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New insights on the interaction between the isoforms 1 and 2 of human translation elongation factor 1A

Migliaccio, Nunzia, Ruggiero, Immacolata, Martucci, Nicola M., Sanges, Carmen, Arbucci, Salvatore, Tatè, Rosarita, Rippa, Emilia, Arcari, Paolo, Lamberti, Annalisa
Biochimie 2015 v.118 pp. 1-7
apoptosis, cell membranes, cell viability, cytoplasm, fluorescence, half life, humans, neoplasm cells, neoplasms, phosphorylation, protein synthesis, recombinant fusion proteins
The eukaryotic translation elongation factor 1A (eEF1A) is a moonlighting protein that besides to its canonical role in protein synthesis is also involved in many other cellular processes such as cell survival and apoptosis. In a previous work, we identified eEF1A Raf-mediated phosphorylation sites and defined their role in the regulation of eEF1A half-life and apoptosis of human cancer cells. We proposed that the phosphorylation of eEF1A by C-Raf required the presence of both eEF1A isoforms thus suggesting the formation of a potential eEF1A heterodimer owning regulatory properties. This study aimed at investigating the cellular localization and interaction between two eEF1A isoforms. To this end, we developed chimera proteins by adding at the N-terminal end of both eEF1A1 and eEF1A2 cyan fluorescence protein (mCerulean) and yellow fluorescence protein (mVenus), respectively. The fluorescent eEF1A1 and eEF1A2 chimeras were both addressed to COS-7 cells and found co-localized in the cytoplasm at the level of cellular membranes. We highlighted FRET between the labeled N-termini of eEF1A isoforms. The intra-molecular FRET of this chimera was about 17%. Our results provide novel information on the intracellular distribution and interaction of eEF1A isoforms.