Recycling and phosphorylation of eukaryotic initiation factor 2 on 60S subunits of 80S initiation complexes and polysomes.
Phosphorylation of the alpha-subunit (38 kDa) of eukaryotic initiation factor 2 (eIF-2 alpha) regulates initiation of protein synthesis in eukaryotic cells. This phosphorylation is enhanced in cycloheximide-treated heme-deficient reticulocyte lysates in which polysomes are maintained. In early heme deficiency prior to polysome disaggregation, eIF-2(alpha P) accumulates primarily on the 60S subunits of polysomes. Further, isolated polysomes contain eIF-2 alpha that is efficiently phosphorylated in vitro by heme-regulated inhibitor (HRI). Immunoblot analysis of eIF-2 distribution in sucrose gradients of actively protein-synthesizing lysates indicates that eIF-2 is distributed at low levels throughout the polysome profiles. These findings suggest that polysome-bound eIF-2 alpha is a target of HRI under physiological conditions. The presence of eIF-2 on the 60S subunits of polysomes is incompatible with the conventional model in which eIF-2 is recycled during the joining of the 48S preinitiation complex and the 60S subunit to form the 80S initiation complex. A modified model is presented with emphasis on the translocation of eIF-2 from the 40S ribosomal subunit of the 48S preinitiation complex (eIF-2.GTP.Met-tRNA(f).40S.mRNA) to the 60S subunit of the 80S initiation complex.
Harvard-Massachusetts Institute of Technology, Division of Health Sciences and Technology, Cambridge 02139.
K V
KV
Ramaiah
R S
RS
Dhindsa
J J
JJ
Chen
I M
IM
London
D
D
Levin
D000818:Animals; D002474:Cell-Free System; D003513:Cycloheximide / Q000494:pharmacology; D015852:Eukaryotic Initiation Factor-2 / Q000378:metabolism*; D020662:Guanine Nucleotide Exchange Factors; D006418:Heme / Q000378:metabolism; D066298:In Vitro Techniques; D010142:Pactamycin / Q000494:pharmacology; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D011132:Polyribosomes / Q000378:metabolism*; D011494:Protein Kinases / Q000378:metabolism*; D011506:Proteins / Q000378:metabolism*; D011691:Puromycin / Q000494:pharmacology; D011817:Rabbits; D012156:Reticulocytes / Q000378:metabolism; D012270:Ribosomes / Q000378:metabolism* / Q000648:ultrastructure; D019892:eIF-2 Kinase
D015852:Eukaryotic Initiation Factor-2; D020662:Guanine Nucleotide Exchange Factors; D010750:Phosphoproteins; D011506:Proteins; D010142:Pactamycin; D006418:Heme; D011691:Puromycin; D003513:Cycloheximide; D011494:Protein Kinases; D019892:eIF-2 Kinase