Skip to main content
PMC home page
Journal of Virology logoLink to Journal of Virology
. 1972 Sep;10(3):504–515. doi: 10.1128/jvi.10.3.504-515.1972

Membrane-Associated Replication Complex in Arbovirus Infection 1

Robert M Friedman a, Judith G Levin a, Philip M Grimley a, Irene K Berezesky a
PMCID: PMC356492  PMID: 4342056

Abstract

Cytoplasmic extracts of chicken embryo fibroblast cells infected with Semliki Forest virus were subjected to isopycnic centrifugation in discontinuous sucrose gradients. Seven distinct bands were usually formed. The four upper bands contained predominantly smooth membranes and the lowest band was enriched in rough endoplasmic reticulum. One fraction (fraction 5), banding at a density of 1.16 g/cm3, was found to be heavily enriched in pulse-labeled ribonucleic acid (RNA), viral RNA polymerase, and viral RNA forms associated with RNA replication. Thus, fraction 5 evidently contained a membrane-associated viral replication complex of a type previously defined in picornavirus infections. Fraction 5 was also consistently enriched with unique membranous structures previously observed in intact cells as type 1 cytopathic vacuoles (CPV-1). When the CPV-1 in fraction 5 were isolated from cells briefly incubated with 3H-uridine and 3H-adenosine prior to cell disruption, a large proportion was found to be labeled by high-resolution autoradiography. Thus, ultrastructural, biochemical, and biological evidence were all consistent with the interpretation that the CPV-1 membranes represent a significant element of the viral replication complex.

Full text

PDF
506

Images in this article

508Image
on p.508
509Image
on p.509
509Image
on p.509
511Image
on p.511

Selected References

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

  1. Acheson N. H., Tamm I. Purification and properties of Semliki Forest virus nucleocapsids. Virology. 1970 Jun;41(2):306–320. doi: 10.1016/0042-6822(70)90083-8. [DOI] [PubMed] [Google Scholar]
  2. Acheson N. H., Tamm I. Replication of Semliki Forest virus: an electron microscopic study. Virology. 1967 May;32(1):128–143. doi: 10.1016/0042-6822(67)90261-9. [DOI] [PubMed] [Google Scholar]
  3. 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]
  4. Ben-Ishai Z., Goldblum N., Becker Y. The intracellular site and sequence of Sindbis virus replication. J Gen Virol. 1968 May;2(3):365–375. doi: 10.1099/0022-1317-2-3-365. [DOI] [PubMed] [Google Scholar]
  5. Bosmann H. B., Hagopian A., Eylar E. H. Cellular membranes: the isolation and characterization of the plasma and smooth membranes of HeLa cells. Arch Biochem Biophys. 1968 Oct;128(1):51–69. doi: 10.1016/0003-9861(68)90008-8. [DOI] [PubMed] [Google Scholar]
  6. CARO L. G., VAN TUBERGEN R. P., KOLB J. A. High-resolution autoradiography. I. Methods. J Cell Biol. 1962 Nov;15:173–188. doi: 10.1083/jcb.15.2.173. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Caliguiri L. A., Mosser A. G. Proteins associated with the poliovirus RNA replication complex. Virology. 1971 Nov;46(2):375–386. doi: 10.1016/0042-6822(71)90039-0. [DOI] [PubMed] [Google Scholar]
  8. Caliguiri L. A., Tamm I. Characterization of poliovirus-specific structures associated with cytoplasmic membranes. Virology. 1970 Sep;42(1):112–122. doi: 10.1016/0042-6822(70)90243-6. [DOI] [PubMed] [Google Scholar]
  9. Caliguiri L. A., Tamm I. Membranous structures associated with translation and transcription of poliovirus RNA. Science. 1969 Nov 14;166(3907):885–886. doi: 10.1126/science.166.3907.885. [DOI] [PubMed] [Google Scholar]
  10. Caro L. G. A common source of difficulty in high-resolution radioautography. J Cell Biol. 1969 Jun;41(3):918–919. doi: 10.1083/jcb.41.3.918. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Cartwright K. L., Burke D. Virus nucleic acids formed in chick embryo cells infected with Semliki Forest virus. J Gen Virol. 1970 Feb;6(2):231–248. doi: 10.1099/0022-1317-6-2-231. [DOI] [PubMed] [Google Scholar]
  12. DULBECCO R., VOGT M. Plaque formation and isolation of pure lines with poliomyelitis viruses. J Exp Med. 1954 Feb;99(2):167–182. doi: 10.1084/jem.99.2.167. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Friedman R. M., Berezesky I. K. Cytoplasmic fractions associated with Semliki Forest virus ribonucleic acid replication. J Virol. 1967 Apr;1(2):374–383. doi: 10.1128/jvi.1.2.374-383.1967. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Friedman R. M., Fantes K. H., Levy H. B., Carter W. B. Interferon action on parental Semliki forest virus ribonucleic acid. J Virol. 1967 Dec;1(6):1168–1173. doi: 10.1128/jvi.1.6.1168-1173.1967. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Friedman R. M., Grimley P. M. Inhibition of arbovirus assembly by cycloheximide. J Virol. 1969 Sep;4(3):292–299. doi: 10.1128/jvi.4.3.292-299.1969. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Friedman R. M., Levy H. B., Carter W. B. Replication of semliki forest virus: three forms of viral RNA produced during infection. Proc Natl Acad Sci U S A. 1966 Aug;56(2):440–446. doi: 10.1073/pnas.56.2.440. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Friedman R. M. Protein synthesis directed by an arbovirus. J Virol. 1968 Jan;2(1):26–32. doi: 10.1128/jvi.2.1.26-32.1968. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Friedman R. M., Sreevalsan T. Membrane binding of input arbovirus ribonucleic acid: effect of interferon or cycloheximide. J Virol. 1970 Aug;6(2):169–175. doi: 10.1128/jvi.6.2.169-175.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Friedman R. M. Structural and nonstructural proteins of an arbovirus. J Virol. 1968 Oct;2(10):1076–1080. doi: 10.1128/jvi.2.10.1076-1080.1968. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Grimley P. M., Berezesky I. K., Friedman R. M. Cytoplasmic structures associated with an arbovirus infection: loci of viral ribonucleic acid synthesis. J Virol. 1968 Nov;2(11):1326–1338. doi: 10.1128/jvi.2.11.1326-1338.1968. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Kaariainen L., Gomatos P. J. A kinetic analysis of the synthesis in BHK 21 cells of RNAs specific for Semliki Forest virus. J Gen Virol. 1969 Sep;5(2):251–265. doi: 10.1099/0022-1317-5-2-251. [DOI] [PubMed] [Google Scholar]
  22. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  23. LUFT J. H. Improvements in epoxy resin embedding methods. J Biophys Biochem Cytol. 1961 Feb;9:409–414. doi: 10.1083/jcb.9.2.409. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Levin J. G., Friedman R. M. Analysis of arbovirus ribonucleic acid forms by polyacrylamide gel electrophoresis. J Virol. 1971 Apr;7(4):504–514. doi: 10.1128/jvi.7.4.504-514.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Martin E. M., Sonnabend J. A. Ribonucleic acid polymerase catalyzing synthesis of double-stranded arbovirus ribonucleic acid. J Virol. 1967 Feb;1(1):97–109. doi: 10.1128/jvi.1.1.97-109.1967. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. 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]
  27. Solymosy F., Fedorcsák I., Gulyás A., Farkas G. L., Ehrenberg L. A new method based on the use of diethyl pyrocarbonate as a nuclease inhibitor for the extraction of undegraded nucleic acid from plant tissues. Eur J Biochem. 1968 Sep 24;5(4):520–527. doi: 10.1111/j.1432-1033.1968.tb00401.x. [DOI] [PubMed] [Google Scholar]
  28. Sonnabend J. A., Martin E. M., Mécs E. Viral specific RNAs in infected cells. Nature. 1967 Jan 28;213(5074):365–367. doi: 10.1038/213365a0. [DOI] [PubMed] [Google Scholar]
  29. Sreevalsan T., Allen P. T. Replication of Western equine encephalomyelitis virus. II. Cytoplasmic structure involved in the synthesis and development of the virions. J Virol. 1968 Oct;2(10):1038–1046. doi: 10.1128/jvi.2.10.1038-1046.1968. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Sreevalsan T. Association of viral ribonucleic acid with cellular membranes in chick embryo cells infected with Sindbis virus. J Virol. 1970 Oct;6(4):438–444. doi: 10.1128/jvi.6.4.438-444.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Sreevalsan T., Lockart R. Z., Jr Heterogeneous RNA's occurring during the replication of Western equine encephalomyelitis virus. Proc Natl Acad Sci U S A. 1966 Apr;55(4):974–981. doi: 10.1073/pnas.55.4.974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Sreevalsan T., Yin F. H. Sindbis virus-induced viral ribonucleic acid polymerase. J Virol. 1969 Jun;3(6):599–604. doi: 10.1128/jvi.3.6.599-604.1969. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Strauss J. H., Jr, Burge B. W., Pfefferkorn E. R., Darnell J. E., Jr Identification of the membrane protein and "core" protein of Sindbis virus. Proc Natl Acad Sci U S A. 1968 Feb;59(2):533–537. doi: 10.1073/pnas.59.2.533. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. VENABLE J. H., COGGESHALL R. A SIMPLIFIED LEAD CITRATE STAIN FOR USE IN ELECTRON MICROSCOPY. J Cell Biol. 1965 May;25:407–408. doi: 10.1083/jcb.25.2.407. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Zhdanov V. M., Yershov F. I., Uryvayev L. V., Ivanovsky D. I. Formation of infectious arbovirus ribonucleoprotein in subcellular structures. Nature. 1970 Dec 19;228(5277):1192–1194. doi: 10.1038/2281192a0. [DOI] [PubMed] [Google Scholar]

Articles from Journal of Virology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES