1887

Abstract

Rubella virus (RUBV) contains a plus-strand RNA genome with two ORFs, one encoding the non-structural replicase proteins (NS-ORF) and the second encoding the virion structural proteins (SP-ORF). This study describes development and use of a -encapsidation system for the assembly of infectious RUBV-like replicon particles (VRPs) containing RUBV replicons (self replicating genomes with the SP-ORF replaced with a reporter gene). First, this system was used to map signals within the RUBV genome that mediate packaging of viral RNA. Mutations within a proposed packaging signal did not significantly affect relative packaging efficiency. The insertion of various fragments derived from the RUBV genome into Sindbis virus replicons revealed that there are several regions within the RUBV genome capable of enhancing encapsidation of heterologous replicon RNAs. Secondly, the -encapsidation system was used to analyse the effect of alterations within the capsid protein (CP) on release of VRPs and subsequent initiation of replication in newly infected cells. Deletion of the N-terminal eight amino acids of the CP reduced VRP titre significantly, which could be partially complemented by native CP provided , indicating that this mutation affected an entry or post-entry event in the replication cycle. To test this hypothesis, the -encapsidation system was used to demonstrate the rescue of a lethal deletion within P150, one of the virus replicase proteins, by CP contained within the virus particle. This novel finding substantiated the functional role of CP in early post-entry replication.

Loading

Article metrics loading...

/content/journal/jgv/10.1099/vir.0.038984-0
2012-03-01
2021-10-17
Loading full text...

Full text loading...

/deliver/fulltext/jgv/93/3/516.html?itemId=/content/journal/jgv/10.1099/vir.0.038984-0&mimeType=html&fmt=ahah

References

  1. Agapov E. V., Frolov I., Lindenbach B. D., Prágai B. M., Schlesinger S., Rice C. M. 1998; Noncytopathic Sindbis virus RNA vectors for heterologous gene expression. Proc Natl Acad Sci U S A 95:12989–12994 [View Article][PubMed]
    [Google Scholar]
  2. Atkins G. J., Fleeton M. N., Sheahan B. J. 2008; Therapeutic and prophylactic applications of alphavirus vectors. Expert Rev Mol Med 10:e33 [View Article][PubMed]
    [Google Scholar]
  3. Bartenschlager R., Junker-Niepmann M., Schaller H. 1990; The P gene product of hepatitis B virus is required as a structural component for genomic RNA encapsidation. J Virol 64:5324–5332[PubMed]
    [Google Scholar]
  4. Beatch M. D., Hobman T. C. 2000; Rubella virus capsid associates with host cell protein p32 and localizes to mitochondria. J Virol 74:5569–5576 [View Article][PubMed]
    [Google Scholar]
  5. Beatch M. D., Everitt J. C., Law L. J., Hobman T. C. 2005; Interactions between rubella virus capsid and host protein p32 are important for virus replication. J Virol 79:10807–10820 [View Article][PubMed]
    [Google Scholar]
  6. Bol J. F. 1999; Alfalfa mosaic virus and ilarviruses: involvement of coat protein in multiple steps of the replication cycle. J Gen Virol 80:1089–1102[PubMed]
    [Google Scholar]
  7. Chen Y., Robinson W. S., Marion P. L. 1992; Naturally occurring point mutation in the C terminus of the polymerase gene prevents duck hepatitis B virus RNA packaging. J Virol 66:1282–1287[PubMed]
    [Google Scholar]
  8. Clever J. L., Miranda D. Jr, Parslow T. G. 2002; RNA structure and packaging signals in the 5′ leader region of the human immunodeficiency virus type 1 genome. J Virol 76:12381–12387 [View Article][PubMed]
    [Google Scholar]
  9. Cristofari G., Ivanyi-Nagy R., Gabus C., Boulant S., Lavergne J. P., Penin F., Darlix J. L. 2004; The hepatitis C virus Core protein is a potent nucleic acid chaperone that directs dimerization of the viral (+) strand RNA in vitro. Nucleic Acids Res 32:2623–2631 [View Article][PubMed]
    [Google Scholar]
  10. Cruceanu M., Urbaneja M. A., Hixson C. V., Johnson D. G., Datta S. A., Fivash M. J., Stephen A. G., Fisher R. J., Gorelick R. J. other authors 2006; Nucleic acid binding and chaperone properties of HIV-1 Gag and nucleocapsid proteins. Nucleic Acids Res 34:593–605 [View Article][PubMed]
    [Google Scholar]
  11. Dalton K., Casais R., Shaw K., Stirrups K., Evans S., Britton P., Brown T. D., Cavanagh D. 2001; cis-Acting sequences required for coronavirus infectious bronchitis virus defective-RNA replication and packaging. J Virol 75:125–133 [View Article][PubMed]
    [Google Scholar]
  12. Derdeyn C. A., Frey T. K. 1995; Characterization of defective-interfering RNAs of rubella virus generated during serial undiluted passage. Virology 206:216–226 [View Article][PubMed]
    [Google Scholar]
  13. Frey T. K. 1994; Molecular biology of rubella virus. Adv Virus Res 44:69–160 [View Article][PubMed]
    [Google Scholar]
  14. Frey T. K., Hemphill M. L. 1988; Generation of defective-interfering particles by rubella virus in Vero cells. Virology 164:22–29 [View Article][PubMed]
    [Google Scholar]
  15. Frolova E., Frolov I., Schlesinger S. 1997; Packaging signals in alphaviruses. J Virol 71:248–258[PubMed]
    [Google Scholar]
  16. Garbutt M., Chan H., Hobman T. C. 1999a; Secretion of rubella virions and virus-like particles in cultured epithelial cells. Virology 261:340–346 [View Article][PubMed]
    [Google Scholar]
  17. Garbutt M., Law L. M., Chan H., Hobman T. C. 1999b; Role of rubella virus glycoprotein domains in assembly of virus-like particles. J Virol 73:3524–3533[PubMed]
    [Google Scholar]
  18. Giessauf A., Flaim M., Walder G., Dierich M. P., Würzner R. 2005; Preparation of immunoblot test stripes from a rubella virus-like particles dye crystal complex as antigen. Arch Virol 150:2077–2090 [View Article][PubMed]
    [Google Scholar]
  19. Grangeot-Keros L., Enders G. 1997; Evaluation of a new enzyme immunoassay based on recombinant rubella virus-like particles for detection of immunoglobulin M antibodies to rubella virus. J Clin Microbiol 35:398–401[PubMed]
    [Google Scholar]
  20. Guogas L. M., Laforest S. M., Gehrke L. 2005; Coat protein activation of alfalfa mosaic virus replication is concentration dependent. J Virol 79:5752–5761 [View Article][PubMed]
    [Google Scholar]
  21. Hobman T. C., Lundstrom M. L., Mauracher C. A., Woodward L., Gillam S., Farquhar M. G. 1994; Assembly of rubella virus structural proteins into virus-like particles in transfected cells. Virology 202:574–585 [View Article][PubMed]
    [Google Scholar]
  22. Ilkow C. S., Weckbecker D., Cho W. J., Meier S., Beatch M. D., Goping I. S., Herrmann J. M., Hobman T. C. 2010; The rubella virus capsid protein inhibits mitochondrial import. J Virol 84:119–130 [View Article][PubMed]
    [Google Scholar]
  23. Ilkow C. S., Goping I. S., Hobman T. C. 2011; The rubella virus capsid is an anti-apoptotic protein that attenuates the pore-forming ability of Bax. PLoS Pathog 7:e1001291 [View Article][PubMed]
    [Google Scholar]
  24. Kim D. Y., Firth A. E., Atasheva S., Frolova E. I., Frolov I. 2011; Conservation of a packaging signal and the viral genome RNA packaging mechanism in alphavirus evolution. J Virol 85:8022–8036 [View Article][PubMed]
    [Google Scholar]
  25. Law L. M., Everitt J. C., Beatch M. D., Holmes C. F., Hobman T. C. 2003; Phosphorylation of rubella virus capsid regulates its RNA binding activity and virus replication. J Virol 77:1764–1771 [View Article][PubMed]
    [Google Scholar]
  26. Law L. J., Ilkow C. S., Tzeng W. P., Rawluk M., Stuart D. T., Frey T. K., Hobman T. C. 2006; Analyses of phosphorylation events in the rubella virus capsid protein: role in early replication events. J Virol 80:6917–6925 [View Article][PubMed]
    [Google Scholar]
  27. Lee J. Y., Hwang D., Gillam S. 1996; Dimerization of rubella virus capsid protein is not required for virus particle formation. Virology 216:223–227 [View Article][PubMed]
    [Google Scholar]
  28. Liang Y., Gillam S. 2001; Rubella virus RNA replication is cis-preferential and synthesis of negative- and positive-strand RNAs is regulated by the processing of nonstructural protein. Virology 282:307–319 [View Article][PubMed]
    [Google Scholar]
  29. Liu Z., Yang D., Qiu Z., Lim K. T., Chong P., Gillam S. 1996; Identification of domains in rubella virus genomic RNA and capsid protein necessary for specific interaction. J Virol 70:2184–2190[PubMed]
    [Google Scholar]
  30. Lundstrom K. 2002; Alphavirus vectors as tools in cancer gene therapy. Technol Cancer Res Treat 1:83–88[PubMed] [CrossRef]
    [Google Scholar]
  31. Magliano D., Marshall J. A., Bowden D. S., Vardaxis N., Meanger J., Lee J. Y. 1998; Rubella virus replication complexes are virus-modified lysosomes. Virology 240:57–63 [View Article][PubMed]
    [Google Scholar]
  32. Mir M. A., Panganiban A. T. 2006; The bunyavirus nucleocapsid protein is an RNA chaperone: possible roles in viral RNA panhandle formation and genome replication. RNA 12:272–282 [View Article][PubMed]
    [Google Scholar]
  33. Nugent C. I., Johnson K. L., Sarnow P., Kirkegaard K. 1999; Functional coupling between replication and packaging of poliovirus replicon RNA. J Virol 73:427–435[PubMed]
    [Google Scholar]
  34. Pappas C. L., Tzeng W. P., Frey T. K. 2006; Evaluation of cis-acting elements in the rubella virus subgenomic RNA that play a role in its translation. Arch Virol 151:327–346 [View Article][PubMed]
    [Google Scholar]
  35. Pollack J. R., Ganem D. 1994; Site-specific RNA binding by a hepatitis B virus reverse transcriptase initiates two distinct reactions: RNA packaging and DNA synthesis. J Virol 68:5579–5587[PubMed]
    [Google Scholar]
  36. Pugachev K. V., Galinski M. S., Frey T. K. 2000; Infectious cDNA clone of the RA27/3 vaccine strain of rubella virus. Virology 273:189–197 [View Article][PubMed]
    [Google Scholar]
  37. Pustowoit B., Grangeot-Keros L., Hobman T. C., Hofmann J. 1996; Evaluation of recombinant rubella-like particles in a commercial immunoassay for the detection of anti-rubella IgG. Clin Diagn Virol 5:13–20 [View Article][PubMed]
    [Google Scholar]
  38. Qiu Z., Ou D., Hobman T. C., Gillam S. 1994; Expression and characterization of virus-like particles containing rubella virus structural proteins. J Virol 68:4086–4091[PubMed]
    [Google Scholar]
  39. Risco C., Carrascosa J. L., Frey T. K. 2003; Structural maturation of rubella virus in the Golgi complex. Virology 312:261–269 [View Article][PubMed]
    [Google Scholar]
  40. Schlesinger S. 2001; Alphavirus vectors: development and potential therapeutic applications. Expert Opin Biol Ther 1:177–191 [View Article][PubMed]
    [Google Scholar]
  41. Schrauf S., Mandl C. W., Bell-Sakyi L., Skern T. 2009; Extension of flavivirus protein C differentially affects early RNA synthesis and growth in mammalian and arthropod host cells. J Virol 83:11201–11210 [View Article][PubMed]
    [Google Scholar]
  42. Swanson M. M., Ansel-McKinney P., Houser-Scott F., Yusibov V., Loesch-Fries L. S., Gehrke L. 1998; Viral coat protein peptides with limited sequence homology bind similar domains of alfalfa mosaic virus and tobacco streak virus RNAs. J Virol 72:3227–3234[PubMed]
    [Google Scholar]
  43. Tzeng W. P., Frey T. K. 2003; Complementation of a deletion in the rubella virus p150 nonstructural protein by the viral capsid protein. J Virol 77:9502–9510 [View Article][PubMed]
    [Google Scholar]
  44. Tzeng W. P., Frey T. K. 2005; Rubella virus capsid protein modulation of viral genomic and subgenomic RNA synthesis. Virology 337:327–334 [View Article][PubMed]
    [Google Scholar]
  45. Tzeng W. P., Frey T. K. 2006; C-E1 fusion protein synthesized by rubella virus DI RNAs maintained during serial passage. Virology 356:198–207 [View Article][PubMed]
    [Google Scholar]
  46. Tzeng W. P., Frey T. K. 2009; Functional replacement of a domain in the rubella virus p150 replicase protein by the virus capsid protein. J Virol 83:3549–3555 [View Article][PubMed]
    [Google Scholar]
  47. Tzeng W. P., Chen M. H., Derdeyn C. A., Frey T. K. 2001; Rubella virus DI RNAs and replicons: requirement for nonstructural proteins acting in cis for amplification by helper virus. Virology 289:63–73 [View Article][PubMed]
    [Google Scholar]
  48. Tzeng W. P., Matthews J. D., Frey T. K. 2006; Analysis of rubella virus capsid protein-mediated enhancement of replicon replication and mutant rescue. J Virol 80:3966–3974 [View Article][PubMed]
    [Google Scholar]
  49. Yao J., Gillam S. 2000; A single-amino-acid substitution of a tyrosine residue in the rubella virus E1 cytoplasmic domain blocks virus release. J Virol 74:3029–3036 [View Article][PubMed]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jgv/10.1099/vir.0.038984-0
Loading
/content/journal/jgv/10.1099/vir.0.038984-0
Loading

Data & Media loading...

Most cited this month Most Cited RSS feed

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error