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J Virol. May 1991; 65(5): 2170–2178.
PMCID: PMC240564

Assembly and transcription of synthetic vesicular stomatitis virus nucleocapsids.

Abstract

The functional template for transcription of vesicular stomatitis virus (VSV) RNA is a ribonucleoprotein particle (nucleocapsid) consisting of the negative-strand sense genomic RNA completely encapsidated by the viral nucleocapsid (N) protein. As an approach to create nucleocapsids in vitro, we demonstrate here the specific encapsidation by purified N protein of in vitro-synthesized RNA sequences representing the 5' end of both the negative- and positive-strand VSV genome-length RNAs. As few as 19 nucleotides from the 5'-end of positive-strand RNA allowed maximal encapsidation, although the 5' terminal 10 nucleotides would allow partial (50%) encapsidation. Sequences downstream of the binding site can be of any origin. Specific encapsidation of VSV sequences was dependent on the presence of uninfected cell cytoplasmic extracts or poly(A). The synthetic nucleocapsids have the properties of RNase resistance and a buoyant density typical of wild-type VSV nucleocapsids. We have encapsidated a synthetic virionlike RNA species which contained just the terminal sequences of the virion RNA: the N encapsidation signal from the 5' end and the leader gene from the 3' end. This assembled nucleocapsid could function in vitro as a transcription template for the VSV RNA polymerase.

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  • Adam SA, Choi YD, Dreyfuss G. Interaction of mRNA with proteins in vesicular stomatitis virus-infected cells. J Virol. 1986 Feb;57(2):614–622. [PMC free article] [PubMed]
  • Arnheiter H, Davis NL, Wertz G, Schubert M, Lazzarini RA. Role of the nucleocapsid protein in regulating vesicular stomatitis virus RNA synthesis. Cell. 1985 May;41(1):259–267. [PubMed]
  • Ballart I, Eschle D, Cattaneo R, Schmid A, Metzler M, Chan J, Pifko-Hirst S, Udem SA, Billeter MA. Infectious measles virus from cloned cDNA. EMBO J. 1990 Feb;9(2):379–384. [PMC free article] [PubMed]
  • Banerjee AK. Transcription and replication of rhabdoviruses. Microbiol Rev. 1987 Mar;51(1):66–87. [PMC free article] [PubMed]
  • Blumberg BM, Giorgi C, Kolakofsky D. N protein of vesicular stomatitis virus selectively encapsidates leader RNA in vitro. Cell. 1983 Feb;32(2):559–567. [PubMed]
  • Blumberg BM, Giorgi C, Rose K, Kolakofsky D. Preparation and analysis of the nucleocapsid proteins of vesicular stomatitis virus and sendai virus, and analysis of the sendai virus leader-NP gene region. J Gen Virol. 1984 Apr;65(Pt 4):769–779. [PubMed]
  • Blumberg BM, Kolakofsky D. Intracellular vesicular stomatitis virus leader RNAs are found in nucleocapsid structures. J Virol. 1981 Nov;40(2):568–576. [PMC free article] [PubMed]
  • Blumberg BM, Leppert M, Kolakofsky D. Interaction of VSV leader RNA and nucleocapsid protein may control VSV genome replication. Cell. 1981 Mar;23(3):837–845. [PubMed]
  • Carlsen SR, Peluso RW, Moyer SA. In vitro replication of Sendai virus wild-type and defective interfering particle genome RNAs. J Virol. 1985 May;54(2):493–500. [PMC free article] [PubMed]
  • Chattopadhyay D, Banerjee AK. Phosphorylation within a specific domain of the phosphoprotein of vesicular stomatitis virus regulates transcription in vitro. Cell. 1987 May 8;49(3):407–414. [PubMed]
  • Crowley JC, Dowling PC, Menonna J, Silverman JI, Schuback D, Cook SD, Blumberg BM. Sequence variability and function of measles virus 3' and 5' ends and intercistronic regions. Virology. 1988 Jun;164(2):498–506. [PubMed]
  • Davis NL, Arnheiter H, Wertz GW. Vesicular stomatitis virus N and NS proteins form multiple complexes. J Virol. 1986 Sep;59(3):751–754. [PMC free article] [PubMed]
  • Emerson SU. Reconstitution studies detect a single polymerase entry site on the vesicular stomatitis virus genome. Cell. 1982 Dec;31(3 Pt 2):635–642. [PubMed]
  • Emerson SU, Wagner RR. Dissociation and reconstitution of the transcriptase and template activities of vesicular stomatitis B and T virions. J Virol. 1972 Aug;10(2):297–309. [PMC free article] [PubMed]
  • Gill DS, Chattopadhyay D, Banerjee AK. Identification of a domain within the phosphoprotein of vesicular stomatitis virus that is essential for transcription in vitro. Proc Natl Acad Sci U S A. 1986 Dec;83(23):8873–8877. [PMC free article] [PubMed]
  • Giorgi C, Blumberg B, Kolakofsky D. Sequence determination of the (+) leader RNA regions of the vesicular stomatitis virus Chandipura, Cocal, and Piry serotype genomes. J Virol. 1983 Apr;46(1):125–130. [PMC free article] [PubMed]
  • Grubman MJ, Shafritz DA. Identification and characterization of messenger ribonucleoprotein complexes from vesicular stomatitis virus-infected HeLa cells. Virology. 1977 Aug;81(1):1–16. [PubMed]
  • Luytjes W, Krystal M, Enami M, Parvin JD, Palese P. Amplification, expression, and packaging of foreign gene by influenza virus. Cell. 1989 Dec 22;59(6):1107–1113. [PubMed]
  • Masters PS, Banerjee AK. Resolution of multiple complexes of phosphoprotein NS with nucleocapsid protein N of vesicular stomatitis virus. J Virol. 1988 Aug;62(8):2651–2657. [PMC free article] [PubMed]
  • Masters PS, Banerjee AK. Complex formation with vesicular stomatitis virus phosphoprotein NS prevents binding of nucleocapsid protein N to nonspecific RNA. J Virol. 1988 Aug;62(8):2658–2664. [PMC free article] [PubMed]
  • Milligan JF, Groebe DR, Witherell GW, Uhlenbeck OC. Oligoribonucleotide synthesis using T7 RNA polymerase and synthetic DNA templates. Nucleic Acids Res. 1987 Nov 11;15(21):8783–8798. [PMC free article] [PubMed]
  • Mirakhur B, Peluso RW. In vitro assembly of a functional nucleocapsid from the negative-stranded genome RNA of a defective interfering particle of vesicular stomatitis virus. Proc Natl Acad Sci U S A. 1988 Oct;85(20):7511–7515. [PMC free article] [PubMed]
  • Moyer SA. Replication of the genome RNAs of defective interfering particles of vesicular stomatitis and Sendai viruses using heterologous viral proteins. Virology. 1989 Sep;172(1):341–345. [PubMed]
  • Moyer SA, Baker SC, Lessard JL. Tubulin: a factor necessary for the synthesis of both Sendai virus and vesicular stomatitis virus RNAs. Proc Natl Acad Sci U S A. 1986 Aug;83(15):5405–5409. [PMC free article] [PubMed]
  • Moyer SA, Gatchell SH. Intracellular events in the replication of defective interfering particles of vesicular stomatitis virus. Virology. 1979 Jan 15;92(1):168–179. [PubMed]
  • Nichol ST, Holland JJ. Genome RNA terminus conservation and diversity among vesiculoviruses. J Virol. 1987 Jan;61(1):200–205. [PMC free article] [PubMed]
  • Peluso RW. Kinetic, quantitative, and functional analysis of multiple forms of the vesicular stomatitis virus nucleocapsid protein in infected cells. J Virol. 1988 Aug;62(8):2799–2807. [PMC free article] [PubMed]
  • Peluso RW, Moyer SA. Initiation and replication of vesicular stomatitis virus genome RNA in a cell-free system. Proc Natl Acad Sci U S A. 1983 Jun;80(11):3198–3202. [PMC free article] [PubMed]
  • Peluso RW, Moyer SA. Viral proteins required for the in vitro replication of vesicular stomatitis virus defective interfering particle genome RNA. Virology. 1988 Feb;162(2):369–376. [PubMed]
  • Rosen CA, Ennis HL, Cohen PS. Translational control of vesicular stomatitis virus protein synthesis: isolation of an mRNA-sequestering particle. J Virol. 1982 Dec;44(3):932–938. [PMC free article] [PubMed]
  • Sprague J, Condra JH, Arnheiter H, Lazzarini RA. Expression of a recombinant DNA gene coding for the vesicular stomatitis virus nucleocapsid protein. J Virol. 1983 Feb;45(2):773–781. [PMC free article] [PubMed]
  • Thomas D, Newcomb WW, Brown JC, Wall JS, Hainfeld JF, Trus BL, Steven AC. Mass and molecular composition of vesicular stomatitis virus: a scanning transmission electron microscopy analysis. J Virol. 1985 May;54(2):598–607. [PMC free article] [PubMed]
  • Zoller MJ, Smith M. Oligonucleotide-directed mutagenesis: a simple method using two oligonucleotide primers and a single-stranded DNA template. DNA. 1984 Dec;3(6):479–488. [PubMed]

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