in progress.