Characterization of the in vivo phosphorylation sites of the mRNA.cap-binding complex proteins eukaryotic initiation factor-4E and p20 in Saccharomyces cerevisiae.
Eukaryotic translation is believed to be regulated via the phosphorylation of specific eukaryotic initiation factors (eIFs), including one of 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 its sequence (Ser2 and Ser15) and are thus in a different part of the protein from the main phosphorylation sites (Ser53 and Ser209) proposed previously for mammalian eIF-4E. The most likely sites of p20 phosphorylation are at Ser91 and/or Ser154. All of the major sites in the two yeast proteins are phosphorylated by casein kinase II in vitro. Casein kinase II phosphorylation of cap-complex proteins should therefore be considered as potentially involved in the control of yeast protein synthesis. Mutagenesis experiments revealed that yeast eIF-4E activity is not dependent on the presence of Ser2 or Ser15. On the other hand, we observed variations in the amount of (phosphorylated) p20 associated with the cap-binding complex as a function of cell growth conditions. Our results suggest that interactions of yeast eIF-4E with other phosphorylatable proteins, such as p20, could play a pivotal role in translational control.
Zanchin
N I
NI
Department of Gene Expression, National Biotechnology Research Centre (GBF), Braunschweig, Federal Republic of Germany.
McCarthy
J E
JE
D000595:Amino Acid Sequence; D047390:Casein Kinase II; D003001:Cloning, Molecular; D039561:Eukaryotic Initiation Factor-4E; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D039502:Nuclear Cap-Binding Protein Complex; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification; D010448:Peptide Initiation Factors / Q000737:chemistry / Q000378:metabolism*; D010748:Phosphopeptides / Q000737:chemistry / Q000302:isolation & purification; D010750:Phosphoproteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D012315:RNA Caps / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D012694:Serine; D013379:Substrate Specificity
C100383:CBC2 protein, S cerevisiae; D039561:Eukaryotic Initiation Factor-4E; D039502:Nuclear Cap-Binding Protein Complex; D010446:Peptide Fragments; D010448:Peptide Initiation Factors; D010748:Phosphopeptides; D010750:Phosphoproteins; D012315:RNA Caps; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D012694:Serine; D047390:Casein Kinase II; D017346:Protein Serine-Threonine Kinases