Translational activation of uncapped mRNAs by the central part of human elF4G is 5' end-dependent

Translation initiation factor (elF) 4G represents a critical link between mRNAs and 40S ribosomal subunits during translation initiation. It interacts directly with the cap-binding protein elF4E through its N-terminal part, and binds elF3 and elF4A through the central and C-terminal region. We expressed and purified recombinant variants of human elF4G lacking the N- terminal domain as GST-fusion proteins, and studied their function in cell- free translation reactions. Both elF4G lacking its N-terminal part (aa 486- 1404) and the central part alone (aa 486-935) exert a dominant negative effect on the translation of capped mRNAs. Furthermore, these polypeptides potently stimulate the translation of uncapped mRNAs. Although this stimulation is cap-independent, it is shown to be dependent on the accessibility of the mRNA 5' end. These results reveal two unexpected features of elF4G-mediated translation. First, the C-terminal elF4A binding site is dispensable for activation of uncapped mRNA translation. Second, translation of uncapped mRNA still requires 5' end-dependent ribosome binding. These new findings are incorporated into existing models of mammalian translation initiation.