Regulation of translation initiation by the elF4E-binding proteins

Most eukaryotic mRNAs possess a 5' cap structure which is important for their translation. The eukaryotic translation initiation factor 4E (eIF4E) binds this structure directly and is repressed by the eIF4E-binbing proteins (4E-BPs), which in turn are regulated through the FRAP/mTOR pathway by a multitude of extracellular stimuli. The focus of this thesis is to investigate the function of the 4E-BPs in the cell by the use of mouse embryonic fibroblasts (MEFs) in which both the genes for 4E-BP1 (Eif4ebp1) and 4E-BP2 (Eif4ebp2) were disrupted. We first examined the role of 4E-BPs in encephalomyocarditis virus (EMCV) infection. 35S-methionine metabolic labeling of MEF proteins revealed that 4E-BPs are not essential for EMCV replication. We then decided to identify mRNAs regulated by 4E-BPs. This was achieved by polyribosomal analysis of specific mRNAs. Our results showed that translation efficiency of c-myc and ornithine decarboxylase (ODC) mRNAs is enhanced in double knockout cells. We also tested MEFs for their sensitivity to rapamycin, a drug known to inhibit FRAP/mTOR. Fluorescence-activated cell sorting (FACS) analysis demonstrated that 4E-BPs contribute to rapamycin-induced growth arrest. Finally, we studied Eif4ebpl-/- ;Eif4ebp2-/- mice in order to test whether they show a phenotype similar to Eif4ebp1 -/- which are known to be smaller and display a lower blood glucose compared with their wild type counterparts. Surprisingly, Eif4ebp1-/-;Eif4ebp2 -/- mice were neither small nor hypoglycemic. The above results emphasize the importance of the 4E-BPs in the cell, thus encouraging further investigations in order to better understand their function