INTRACELLULAR TARGETING AND MESSENGER-RNA INTERACTIONS OF THE EUKARYOTIC TRANSLATION INITIATION-FACTOR EIF4E IN THE YEAST SACCHAROMYCES-CEREVISIAE

The 5' cap structure of eukaryotic mRNAs is believed to play a role in a number of cellular processes, including pre-mRNA splicing, nuclear export and translation. An essential cap-binding protein that is likel y to mediate the participation of the cap in at least one of these pro cesses is the eukaryotic translation initiation factor eIF4E. This pro tein is thought to facilitate the initial ribosomal interaction with t he 5' end of the mRNA, involving the binding of eIF4E to the cap in th e cytoplasm. Yet the subcellular distribution and mechanism of targeti ng of eIF4E has been an unresolved issue. We have therefore examined w hether eIF4E in the yeast Saccharomyces cerevisiae is directed to the nucleus by virtue of a nuclear localization sequence (NLS) in its amin o acid sequence. eIF4E was fused with the 'marker proteins' yeast inve rtase and jellyfish green fluorescent protein. The distribution of the se fusions could be followed using immunofluorescence and confocal mic roscopy of protoplasts and whole cells. These and other fusions were u sed to show that while yeast eIF4E does not possess an efficiently fun ctioning NLS, it can be transported into the nucleus if provided with a known active NLS. However, an NLS-eIF4E fusion of this type cannot b e stably supported by the cell, most likely because of its inhibitory effects when present in large quantities in the nucleus, whereas an NL S fusion with a mutant form of eIF4E that has reduced cap-affinity is tolerated.