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. 1975 Oct;72(10):4157–4161. doi: 10.1073/pnas.72.10.4157

Complete translation of poliovirus RNA in a eukaryotic cell-free system.

L Villa-Komaroff, N Guttman, D Baltimore, H F Lodishi
PMCID: PMC433159  PMID: 172898

Abstract

Poliovirus RNA stimulates imcorporation of 35S from both [35S]methionine and formyl-[35S]methionyl-tRNAfMet in cell-free systems derived from HeLa cells or from poliovirus-infected HeLa cells. The largest product formed under the direction of the viral RNA is the same size as the polyprotein thought to represent translation of the entire RNA. Synthesis of this polyprotein and other large products was stimulated greatly by increasing the salt concentration during the reaction from the optimum for initiation (90 mM) to the optimum for elongation (155 mM). Only one initiation peptide could be identified, and a tryptic digest of the product contained mainly peptides that cochromatographed with peptides from authentic viral proteins. The RNA from a deletion mutant of poliovirus initiated protein synthesis at the same site used by standard RNA and programmed synthesis of an appropriately deleted set of polypeptides. The results strongly support the model of translation of poliovirus RNA from a single initiation site into a continuous polyprotein that is cleaved to form the functional proteins. It is suggested that uninfected HeLa cell extracts can carry out the cleavages of nascent polyprotein.

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Selected References

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

  1. Baltimore D., Girard M., Darnell J. E. Aspects of the synthesis of poliovirus RNA and the formation of virus particles. Virology. 1966 Jun;29(2):179–189. doi: 10.1016/0042-6822(66)90024-9. [DOI] [PubMed] [Google Scholar]
  2. Boime I., Leder P. Protein synthesis directed by encephalomyocarditis virus mRNA. 3. Discrete polypeptides translated from a monocistronic messenger in vitro. Arch Biochem Biophys. 1972 Dec;153(2):706–713. doi: 10.1016/0003-9861(72)90389-x. [DOI] [PubMed] [Google Scholar]
  3. Cole C. N., Baltimore D. Defective interfering particles of poliovirus. II. Nature of the defect. J Mol Biol. 1973 May 25;76(3):325–343. doi: 10.1016/0022-2836(73)90508-1. [DOI] [PubMed] [Google Scholar]
  4. Cole C. N., Smoler D., Wimmer E., Baltimore D. Defective interfering particles of poliovirus. I. Isolation and physical properties. J Virol. 1971 Apr;7(4):478–485. doi: 10.1128/jvi.7.4.478-485.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Eggen K. L., Shatkin A. J. In vitro translation of cardiovirus ribonucleic acid by mammalian cell-free extracts. J Virol. 1972 Apr;9(4):636–645. doi: 10.1128/jvi.9.4.636-645.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Esteban M., Kerr I. M. The synthesis of encephalomyocarditis virus polypeptides in infected L-cells and cell-free systems. Eur J Biochem. 1974 Jun 15;45(2):567–576. doi: 10.1111/j.1432-1033.1974.tb03583.x. [DOI] [PubMed] [Google Scholar]
  7. Granboulan N., Girard M. Molecular weight of poliovirus ribonucleic acid. J Virol. 1969 Oct;4(4):475–479. doi: 10.1128/jvi.4.4.475-479.1969. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Housman D., Jacobs-Lorena M., Rajbhandary U. L., Lodish H. F. Initiation of haemoglobin synthesis by methionyl-tRNA. Nature. 1970 Aug 29;227(5261):913–918. doi: 10.1038/227913a0. [DOI] [PubMed] [Google Scholar]
  9. Huang A. S., Balitmore D. Initiation of polyribosome formation in poliovirus-infected HeLa cells. J Mol Biol. 1970 Feb 14;47(3):275–291. doi: 10.1016/0022-2836(70)90302-5. [DOI] [PubMed] [Google Scholar]
  10. Jacobson M. F., Asso J., Baltimore D. Further evidence on the formation of poliovirus proteins. J Mol Biol. 1970 May 14;49(3):657–669. doi: 10.1016/0022-2836(70)90289-5. [DOI] [PubMed] [Google Scholar]
  11. Jacobson M. F., Baltimore D. Polypeptide cleavages in the formation of poliovirus proteins. Proc Natl Acad Sci U S A. 1968 Sep;61(1):77–84. doi: 10.1073/pnas.61.1.77. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Kerr I. M., Brown R. E., Tovell D. R. Characterization of the polypeptides formed in response to encephalomyocarditis virus ribonucleic acid in a cell-free system from mouse ascites tumor cells. J Virol. 1972 Jul;10(1):73–81. doi: 10.1128/jvi.10.1.73-81.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. 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]
  14. Mathews M. B., Osborn M. The rate of polypeptide chain elongation in a cell-free system from Krebs II ascites cells. Biochim Biophys Acta. 1974 Mar 8;340(2):147–152. doi: 10.1016/0005-2787(74)90107-5. [DOI] [PubMed] [Google Scholar]
  15. McDowell M. J., Joklik W. K., Villa-Komaroff L., Lodish H. F. Translation of reovirus messenger RNAs synthetesized in vitro into reovirus polypeptides by several mammalian cell-free extracts. Proc Natl Acad Sci U S A. 1972 Sep;69(9):2649–2653. doi: 10.1073/pnas.69.9.2649. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Oberg B. F., Shatkin A. J. Initiation of picornavirus protein synthesis in ascites cell extracts. Proc Natl Acad Sci U S A. 1972 Dec;69(12):3589–3593. doi: 10.1073/pnas.69.12.3589. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Osborn M., Weber K., Lodish H. F. Amino terminal peptides of RNA phage proteins synthesized in the cell free system. Biochem Biophys Res Commun. 1970 Nov 9;41(3):748–756. doi: 10.1016/0006-291x(70)90077-x. [DOI] [PubMed] [Google Scholar]
  18. PENMAN S., BECKER Y., DARNELL J. E. A CYTOPLASMIC STRUCTURE INVOLVED IN THE SYNTHESIS AND ASSEMBLY OF POLIOVIRUS COMPONENTS. J Mol Biol. 1964 Apr;8:541–555. doi: 10.1016/s0022-2836(64)80010-3. [DOI] [PubMed] [Google Scholar]
  19. Rekosh D. Gene order of the poliovirus capsid proteins. J Virol. 1972 Mar;9(3):479–487. doi: 10.1128/jvi.9.3.479-487.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Smith A. E. The initiation of protein synthesis directed by the RNA from encephalomyocarditis virus. Eur J Biochem. 1973 Mar 1;33(2):301–313. doi: 10.1111/j.1432-1033.1973.tb02684.x. [DOI] [PubMed] [Google Scholar]
  21. Spector D. H., Baltimore D. Polyadenylic acid on poliovirus RNA. II. poly(A) on intracellular RNAs. J Virol. 1975 Jun;15(6):1418–1431. doi: 10.1128/jvi.15.6.1418-1431.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Summers D. F., Maizel J. V., Jr Determination of the gene sequence of poliovirus with pactamycin. Proc Natl Acad Sci U S A. 1971 Nov;68(11):2852–2856. doi: 10.1073/pnas.68.11.2852. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Taber R., Rekosh D., Baltimore D. Effect of pactamycin on synthesis of poliovirus proteins: a method for genetic mapping. J Virol. 1971 Oct;8(4):395–401. doi: 10.1128/jvi.8.4.395-401.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Villa-Komaroff L., McDowell M., Baltimore D., Lodish H. F. Translation of reovirus mRNA, poliovirus RNA and bacteriophage Qbeta RNA in cell-free extracts of mammalian cells. Methods Enzymol. 1974;30:709–723. doi: 10.1016/0076-6879(74)30068-7. [DOI] [PubMed] [Google Scholar]

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