The joining of ribosomal subunits in eukaryotes requires eIF5B (original) (raw)

Nature volume 403, pages 332–335 (2000)Cite this article

Abstract

Initiation of eukaryotic protein synthesis begins with the ribosome separated into its 40S and 60S subunits1. The 40S subunit first binds eukaryotic initiation factor (eIF) 3 and an eIF2–GTP–initiator transfer RNA ternary complex. The resulting complex requires eIF1, eIF1A, eIF4A, eIF4B and eIF4F to bind to a messenger RNA and to scan to the initiation codon2. eIF5 stimulates hydrolysis of eIF2-bound GTP and eIF2 is released from the 48S complex formed at the initiation codon before it is joined by a 60S subunit to form an active 80S ribosome3,4,5,6,7,8. Here we show that hydrolysis of eIF2-bound GTP induced by eIF5 in 48S complexes is necessary but not sufficient for the subunits to join. A second factor termed eIF5B (relative molecular mass 175,000) is essential for this process. It is a homologue of the prokaryotic initiation factor IF2 (refs 6, 7) and, like it8,9,10,11,12, mediates joining of subunits and has a ribosome-dependent GTPase activity that is essential for its function.

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References

  1. Merrick, W. C. Mechanism and regulation of eukaryotic protein synthesis. Microbiol. Rev. 56, 291–315 ( 1992).
    CAS PubMed PubMed Central Google Scholar
  2. Pestova, T. V., Borukhov, S. I. & Hellen, C. U. T. Eukaryotic ribosomes require initiation factors 1 and 1A to locate initiation codons. Nature 394, 854–859 (1998).
    Article ADS CAS Google Scholar
  3. Chakrabarti, A. & Maitra, U. Functions of eukaryotic initiation factor 5 in the formation of an 80S ribosomal polypeptide chain initiation complex. J. Biol. Chem. 266, 14039–14045 (1991).
    CAS PubMed Google Scholar
  4. Das, K., Chesevich, J. & Maitra, U. Molecular cloning and expression of cDNA for mammalian translation initiation factor 5. Proc. Natl Acad. Sci. USA 90, 3058–3062 (1993).
    Article ADS CAS PubMed Google Scholar
  5. Huang, H.-K., Yoon, H., Hannig, E. M. & Donahue, T. F. GTP hydrolysis controls stringent selection of the AUG start codon during translation initiation in Saccharomyces cerevisiae. Genes Dev. 11, 2396–2413 (1997).
    Article CAS PubMed PubMed Central Google Scholar
  6. Choi, S. K., Lee, J. H., Zoll, W. L., Merrick, W. C. & Dever, T. E. Promotion of Met-tRNAMet binding to ribosomes by yIF2, a bacterial IF2 homolog in yeast. Science 280, 1757–1760 (1998).
    Article ADS CAS PubMed Google Scholar
  7. Lee, J. H., Choi, S. K., Roll-Mecak, A., Burley, S. K. & Dever, T. E. Universal conservation in translation initiation revealed by human and archaeal homologs of bacterial translation factor IF2. Proc. Natl Acad. Sci. USA 96, 4342–4347 (1999).
    Article ADS CAS PubMed Google Scholar
  8. Sacerdot, C., Dessen, P., Hershey, J. W. B., Plumbridge, J. A. & Grunberg-Manago, M. Sequence of the initiation factor IF2 gene; unusual protein features and homologies with elongation factors. Proc. Natl Acad. Sci. USA 81, 7787– 7791 (1984).
    Article ADS CAS PubMed Google Scholar
  9. Kolakofsky, D., Dewey, K. F., Hershey, J. W. B. & Thach, R. E. Guanosine 5′-triphosphatase activity of initiation factor f2. Proc. Natl Acad. Sci. USA 61, 1066– 1070 (1968).
    Article ADS CAS PubMed Google Scholar
  10. Godefroy-Colburn, T. et al. Light-scattering studies showing the effect of initiation factors on the reversible dissociation of Escherichia coli ribosomes. J. Mol. Biol. 94, 461– 478 (1975).
    Article PubMed Google Scholar
  11. Luchin, S. et al. In vitro study of two dominant inhibitory GTPase mutants of Escherichia coli translation initiation factor IF2. Direct evidence that GTP hydrolysis is necessary for factor recycling. J. Biol. Chem. 274, 6074–6079 ( 1999).
    Article CAS PubMed Google Scholar
  12. Lockwood, A. H., Sarkar, P. & Maitra, U. Release of polypeptide chain initiation factor IF-2 during initiation complex formation. Proc. Natl Acad. Sci. USA 69, 3602–3605 ( 1972).
    Article ADS CAS PubMed Google Scholar
  13. Merrick, W. C., Kemper, W. M. & Anderson, W. F. Purification and characterization of homogenous initiation factor M2A from rabbit reticulocytes. J. Biol. Chem. 250, 5556–5562 (1975).
    CAS PubMed Google Scholar
  14. Trachsel, H., Emi, B., Schreier, M. H. & Staehelin, T. Initiation of mammalian protein synthesis. II. The assembly of the initiation complex with purified initiation factors. J. Mol. Biol. 116, 755–767 (1977).
    Article CAS PubMed Google Scholar
  15. Benne, R., Brown-Luedi, M. L. & Hershey, J. W. B. Purification and characterization of protein synthesis initiation factors eIF-1, eIF-4C, eIF-4D, and eIF-5 from rabbit reticulocytes. J. Biol. Chem. 253, 3070– 3077 (1978).
    CAS PubMed Google Scholar
  16. Peterson, D. T., Safer, B. & Merrick, W. C. Role of eukaryotic initiation factor 5 in the formation of 80S initiation complexes. J. Biol. Chem. 254, 7730–7735 (1979).
    CAS PubMed Google Scholar
  17. Pestova, T. V., Shatsky, I. N., Fletcher, S. P., Jackson, R. J. & Hellen, C. U. T. A prokaryotic-like mode of binding of cytoplasmic eukaryotic ribosomes to the initiation codon during internal initiation of translation of Hepatitis C and Classical Swine fever virus RNAs. Genes Dev. 12, 67– 83 (1998).
    Article CAS PubMed PubMed Central Google Scholar

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Acknowledgements

We thank W. Merrick for discussions, D. Etchison and R. Schneider for antibodies, and L. Siconolfi-Baez for sequencing eIF5B. These studies were supported by grants from the NIH to C.U.T.H. and T.V.P.

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Authors and Affiliations

  1. Department of Microbiology and Immunology State University of New York Health Science Center at Brooklyn, 450 Clarkson Avenue, Brooklyn, 11203, New York, USA
    Tatyana V. Pestova, Ivan B. Lomakin & Christopher U. T. Hellen
  2. A.N. Belozersky Institute of Physico-Chemical Biology, Moscow State University, Moscow, 119899, Russia
    Tatyana V. Pestova
  3. Laboratory of Eukaryotic Gene Regulation, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, 20892-2716, Maryland, USA
    Joon H. Lee, Sang Ki Choi & Thomas E. Dever

Authors

  1. Tatyana V. Pestova
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  2. Ivan B. Lomakin
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  3. Joon H. Lee
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  4. Sang Ki Choi
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  5. Thomas E. Dever
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  6. Christopher U. T. Hellen
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Correspondence toTatyana V. Pestova.

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Pestova, T., Lomakin, I., Lee, J. et al. The joining of ribosomal subunits in eukaryotes requires eIF5B.Nature 403, 332–335 (2000). https://doi.org/10.1038/35002118

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