A eukaryotic translation initiation factor 2-associated 67 kDa glycoprotein partially reverses protein synthesis inhibition by activated double-stranded RNA-dependent protein kinase in intact cells.
A eukaryotic translation initiation factor 2 (eIF-2)-associated 67 kDa glycoprotein (p67) protects the eIF-2 alpha-subunit from inhibitory phosphorylation by eIF-2 kinases, and this promotes protein synthesis in the presence of active eIF-2 alpha kinases in vitro [Ray, M. K., et al. (1992) Proc. Natl. Acad. Sci. U.S.A. 89, 539-543]. We have now examined the effect of overexpression of this cellular eIF-2 kinase inhibitor in an in vivo system using transiently transfected COS-l cells. In this system, coexpression of genes that inhibit PKR activity restores translation of plasmid-derived mRNA. We now report the following. (1) Transient transfection of COS-1 cells with a p67 expression vector increased p67 synthesis by 20-fold over endogenous levels in the isolated subpopulation of transfected cells. (2) Cotransfection of p67 cDNA increased translation of plasmid-derived mRNAs. (3) Overexpression of p67 reduced phosphorylation of coexpressed eIF-2 alpha. (4) p67 synthesis was inhibited by cotransfection with an eIF-2 alpha mutant S51D, a mutant that mimics phosphorylated eIF-2 alpha, indicating that p67 cannot bypass translational inhibition mediated by phosphorylation of the eIF-2 alpha-subunit. These results show that the cellular protein p67 can reverse PKR-mediated translational inhibition in intact cells.
Wu
S
S
Department of Chemistry, University of Nebraska, Lincoln 68588, USA.
Rehemtulla
A
A
Gupta
N K
NK
Kaufman
R J
RJ
D000626:Aminopeptidases; D004789:Enzyme Activation; D005057:Eukaryotic Cells / Q000378:metabolism; D015852:Eukaryotic Initiation Factor-2 / Q000737:chemistry*; D006023:Glycoproteins / Q000494:pharmacology*; D063208:Methionyl Aminopeptidases; D010766:Phosphorylation / Q000187:drug effects; D014176:Protein Biosynthesis / Q000187:drug effects*; D017346:Protein Serine-Threonine Kinases / Q000494:pharmacology*; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology*; D011994:Recombinant Proteins / Q000494:pharmacology; D019892:eIF-2 Kinase
D015852:Eukaryotic Initiation Factor-2; D006023:Glycoproteins; D011500:Protein Synthesis Inhibitors; D011994:Recombinant Proteins; D017346:Protein Serine-Threonine Kinases; D019892:eIF-2 Kinase; D000626:Aminopeptidases; C081311:METAP2 protein, human; D063208:Methionyl Aminopeptidases