Evidence that phosphorylation of eIF-2(alpha) prevents the eIF-2B-mediated dissociation of eIF-2 X GDP from the 60 S subunit of complete initiation complexes.
Recent observations have indicated that eukaryotic initiation factor (eIF)-2 and GTP or GDP normally bind to 60 S ribosomal subunits in rabbit reticulocyte lysate and that when eIF-2 alpha is phosphorylated and polypeptide chain initiation is inhibited, eIF-2 X GDP accumulates on 60 S subunits due to impaired dissociation that is normally mediated by the reversing factor (eIF-2B). Current findings now indicate that inhibition due to phosphorylation of eIF-2 alpha is mediated, at least in part, by the inability to dissociate eIF-2 X GDP from the 60 S subunit of complete initiation complexes. At the onset of inhibition, there is an accumulation of Met-tRNA(f) and eIF-2 on the polysomes, despite a marked reduction in Met-tRNA(f) bound to 40 S subunits and Met-peptidyl-tRNA bound to the polysomes. This initial effect is not associated with the formation of "half-mers" (polysomes containing an extra unpaired 40 S subunit), and the 40 S X Met-tRNA(f) complexes, though reduced, still sediment at 43 S. When inhibition is maximal and the polysomes are largely disaggregated, there is an accumulation of 48 S complexes consisting of a 40 S subunit and Met-tRNA(f) bound to globin mRNA as well as small polysomal half-mers, such that residual protein synthesis occurs to about the same degree on "1 1/2"s and "2 1/2"s as on mono-, di-, and triribosomes. Exogenous eIF-2B increases protein synthesis on mono-, di-, and triribosomes and decreases that on half-mers. This is associated with reduced binding of Met-tRNA(f) and eIF-2 to ribosomal particles sedimenting at 80 S and greater and a shift from 48 S to 43 S complexes. These results suggest that eIF-2B must normally promote dissociation of eIF-2 X GDP from the 60 S subunit of complete initiation complexes before they can elongate but cannot when eIF-2 alpha is phosphorylated, resulting in the accumulation of these complexes, some of which dissociate into Met-tRNA(f) X 40 S X mRNA and 60 S X eIF-2 X GDP.
D000818:Animals; D015852:Eukaryotic Initiation Factor-2; D005914:Globins / Q000235:genetics; D006150:Guanine Nucleotides / Q000378:metabolism*; D006153:Guanosine Diphosphate / Q000378:metabolism*; D006418:Heme / Q000378:metabolism; D007700:Kinetics; D046911:Macromolecular Substances; D010448:Peptide Initiation Factors / Q000378:metabolism*; D010766:Phosphorylation; D011132:Polyribosomes / Q000378:metabolism; D011485:Protein Binding; D014176:Protein Biosynthesis; D011506:Proteins / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D012346:RNA, Transfer, Amino Acyl / Q000378:metabolism; D012358:RNA, Transfer, Met; D011817:Rabbits; D012156:Reticulocytes / Q000378:metabolism
D015852:Eukaryotic Initiation Factor-2; D006150:Guanine Nucleotides; D046911:Macromolecular Substances; D010448:Peptide Initiation Factors; D011506:Proteins; D012333:RNA, Messenger; D012346:RNA, Transfer, Amino Acyl; D012358:RNA, Transfer, Met; C024988:tRNA, formylmethionine-; D006153:Guanosine Diphosphate; D006418:Heme; D005914:Globins