Mutation of a conserved CDK site converts a metazoan Elongation Factor 1Bbeta subunit into a replacement for yeast eEF1Balpha.
Elongation factor subunit eEF1Bbeta (formerly EF-1beta in plants and EF-1delta in animals) was identified and cloned in a screen for proteins from pea that interact with a cyclin-dependent kinase (CDK). CDKs are enzymes that regulate progression through meiotic and mitotic cell cycles in eukaryotes. eEF1Bbeta and the related protein eEF1Balpha (formerly EF-1beta' in plants and EF-1beta in animals and fungi) can catalyze GTP/GDP exchange on the G-protein eEF1A (formerly EF-1alpha in plants, animals and fungi) during the elongation phase of protein synthesis in eukaryotes. Recombinant Cdc2 and its native homologues from pea extracts associated both in vitro and in vivo with eEF1Bbeta. A Cdc2-cyclin B complex phosphorylated recombinant plant eEF1Bbetas, but not eEF1Balpha. These interactions between CDK and eEF1Bbeta prompted investigations into the in vivo consequences of this relationship. Expression of cDNAs encoding rice or pea eEF1Bbeta subunits failed to complement a Saccharomyces cerevisiae mutant deleted for the eEF1Balpha gene, as was previously observed for the human eEF1Bbeta. However, replacement of Thr91, the sole consensus CDK phosphorylation site in pea eEF1Bbeta, with alanine allowed the pea protein to substitute for eEF1Balpha function in vivo. In addition, this rescued strain was severely cold sensitive, and more sensitive to translational inhibitors than wild-type yeast. Taken together, these results suggest a physiological connection between the cyclin-dependent class of kinases and a translational elongation factor in mitotic cells, and provide the first in vivo evidence that an altered form of eEF1Bbeta can serve as the guanine nucleotide exchange factor for eEF1A.
Pomerening
J R
JR
Department of Molecular Pharmacology, Stanford University School of Medicine, 269 West Campus Drive, CCSR 3160, Stanford, CA 94305-5174, USA. pomereni@stanford.edu
Valente
L
L
Kinzy
T G
TG
Jacobs
T W
TW
D000595:Amino Acid Sequence; D001665:Binding Sites; D015153:Blotting, Western; D017124:Conserved Sequence; D018844:Cyclin-Dependent Kinases / Q000737:chemistry / Q000378:metabolism*; D017931:DNA Primers / Q000737:chemistry; D015723:Gene Library; D020662:Guanine Nucleotide Exchange Factors / Q000378:metabolism; D006153:Guanosine Diphosphate / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism*; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics*; D012275:Oryza / Q000235:genetics / Q000378:metabolism*; D018532:Pisum sativum / Q000235:genetics / Q000378:metabolism*; D020648:Peptide Elongation Factor 1 / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D016133:Polymerase Chain Reaction; D011233:Precipitin Tests; D021122:Protein Subunits; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid
D017931:DNA Primers; D020662:Guanine Nucleotide Exchange Factors; D020648:Peptide Elongation Factor 1; D021122:Protein Subunits; D006153:Guanosine Diphosphate; D006160:Guanosine Triphosphate; D018844:Cyclin-Dependent Kinases