Home LiteratureArticle Details
PMID: 1361233 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

Recycling and phosphorylation of eukaryotic initiation factor 2 on 60S subunits of 80S initiation complexes and polysomes.

Proceedings of the National Academy of Sciences of the United States of America ·Vol. 89 ·No. 24 ·1992-12-15 ·Pages 12063-7

Ramaiah KV, Dhindsa RS, Chen JJ, London IM, Levin D

Abstract

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.

MeSH Terms
Animals Cell-Free System Cycloheximide/pharmacology Eukaryotic Initiation Factor-2/metabolism Guanine Nucleotide Exchange Factors Heme/metabolism In Vitro Techniques Pactamycin/pharmacology Phosphoproteins/metabolism Phosphorylation Polyribosomes/metabolism Protein Kinases/metabolism Proteins/metabolism Puromycin/pharmacology Rabbits Reticulocytes/metabolism Ribosomes/metabolism,ultrastructure eIF-2 Kinase
Chemicals
Eukaryotic Initiation Factor-2 Guanine Nucleotide Exchange Factors Phosphoproteins Proteins Pactamycin Heme Puromycin Cycloheximide Protein Kinases eIF-2 Kinase
Authors & Affiliations
5 authors, click to expand affiliations / ORCID
Ramaiah K V
Harvard-Massachusetts Institute of Technology, Division of Health Sciences and Technology, Cambridge 02139.
Dhindsa R S
Chen J J
London I M
Levin D
References (31)
31 references, click to expand
  1. Poliovirus replicase stimulation by terminal uridylyl transferase.
    J Biol Chem. 1985 Jun 25;260(12):7628-35 PMID: 2987262
  2. Regulation of a protein synthesis initiation factor by adenovirus virus-associated RNA.
    Nature. 1985 Jan 17-23;313(5999):196-200 PMID: 2578613
  3. The association of NADPH with the guanine nucleotide exchange factor from rabbit reticulocytes: a role of pyridine dinucleotides in eukaryotic polypeptide chain initiation.
    Proc Natl Acad Sci U S A. 1986 Sep;83(18):6746-50 PMID: 3462724
  4. Regulation of polypeptide chain initiation in Chinese hamster ovary cells with a temperature-sensitive leucyl-tRNA synthetase. Changes in phosphorylation of initiation factor eIF-2 and in the activity of the guanine nucleotide exchange factor GEF.
    J Biol Chem. 1987 Jan 15;262(2):767-71 PMID: 3805006
  5. The 60 S ribosomal subunit as a carrier of eukaryotic initiation factor 2 and the site of reversing factor activity during protein synthesis.
    J Biol Chem. 1985 Aug 15;260(17):9860-6 PMID: 3926772
  6. Assembly and breakdown of mammalian protein synthesis initiation complexes: regulation by guanine nucleotides and by phosphorylation of initiation factor eIF-2.
    Biochemistry. 1983 Feb 15;22(4):726-33 PMID: 6551177
  7. A GDP/GTP exchange factor essential for eukaryotic initiation factor 2 cycling in Ehrlich ascites tumor cells and its regulation by eukaryotic initiation factor 2 phosphorylation.
    J Biol Chem. 1983 Jul 10;258(13):7928-34 PMID: 6553052
  8. Distribution of reversing factor in reticulocyte lysates during active protein synthesis and on inhibition by heme deprivation or double-stranded RNA.
    Proc Natl Acad Sci U S A. 1984 Nov;81(22):6998-7002 PMID: 6594676
  9. Mechanism of polypeptide chain initiation in eukaryotes and its control by phosphorylation of the alpha subunit of initiation factor 2.
    Proc Natl Acad Sci U S A. 1982 Apr;79(8):2537-40 PMID: 6953412
  10. Protein synthesis in rabbit reticulocytes: characteristics of the protein factor RF that reverses inhibition of protein synthesis in heme-deficient reticulocyte lysates.
    Proc Natl Acad Sci U S A. 1982 Nov;79(21):6517-21 PMID: 6959132
  11. Specificity of the protein kinase activity associated with the hemin-controlled repressor of rabbit reticulocyte.
    Proc Natl Acad Sci U S A. 1976 Sep;73(9):3078-82 PMID: 184458
  12. Purification and characterization of a protein factor that reverses the inhibition of protein synthesis by the heme-regulated translational inhibitor in rabbit reticulocyte lysates.
    Eur J Biochem. 1979 Aug 1;98(2):513-20 PMID: 488111
  13. Additional evidence that the hemin-controlled translational repressor from rabbit reticulocytes is a protein kinase.
    Biochem Biophys Res Commun. 1977 Jan 24;74(2):559-69 PMID: 836310
  14. Translational control in mammalian cells.
    Annu Rev Biochem. 1991;60:717-55 PMID: 1883206
  15. Complex formation by positive and negative translational regulators of GCN4.
    Mol Cell Biol. 1991 Jun;11(6):3217-28 PMID: 2038327
  16. Disulfide bond formation in the regulation of eIF-2 alpha kinase by heme.
    J Biol Chem. 1989 Jun 5;264(16):9559-64 PMID: 2722851
  17. Correlation between the distribution of the reversing factor and eukaryotic initiation factor 2 in heme-deficient or double-stranded RNA-inhibited reticulocyte lysates.
    FEBS Lett. 1988 Aug 15;236(1):179-84 PMID: 3402612
  18. 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.
    J Biol Chem. 1987 May 15;262(14):6899-907 PMID: 3646234
  19. Evidence that the primary effect of phosphorylation of eukaryotic initiation factor 2(alpha) in rabbit reticulocyte lysate is inhibition of the release of eukaryotic initiation factor-2.GDP from 60 S ribosomal subunits.
    J Biol Chem. 1985 Aug 5;260(16):9491-500 PMID: 3848434
  20. Control of globin synthesis: the role of heme.
    J Mol Biol. 1972 May 28;66(3):471-81 PMID: 5037023
  21. Effect of phosphorylation of the alpha-subunit of eukaryotic initiation factor 2 on the function of reversing factor in the initiation of protein synthesis.
    Proc Natl Acad Sci U S A. 1983 May;80(9):2559-63 PMID: 6573671
  22. Purification of the eukaryotic initiation factor 2-eukaryotic initiation factor 2B complex and characterization of its guanine nucleotide exchange activity during protein synthesis initiation.
    J Biol Chem. 1983 Mar 10;258(5):3402-8 PMID: 6826566
  23. Phosphorylation inhibits guanine nucleotide exchange on eukaryotic initiation factor 2.
    Nature. 1982 Mar 4;296(5852):93-5 PMID: 7063012
  24. Regulation of protein synthesis in reticulocyte lysates: phosphorylation of methionyl-tRNAf binding factor by protein kinase activity of translational inhibitor isolated from hemedeficient lysates.
    Proc Natl Acad Sci U S A. 1976 Sep;73(9):3112-6 PMID: 184460
  25. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications.
    Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350-4 PMID: 388439
  26. Phosphorylation of initiation factor elF-2 and the control of reticulocyte protein synthesis.
    Cell. 1977 May;11(1):187-200 PMID: 559547
  27. Evidence that glucose 6-phosphate regulates protein synthesis initiation in reticulocyte lysates.
    J Biol Chem. 1978 Oct 25;253(20):7163-72 PMID: 701241
  28. Release and recycling of eukaryotic initiation factor 2 in the formation of an 80 S ribosomal polypeptide chain initiation complex.
    J Biol Chem. 1992 Jun 25;267(18):12964-72 PMID: 1618793
  29. Ribosome association of GCN2 protein kinase, a translational activator of the GCN4 gene of Saccharomyces cerevisiae.
    Mol Cell Biol. 1991 Jun;11(6):3027-36 PMID: 2038314
  30. A novel role for aminoacyl-tRNA synthetases in the regulation of polypeptide chain initiation.
    Eur J Biochem. 1989 Jun 1;182(1):1-9 PMID: 2543569
  31. Fate of reversing factor during restoration of protein synthesis by hemin or GTP in heme-deficient reticulocyte lysates.
    Proc Natl Acad Sci U S A. 1986 Mar;83(5):1217-21 PMID: 3456582
Article Info
Journal
Proceedings of the National Academy of Sciences of the United States of America
Abbr.
Proc Natl Acad Sci U S A
ISSN
0027-8424
DOI
10.1073/pnas.89.24.12063
Published
1992-12-15
Pages
12063-7
Language
English
Region
United States
NLM ID
7505876
PMCID
PMC50698
Subset
IM
Grants
NIDDK NIH HHS · R01 DK016272 · United States
NIDDK NIH HHS · DK-16272 · United States
NIGMS NIH HHS · GM-24825 · United States
External Links
PubMed source DOI: 10.1073/pnas.89.24.12063 PMC Full Text View journal details
Analysis Services
Analysis Services

Contact

No. 2 Wenbo Road, Zhangqiu District, Jinan, Shandong

Qilu Normal University · Genelibs Bioinformatics Lab

750 Shunhua Rd, Jinan

2F, Bldg F, University Science Park

Tel: 0531-88819269

WeChat Official Account

Follow our WeChat subscription account for real-time updates and the latest in medical and biological research.


Business Email

E-mail: product@genelibs.com

Copyright © 2026.Genelibs Jinan Honghui Biotechnology Co., Ltd. 鲁ICP备20007159号-2