PURIFICATION AND CHARACTERIZATION OF EUKARYOTIC POLYPEPTIDE-CHAIN INITIATION-FACTOR 4F FROM DROSOPHILA-MELANOGASTER EMBRYOS

The eukaryotic polypeptide chain initiation factor 4F (eIF-4F), purifi ed by m(7)GTP-Sepharose chromatography from whole extracts of Drosophi la melanogaster embryos, consists of two subunits, the previously iden tified eIF-4E (35 kDa) (Maroto, F. G., and Sierra, J. M. (1989) Mol. C ell. Biol. 9, 2181-2190) and another of 200 kDa. Both subunits cosedim ented through a sucrose density gradient containing 0.5 M KCl. In cont rast to rabbit reticulocyte eIF-4F, we did not find any RNA-dependent ATPase associated with the Drosophila factor. As shown previously for eIF-4E, the p200 subunit was also required for the translation of endo genous mRNAs in cell-free systems from Drosophila embryos. Only the eI F-4E subunit was able to cross-link to the m(7)G cap structure. Howeve r, an efficient cross-linking of the p200 subunit to an uncapped mRNA was observed. Both subunits were phosphorylated in vitro by protein ki nase C from rat brain. As an extension of our previous results (Zapata , J. M., Maroto, F. G., and Sierra, J. M. (1991) J. Biol. Chem. 266, 1 6007-16014) we found that the translation of the heat shock mRNAs was independent of both of the eIF-4F subunits.