Casein kinase II phosphorylates translation initiation factor 5 (eIF5) in Saccharomyces cerevisiae.
Eukaryotic translation initiation factor 5 (eIF5) interacts with the 40S initiation complex (40S-eIF3-mRNA-Met-tRNA(f)-eIF2-GTP) to promote the hydrolysis of ribosome-bound GTP. In Saccharomyces cerevisiae, eIF5 is encoded by a single-copy essential gene, TIF5, that is required for cell growth and viability. In this work, we show that eIF5 immunoprecipitated from cell-free extracts of (32)P-labelled yeast cells is phosphorylated on multiple serine residues. Phosphopeptide mapping reveals four major sites of phosphorylation that appear to be identical to recombinant yeast eIF5 sites phosphorylated in vitro by casein kinase II. Furthermore, analysis of eIF5 isolated from a yeast strain having a conditional mutant of casein kinase II indicates that phosphorylation of eIF5 is completely abolished at the non-permissive temperature. Additionally, haploid yeast strains were constructed to contain Ser-to-Ala mutations at the five casein kinase II consensus sequences in eIF5; in these cells, eIF5 phosphorylation was absent. Surprisingly, substitution of the TIF5 gene mutated at these sites for the wild-type gene had no obvious effect on cell growth under normal growth conditions. The implications of these results in eIF5 function are discussed.
Maiti
Tapan
T
Department of Developmental and Molecular Biology, Albert Einstein College of Medicine of Yeshiva University, Bronx, New York 10461, USA.
Bandyopadhyay
Amitabha
A
Maitra
Umadas
U
D000595:Amino Acid Sequence; D047390:Casein Kinase II; D016384:Consensus Sequence; D004926:Escherichia coli / Q000235:genetics; D039642:Eukaryotic Initiation Factors / Q000378:metabolism*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010449:Peptide Mapping; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000254:growth & development / Q000378:metabolism; D016415:Sequence Alignment; D012694:Serine / Q000378:metabolism
D039642:Eukaryotic Initiation Factors; D011994:Recombinant Proteins; C451459:eukaryotic initiation factor-5B; D012694:Serine; D047390:Casein Kinase II; D017346:Protein Serine-Threonine Kinases