Eukaryotic Initiation Factors 4A (eIF4A) and 4G (eIF4G) Mutually Interact in a 1:1 Ratio in Vivo

Abstract

mRNA translation in eukaryotic cells involves a set of proteins termed translation initiation factors (eIFs), several of which are involved in the binding of ribosomes to mRNA. These include eIF4G, a modular scaffolding protein, and eIF4A, an RNA helicase, of which two closely related forms are known in mammals, eIF4AI and eIF4AII. In mammals, eIF4G possesses two independent sites for binding eIF4A, whereas in other eukaryotes (e.g. yeast) only one site appears to be present, thus raising the issue of the stoichiometry of eIF4G·eIF4A complexes in different eukaryotes. We show that in human embryonic kidney cells eIF4G is associated with eIF4AI or eIF4AII but not with both simultaneously, suggesting a stoichiometry of 1:1 rather than 1:2. To confirm this, eIF4AI or eIF4AII was expressed in a tagged form in these cells, and complexes with eIF4G were again isolated. Complexes containing tagged eIF4AI or eIF4AII contained no endogenous eIF4A, supporting the notion that eIF4G binds only one molecule of eIF4A. Each binding site in eIF4G can bind either eIF4AI or eIF4AII. The data imply that the second binding site in mammalian eIF4A does not bind an additional eIF4A molecule and that initiation factor complexes in different eukaryotes contain one eIF4A per eIF4G.