The effect of ethanol on polypeptide chain initiation in reticulocyte lysates. Inhibition of recycling of initiation factor eIF-2.

Using the reticulocyte cell-free system, we have investigated the mechanism by which ethanol inhibits the initiation of protein synthesis. Ethanol inhibited the formation of 40S-initiation complexes, and this effect correlated well with the inhibition by ethanol of overall peptide-chain initiation. Ethanol was a more potent inhibitor of translation at 37 degrees than at 30 degrees. The inhibition of peptide-chain initiation and 40S-initiation complex formation in reticulocyte lysates under other conditions is associated with increased phosphorylation of the alpha-subunit of protein synthesis initiation factor-2 (eIF-2 alpha) and the inhibition of recycling of this factor. Recycling of eIF-2 is mediated by another protein factor GEF (= guanine nucleotide-exchange factor). The addition of ethanol to reticulocyte lysates led to increased phosphorylation of eIF-2 alpha and to a decrease in the rate of exchange of guanine nucleotides bound to eIF-2. This second finding indicated that recycling of eIF-2 was impaired probably due to decreased availability of GEF. Using purified components it was found that ethanol inhibited the ability of GEF to stimulate eIF-2 and that this inhibition showed a similar temperature dependence to the effect of ethanol on overall protein synthesis. Taken together, these results suggest that ethanol leads to inhibition of peptide-chain initiation both through increased phosphorylation of eIF-2 alpha and by directly inhibiting the productive interaction of eIF-2 and GEF.