Distinct mechanisms of translation of members of the mammalian elongation factor family, eEF1A-1 and eEF1A-2, during neuronal differentiation

Eukaryotic peptide elongation factor-1 alpha (eEF1A) is an abundantly expressed protein involved in the elongation step of protein synthesis. Its protein expression is regulated at the translational level, as its mRNA shifts from ribonucleoprotein (RNP) complexes to polyribosomes upon growth factor stimulation. The regulation of this shift is mediated by a terminal oligopyrimidine (TOP) element in the 5' untranslated region (UTR), of the eEF1A-1 mRNA. This shifting is also involved during differentiation and mammalian development. During development the expression of eEF1A-1 declines and eEF1A-2, a highly conserved homologue, becomes the dominant eEF1A isoform expressed in brain, heart, and skeletal muscle. In this thesis, three studies were undertaken to examine the regulation and involvement of eEF1A-1 and eEF1A-2 in protein synthesis in neurons. In the first study, the steady-state expression of eEF1A-1 and eEF1A-2 mRNAs and proteins was studied during retinoic acid-induced neuronal differentiation of P19 embryonic teratocarcinoma cells. The decline of eEF1A-1 and the increase in eEF1A-2 expression during neuronal differentiation were found to be regulated primarily at the transcriptional level, since there was a correlation between the levels of mRNA and protein expression. In differentiated P19 neurons translational repression of eEF1A-1 mRNA also contributed to the decrease of eEF1A-1 protein. This indicated that transcriptional and translational repression of eEF1A-1 mRNA occurred during neuronal differentiation. Unlike eEF1A-1, eEF1A-2 mRNA was associated primarily with polyribosomes, suggesting that eEFIA-2 is preferentially translated during P19 neuronal differentiation. In the second study, the eEF1A-1 isoform was shown to be preferentially synthesized in nerve growth factor (NGF) stimulated PC12 cells. The increased synthesis of eEF1A-1 was mediated at the translational level, as shown by the shifting of eEF1A-1 mRNA to polyribosomes after NGF stimul