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. 1982 Jan;41(1):30–41. doi: 10.1128/jvi.41.1.30-41.1982

Genomic RNA of mengovirus V. Recognition of common features by ribosomes and eucaryotic initiation factor 2.

R Perez-Bercoff, R Kaempfer
PMCID: PMC256723  PMID: 6283122

Abstract

Binding of ribosomes to the 32P-labeled genomic RNA of mengovirus was studied in lysates of mouse L929 and Krebs ascites cells under conditions for initiation of translation. Upon total digestion with RNase T1, the 32P-labeled RNA protected in either 40S or 80S initiation complexes yielded four unique, large oligonucleotides. Each of these oligonucleotides occurred once in the viral RNA molecule. The same four oligonucleotides were recovered from 80S initiation complexes formed in lysates in which unlabeled mengovirus RNA had been translated extensively, indicating that recognition by ribosomes was not modulated detectably by a viral translation product. The recognition of intact, 32P-labeled mengovirus RNA by eucaryotic initiation factor 2 (eIF-2) was examined by direct complex formation. Fingerprint analysis of the RNA protected by eIF-2 against RNase T1 digestion yielded three T1 oligonucleotides that were identical to three of the four oligonucleotides protected in either 40S or 80S initiation complexes. A physical map of the large T1 oligonucleotides of the mengovirus RNA molecule was constructed, and the four protected oligonucleotides were found to map internally, within the region between the polycytidylate tract and the 3' end. For either ribosomes or eIF-2, the protected oligonucleotides could not be arranged in a continuous sequence, suggesting that they constitute at least two widely separated domains. These results show that ribosomes recognize and blind to more than a single sequence in mengovirus RNA, located internally in regions that are far removed from the 5' end of the molecule. eIF-2 itself binds with high specificity to mengovirus RNA, recognizing apparently three of the four sequences recognized by ribosomes.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Ahlquist P., Dasgupta R., Shih D. S., Zimmern D., Kaesberg P. Two-step binding of eukaryotic ribosomes to brome mosaic virus RNA3. Nature. 1979 Sep 27;281(5729):277–282. doi: 10.1038/281277a0. [DOI] [PubMed] [Google Scholar]
  2. Barrieux A., Rosenfeld M. G. Characterization of GTP-dependent Met-tRNAf binding protein. J Biol Chem. 1977 Jun 10;252(11):3843–3847. [PubMed] [Google Scholar]
  3. Barrieux A., Rosenfeld M. G. mRNA-induced dissociation of initiation factor 2. J Biol Chem. 1978 Sep 25;253(18):6311–6314. [PubMed] [Google Scholar]
  4. Browning K. S., Leung D. W., Clark J. M., Jr Protection of satellite tobacco necrosis virus ribonucleic acid by wheat germ 40S and 80S ribosomes. Biochemistry. 1980 May 13;19(10):2276–2283. doi: 10.1021/bi00551a044. [DOI] [PubMed] [Google Scholar]
  5. Celma M. L., Ehrenfeld E. Translation of poliovirus RNA in vitro: detection of two different initiation sites. J Mol Biol. 1975 Nov 15;98(4):761–780. doi: 10.1016/s0022-2836(75)80009-x. [DOI] [PubMed] [Google Scholar]
  6. Chen Y. C., Woodley C. L., Bose K. K., Gupta K. K. Protein synthesis in rabbit reticulocytes: characteristics of a Met-tRNA Met f binding factor. Biochem Biophys Res Commun. 1972 Jul 11;48(1):1–9. doi: 10.1016/0006-291x(72)90335-x. [DOI] [PubMed] [Google Scholar]
  7. Coffin J. M., Billeter M. A. A physical map of the Rous sarcoma virus genome. J Mol Biol. 1976 Jan 25;100(3):293–318. doi: 10.1016/s0022-2836(76)80065-4. [DOI] [PubMed] [Google Scholar]
  8. Darlix J. L., Spahr P. F., Bromley P. A., Jaton J. C. In vitro, the major ribosome binding site on Rous sarcoma virus RNA does not contain the nucleotide sequence coding for the N-terminal amino acids of the gag gene product. J Virol. 1979 Feb;29(2):597–611. doi: 10.1128/jvi.29.2.597-611.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Darnbrough C., Legon S., Hunt T., Jackson R. J. Initiation of protein synthesis: evidence for messenger RNA-independent binding of methionyl-transfer RNA to the 40 S ribosomal subunit. J Mol Biol. 1973 May 25;76(3):379–403. doi: 10.1016/0022-2836(73)90511-1. [DOI] [PubMed] [Google Scholar]
  10. Dasgupta R., Shih D. S., Saris C., Kaesberg P. Nucleotide sequence of a viral RNA fragment that binds to eukaryotic ribosomes. Nature. 1975 Aug 21;256(5519):624–628. doi: 10.1038/256624a0. [DOI] [PubMed] [Google Scholar]
  11. Dettman G. L., Stanley W. M., Jr Recognition of eukaryotic initiator tRNA by an initiation factor and the transfer of the methionine moiety into peptide linkage. Biochim Biophys Acta. 1972 Nov 16;287(1):124–133. doi: 10.1016/0005-2787(72)90336-x. [DOI] [PubMed] [Google Scholar]
  12. Di Segni G., Rosen H., Kaempfer R. Competition between alpha- and beta-globin messenger ribonucleic acids for eucaryotic initiation factor 2. Biochemistry. 1979 Jun 26;18(13):2847–2854. doi: 10.1021/bi00580a027. [DOI] [PubMed] [Google Scholar]
  13. Domingo E., Dávila M., Ortín J. Nucleotide sequence heterogeneity of the RNA from a natural population of foot-and-mouth-disease virus. Gene. 1980 Nov;11(3-4):333–346. doi: 10.1016/0378-1119(80)90073-6. [DOI] [PubMed] [Google Scholar]
  14. Egberts E., Hackett P. B., Traub P. Shutoff of histone synthesis by Mengovirus infection of Ehrlich ascites tumor cells. Hoppe Seylers Z Physiol Chem. 1977 Apr;358(4):463–474. doi: 10.1515/bchm2.1977.358.1.463. [DOI] [PubMed] [Google Scholar]
  15. Ehrenfeld E., Brown D. Stability of poliovirus RNA in cell-free translation systems utilizing two initiation sites. J Biol Chem. 1981 Mar 25;256(6):2656–2661. [PubMed] [Google Scholar]
  16. Golini F., Thach S. S., Birge C. H., Safer B., Merrick W. C., Thach R. E. Competition between cellular and viral mRNAs in vitro is regulated by a messenger discriminatory initiation factor. Proc Natl Acad Sci U S A. 1976 Sep;73(9):3040–3044. doi: 10.1073/pnas.73.9.3040. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hackett P. B., Egberts E., Traub P. Selective translation of mengovirus RNA over Host mRNA in homologous, fractionated, cell-free translational systems from Ehrlich-ascites-tumor cells. Eur J Biochem. 1978 Feb;83(2):353–361. doi: 10.1111/j.1432-1033.1978.tb12101.x. [DOI] [PubMed] [Google Scholar]
  18. Hackett P. B., Egberts E., Traub P. Translation of ascites and mengovirus RNA in fractionated cell-free systems from uninfected and mengovirus-infected Ehrlich-ascites-tumor cells. Eur J Biochem. 1978 Feb;83(2):341–352. doi: 10.1111/j.1432-1033.1978.tb12100.x. [DOI] [PubMed] [Google Scholar]
  19. Hellerman J. G., Shafritz D. A. Interaction of poly(A) and mRNA with eukaryotic initiator met-tRNA-f binding factor: identification of this activity on reticulocyte ribonucleic acid protein particles. Proc Natl Acad Sci U S A. 1975 Mar;72(3):1021–1025. doi: 10.1073/pnas.72.3.1021. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Horst J., Fraenkel-Conrat H., Mandeles S. Terminal heterogeneity at both ends of the satellite tobacco necrosis virus ribonucleic acid. Biochemistry. 1971 Dec 7;10(25):4748–4752. doi: 10.1021/bi00801a022. [DOI] [PubMed] [Google Scholar]
  21. Jense H., Knauert F., Ehrenfeld E. Two initiation sites for translation of poliovirus RNA in vitro: comparison of LSc and Mahoney strains. J Virol. 1978 Oct;28(1):387–394. doi: 10.1128/jvi.28.1.387-394.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Kaempfer R. Binding of messenger RNA in initiation of eukaryotic translation. Methods Enzymol. 1979;60:380–392. doi: 10.1016/s0076-6879(79)60036-8. [DOI] [PubMed] [Google Scholar]
  23. Kaempfer R., Hollender R., Abrams W. R., Israeli R. Specific binding of messenger RNA and methionyl-tRNAfMet by the same initiation factor for eukaryotic protein synthesis. Proc Natl Acad Sci U S A. 1978 Jan;75(1):209–213. doi: 10.1073/pnas.75.1.209. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Kaempfer R., Hollender R., Soreq H., Nudel U. Recognition of messenger RNA in eukaryotic protein synthesis. Equilibrium studies of the interaction between messenger RNA and the initiation factor that binds methionyl-tRNAf. Eur J Biochem. 1979 Mar;94(2):591–600. doi: 10.1111/j.1432-1033.1979.tb12929.x. [DOI] [PubMed] [Google Scholar]
  25. Kaempfer R., Rosen H., Israeli R. Translational control: recognition of the methylated 5' end and an internal sequence in eukaryotic mRNA by the initiation factor that binds methionyl-tRNAfMet. Proc Natl Acad Sci U S A. 1978 Feb;75(2):650–654. doi: 10.1073/pnas.75.2.650. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Knauert F., Ehrenfeld E. Translation of poliovirus RNA in vitro: studies on n-formylmethionine-labeled polypeptides initiated in cell-free extracts prepared from poliovirus infected HeLa cells. Virology. 1979 Mar;93(2):537–546. doi: 10.1016/0042-6822(79)90256-3. [DOI] [PubMed] [Google Scholar]
  27. Kozak M. How do eucaryotic ribosomes select initiation regions in messenger RNA? Cell. 1978 Dec;15(4):1109–1123. doi: 10.1016/0092-8674(78)90039-9. [DOI] [PubMed] [Google Scholar]
  28. Kozak M. Role of ATP in binding and migration of 40S ribosomal subunits. Cell. 1980 Nov;22(2 Pt 2):459–467. doi: 10.1016/0092-8674(80)90356-6. [DOI] [PubMed] [Google Scholar]
  29. Kozak M., Shatkin A. J. Characterization of translational initiation regions from eukaryotic messenger RNAs. Methods Enzymol. 1979;60:360–375. doi: 10.1016/s0076-6879(79)60034-4. [DOI] [PubMed] [Google Scholar]
  30. Kozak M., Shatkin A. J. Sequences of two 5'-terminal ribosome-protected fragments from reovirus messenger RNAs. J Mol Biol. 1977 May 5;112(1):75–96. doi: 10.1016/s0022-2836(77)80157-5. [DOI] [PubMed] [Google Scholar]
  31. Lawrence C., Thach R. E. Encephalomyocarditis virus infection of mouse plasmacytoma cells. I. Inhibition of cellular protein synthesis. J Virol. 1974 Sep;14(3):598–610. doi: 10.1128/jvi.14.3.598-610.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Legon S. Characterization of the ribosome-protected regions of 125I-labelled rabbit globin messenger RNA. J Mol Biol. 1976 Sep 5;106(1):37–53. doi: 10.1016/0022-2836(76)90299-0. [DOI] [PubMed] [Google Scholar]
  33. Legon S., Robertson H. D. The binding of 125I-labelled rabbit globin messenger RNA to reticulocyte ribosomes. J Mol Biol. 1976 Sep 5;106(1):23–36. doi: 10.1016/0022-2836(76)90298-9. [DOI] [PubMed] [Google Scholar]
  34. Levin D. H., Kyner D., Acs G. Protein initiation in eukaryotes: formation and function of a ternary complex composed of a partially purified ribosomal factor, methionyl transfer RNA, and guanosine triphosphate. Proc Natl Acad Sci U S A. 1973 Jan;70(1):41–45. doi: 10.1073/pnas.70.1.41. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Lewis J. A., Falcoff E., Falcoff R. Dual action of double-stranded RNA inhibiting protein synthesis in extracts of interferon-treated mouse L cells. Translation is impaired at the level of initiation and by mRNA degradation. Eur J Biochem. 1978 May 16;86(2):497–509. doi: 10.1111/j.1432-1033.1978.tb12333.x. [DOI] [PubMed] [Google Scholar]
  36. Lodish H. F., Rose J. K. Relative importance of 7-methylguanosine in ribosome binding and translation of vesicular stomatitis virus mRNA in wheat germ and reticulocyte cell-free systems. J Biol Chem. 1977 Feb 25;252(4):1181–1188. [PubMed] [Google Scholar]
  37. Nomoto A., Kitamura N., Golini F., Wimmer E. The 5'-terminal structures of poliovirion RNA and poliovirus mRNA differ only in the genome-linked protein VPg. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5345–5349. doi: 10.1073/pnas.74.12.5345. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Perez-Bercoff R., Gander M., Preisig E. The genomic RNA of mengovirus. II: Comparative analysis of the standard and denser virions. Virology. 1978 Apr;85(2):378–386. doi: 10.1016/0042-6822(78)90446-4. [DOI] [PubMed] [Google Scholar]
  39. Perez-Bercoff R., Gander M. The genomic RNA of mengovirus. I. Location of the poly(C) tract. Virology. 1977 Jul 15;80(2):426–429. doi: 10.1016/s0042-6822(77)80018-4. [DOI] [PubMed] [Google Scholar]
  40. Pettersson R. F., Flanegan J. B., Rose J. K., Baltimore D. 5'-Terminal nucleotide sequences of polio virus polyribosomal RNA and virion RNA are identical. Nature. 1977 Jul 21;268(5617):270–272. doi: 10.1038/268270a0. [DOI] [PubMed] [Google Scholar]
  41. Rose J. K., Trachsel H., Leong K., Baltimore D. Inhibition of translation by poliovirus: inactivation of a specific initiation factor. Proc Natl Acad Sci U S A. 1978 Jun;75(6):2732–2736. doi: 10.1073/pnas.75.6.2732. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Rosen H., Kaempfer R. Mutually exclusive binding of messenger RNA and initiator methionyl transfer RNA to eukaryotic initiation factor 2. Biochem Biophys Res Commun. 1979 Nov 28;91(2):449–455. doi: 10.1016/0006-291x(79)91542-0. [DOI] [PubMed] [Google Scholar]
  43. Sangar D. V., Black D. N., Rowlands D. J., Harris T. J., Brown F. Location of the initiation site for protein synthesis on foot-and-mouth disease virus RNA by in vitro translation of defined fragments of the RNA. J Virol. 1980 Jan;33(1):59–68. doi: 10.1128/jvi.33.1.59-68.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Schreier M. H., Staehelin T. Initiation of eukaryotic protein synthesis: (Met-tRNA f -40S ribosome) initiation complex catalysed by purified initiation factors in the absence of mRNA. Nat New Biol. 1973 Mar 14;242(115):35–38. doi: 10.1038/newbio242035a0. [DOI] [PubMed] [Google Scholar]
  45. Schroeder H. W., Jr, Liarakos C. D., Gupta R. C., Randerath K., O'Malley B. W. Ribosome binding site analysis of ovalbumin messenger ribonucleic acid. Biochemistry. 1979 Dec 25;18(26):5798–5808. doi: 10.1021/bi00593a009. [DOI] [PubMed] [Google Scholar]
  46. Shatkin A. J. Capping of eucaryotic mRNAs. Cell. 1976 Dec;9(4 Pt 2):645–653. doi: 10.1016/0092-8674(76)90128-8. [DOI] [PubMed] [Google Scholar]
  47. Shih D. S., Shih C. T., Kew O., Pallansch M., Rueckert R., Kaesberg P. Cell-free synthesis and processing of the proteins of poliovirus. Proc Natl Acad Sci U S A. 1978 Dec;75(12):5807–5811. doi: 10.1073/pnas.75.12.5807. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Svitkin Y. V., Agol V. I. Complete translation of encephalomyocarditis virus RNA and faithful cleavage of virus-specific proteins in a cell-free system from Krebs-2 cells. FEBS Lett. 1978 Mar 1;87(1):7–11. doi: 10.1016/0014-5793(78)80121-5. [DOI] [PubMed] [Google Scholar]
  49. Wang L. H., Duesberg P., Beemon K., Vogt P. K. Mapping RNase T1-resistant oligonucleotides of avian tumor virus RNAs: sarcoma-specific oligonucleotides are near the poly(A) end and oligonucleotides common to sarcoma and transformation-defective viruses are at the poly(A) end. J Virol. 1975 Oct;16(4):1051–1070. doi: 10.1128/jvi.16.4.1051-1070.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Wimmer E., Chang A. Y., Clark J. M., Jr, Reichmann M. E. Sequence studies of satellite tobacco necrosis virus RNA. Isolation and characterization of a 5'-terminal trinucleotide. J Mol Biol. 1968 Nov 28;38(1):59–73. doi: 10.1016/0022-2836(68)90128-9. [DOI] [PubMed] [Google Scholar]

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