Dominant-negative mutant phenotypes and the regulation of translation elongation factor 2 levels in yeast.
The eukaryotic translation elongation factor 2 (eEF2), a member of the G-protein superfamily, catalyzes the post-peptidyl transferase translocation of deacylated tRNA and peptidyl tRNA to the ribosomal E- and P-sites. eEF2 is modified by a unique post-translational modification: the conversion of His699 to diphthamide at the tip of domain IV, the region proposed to mimic the anticodon of tRNA. Structural models indicate a hinge is important for conformational changes in eEF2. Mutations of V488 in the hinge region and H699 in the tip of domain IV produce non-functional mutants that when co-expressed with the wild-type eEF2 result in a dominant-negative growth phenotype in the yeast Saccharomyces cerevisiae. This phenotype is linked to reduced levels of the wild-type protein, as total eEF2 levels are unchanged. Changes in the promoter, 5'-untranslated region (5'-UTR) or 3'-UTR of the EFT2 gene encoding eEF2 do not allow overexpression of the protein, showing that eEF2 levels are tightly regulated. The H699K mutant, however, also alters translation phenotypes. The observed regulation suggests that the cell needs an optimum amount of active eEF2 to grow properly. This provides information about a new mechanism by which translation is efficiently maintained.
Ortiz
Pedro A
PA
Department of Molecular Genetics, Microbiology and Immunology, UMDNJ Robert Wood Johnson Medical School 675 Hoes Lane, Piscataway, NJ 08854-5635, USA.
Kinzy
Terri Goss
TG
D015966:Gene Expression Regulation, Fungal; D009154:Mutation; D010441:Peptide Chain Elongation, Translational; D020652:Peptide Elongation Factor 2 / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D010641:Phenotype; D011401:Promoter Regions, Genetic; D011499:Protein Processing, Post-Translational; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D020506:Untranslated Regions
D020652:Peptide Elongation Factor 2; D029701:Saccharomyces cerevisiae Proteins; D020506:Untranslated Regions