CHARACTERIZATION OF THE IN-VIVO PHOSPHORYLATION SITES OF THE MESSENGER-RNA-CENTER-DOT-CAP-BINDING COMPLEX PROTEINS EUKARYOTIC INITIATION FACTOR-4E AND P20 IN SACCHAROMYCES-CEREVISIAE

Eukaryotic translation is believed to be regulated via the phosphoryla tion of specific eukaryotic initiation factors (eIFs), including one o f the cap-binding complex proteins, eIF-4E. We show that in the yeast Saccharomyces cerevisiae, both eIF-4E and another cap-binding complex protein, p20, are phosphoproteins. The major sites of phosphorylation of yeast eIF-4E are found to be located in the N-terminal region of it s sequence (Ser(2) and Ser(15)) and are thus in a different part of th e protein from the main phosphorylation sites (Ser(53) and Ser(209)) p roposed previously for mammalian eIF-4E. The most likely sites of p20 phosphorylation are at Ser(91) and/or Ser(154). All of the major sites in the two yeast proteins are phosphorylated by casein kinase II in v itro. Casein kinase II phosphorylation of cap complex proteins should therefore be considered as potentially involved in the control of yeas t protein synthesis. Mutagenesis experiments revealed that yeast eIF-4 E activity is not dependent on the presence of Ser(2) or Ser(15). On t he other hand, we observed variations in the amount of (phosphorylated ) p20 associated with the cap binding complex as a function of cell gr owth conditions. Our results suggest that interactions of yeast eIF-4E with other phosphorylatable proteins, such as p20, could play a pivot al role in translational control.