In vivo interactions of eIF4E in Drosophila cytoplasmic foci. Rolando V. Rivera-Pomar^1,2, Carla Layana^1,2, Paola Ferrero^1,2, Ezequiel Paulucci¹, Pablo Gutierrez¹. 1) Centro Reg Estudios Genomicos, Univ Nacional de La Plata, Florencio Varela, Buenos Aires, Argentina; 2) Departamento de Ciencias Básicas y Experimentales, Universidad Nacional del Noroeste de Buenos Aires. Pergamino, Argentina.

   Eukaryotic translation initiation factor 4E (eIF4E) is required for cap-dependent initiation. In addition, eIF4E occurs in cytoplasmic foci such as processing bodies (PB) and stress granules (SG). We examined the role of key functional amino acid residues of eIF4E in the recruitment of this protein to cytoplasmic foci. We demonstrate that tryptophan residues required for mRNA cap recognition are not required for the recruitment of eIF4E to SG or PB. We show that a tryptophan residue different than the eIF4E-BP interacting domain required for protein-protein interactions is essential for the accumulation of eIF4E in granules. We conclude that protein-protein interactions rather than interactions with the mRNA are essential for the recruitment of eIF4E and for a putative nucleation function. We have also analyzed the interaction of eIF4E with the non-canonical partner Me31B -an ortholog of the helicase rck/p54, Lsm-1 and Cup. We demonstrate by in vivo FRET that the interaction occurs in the cytoplasmic foci between each of the proteins and eIF4E, but not among the interacting proteins. This suggests that eIF4E acts as a simultaneous interactor. With the method used we could not establish id the interactions are simultaneous or they represent subpopulation of interacting molecules in the foci. However, we hypothesize that the interaction may be simultaneous and propose a model for the sequential assembly of the foci in which eIF4E is the common factor for each step. This work has been supported by grants from ANPCyT and CONICET.