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J Virol. Mar 1996; 70(3): 1981–1989.
PMCID: PMC190027

Deduced consensus sequence of Sindbis virus strain AR339: mutations contained in laboratory strains which affect cell culture and in vivo phenotypes.

Abstract

The consensus sequence of the Sindbis virus AR339 isolate, the prototype alphavirus, has been deduced. THe results presented here suggest (i) that a substantial proportion of the sequence divergence evident between the consensus sequence and sequences of laboratory strains of AR339 has resulted from selection for efficient growth in cell culture, (ii) that many of these changes affect the virulence of the virus in animal models, and (iii) that such modified genetic backgrounds present in laboratory strains can exert a significant influence on genetic studies of virus pathogenesis and host range. A laboratory strain of Sindbis virus AR339 was sequenced and cloned as a cDNA (pTRSB) from which infectious virus (TRSB) could be derived. The consensus sequence was deduced from the complete sequences of pTRSB and HRsp (E. G. Strauss, C. M. Rice, and J. H. Strauss, Virology 133:92-110, 1984), from partial sequences of the glycoprotein genes of three other AR339 laboratory strains, and by comparison with the sequences of the glycoprotein genes of three other AR339 sequence. HRsp differed form the consensus sequence by eight coding changes, and TRSB differed by three coding changes. In the 5' untranslated region, HRsp differed from the consensus sequence at nucleotide (nt) 5. These differences were likely the result of cell culture passage of the original AR339 isolate. At three of the difference loci (one in TRSB and two in HRsp), selection of cell-culture-adaptive mutations was documented with Sindbis virus or other alphaviruses. Selection in cell culture often results in attenuation of virulence in animals. Considering the TRSB and HRsp sequences together, one noncoding difference from the consensus (an A-for-G substitution in the 5' untranslated region at nt 5) and six coding differences in the glycoprotein genes (at E2 amino acids 1, 3, 70, and 172 and at E1 amino acids 72 and 237) were at loci which, either individually or in combination, significantly affected alphavirus virulence in mice. Although the levels of virulence of isogenic strains containing either nt 5 A or nt 5 G did not differ significantly in neonatal mice, the presence of nt 5 A greatly enhanced the effect of a second attenuating mutation in the E2 gene. These results suggest that minimal differences in the "wild type" genetic background into which an additional mutation is introduced can have a dramatic effect on apparent virulence and pathogenesis phenotypes. A cDNA clone of the consensus AR339 sequence, a sequence devoid of occult attenuating mutations introduced by cell culture passage, will allow the molecular genetic examination of cell culture and in vivo phenotypes of a virus which may best reflect the sequence of Sindbis virus AR339 at the time of its isolation.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Ahlquist P, French R, Janda M, Loesch-Fries LS. Multicomponent RNA plant virus infection derived from cloned viral cDNA. Proc Natl Acad Sci U S A. 1984 Nov;81(22):7066–7070. [PMC free article] [PubMed]
  • Baric RS, Trent DW, Johnston RE. A Sindbis virus variant with a cell-determined latent period. Virology. 1981 Apr 15;110(1):237–242. [PubMed]
  • Boggs WM, Hahn CS, Strauss EG, Strauss JH, Griffin DE. Low pH-dependent Sindbis virus-induced fusion of BHK cells: differences between strains correlate with amino acid changes in the E1 glycoprotein. Virology. 1989 Apr;169(2):485–488. [PubMed]
  • Burge BW, Pfefferkorn ER. Isolation and characterization of conditional-lethal mutants of Sindbis virus. Virology. 1966 Oct;30(2):204–213. [PubMed]
  • Davis NL, Fuller FJ, Dougherty WG, Olmsted RA, Johnston RE. A single nucleotide change in the E2 glycoprotein gene of Sindbis virus affects penetration rate in cell culture and virulence in neonatal mice. Proc Natl Acad Sci U S A. 1986 Sep;83(18):6771–6775. [PMC free article] [PubMed]
  • Davis NL, Pence DF, Meyer WJ, Schmaljohn AL, Johnston RE. Alternative forms of a strain-specific neutralizing antigenic site on the Sindbis virus E2 glycoprotein. Virology. 1987 Nov;161(1):101–108. [PubMed]
  • Davis NL, Powell N, Greenwald GF, Willis LV, Johnson BJ, Smith JF, Johnston RE. Attenuating mutations in the E2 glycoprotein gene of Venezuelan equine encephalitis virus: construction of single and multiple mutants in a full-length cDNA clone. Virology. 1991 Jul;183(1):20–31. [PubMed]
  • Davis NL, Willis LV, Smith JF, Johnston RE. In vitro synthesis of infectious venezuelan equine encephalitis virus RNA from a cDNA clone: analysis of a viable deletion mutant. Virology. 1989 Jul;171(1):189–204. [PubMed]
  • Evans DM, Dunn G, Minor PD, Schild GC, Cann AJ, Stanway G, Almond JW, Currey K, Maizel JV., Jr Increased neurovirulence associated with a single nucleotide change in a noncoding region of the Sabin type 3 poliovaccine genome. Nature. 1985 Apr 11;314(6011):548–550. [PubMed]
  • Griffin DE, Johnson RT. Role of the immune response in recovery from Sindbis virus encephalitis in mice. J Immunol. 1977 Mar;118(3):1070–1075. [PubMed]
  • Hahn CS, Lustig S, Strauss EG, Strauss JH. Western equine encephalitis virus is a recombinant virus. Proc Natl Acad Sci U S A. 1988 Aug;85(16):5997–6001. [PMC free article] [PubMed]
  • Heidner HW, Johnston RE. The amino-terminal residue of Sindbis virus glycoprotein E2 influences virus maturation, specific infectivity for BHK cells, and virulence in mice. J Virol. 1994 Dec;68(12):8064–8070. [PMC free article] [PubMed]
  • Heidner HW, McKnight KL, Davis NL, Johnston RE. Lethality of PE2 incorporation into Sindbis virus can be suppressed by second-site mutations in E3 and E2. J Virol. 1994 Apr;68(4):2683–2692. [PMC free article] [PubMed]
  • Holland J, Spindler K, Horodyski F, Grabau E, Nichol S, VandePol S. Rapid evolution of RNA genomes. Science. 1982 Mar 26;215(4540):1577–1585. [PubMed]
  • Janssen R, Gonzalez-Scarano F, Nathanson N. Mechanisms of bunyavirus virulence. Comparative pathogenesis of a virulent strain of La Crosse and an avirulent strain of Tahyna virus. Lab Invest. 1984 Apr;50(4):447–455. [PubMed]
  • Janssen RS, Nathanson N, Endres MJ, Gonzalez-Scarano F. Virulence of La Crosse virus is under polygenic control. J Virol. 1986 Jul;59(1):1–7. [PMC free article] [PubMed]
  • Johnston RE, Smith JF. Selection for accelerated penetration in cell culture coselects for attenuated mutants of Venezuelan equine encephalitis virus. Virology. 1988 Feb;162(2):437–443. [PubMed]
  • Kaye KM, Spriggs DR, Bassel-Duby R, Fields BN, Tyler KL. Genetic basis for altered pathogenesis of an immune-selected antigenic variant of reovirus type 3 (Dearing). J Virol. 1986 Jul;59(1):90–97. [PMC free article] [PubMed]
  • Kinney RM, Chang GJ, Tsuchiya KR, Sneider JM, Roehrig JT, Woodward TM, Trent DW. Attenuation of Venezuelan equine encephalitis virus strain TC-83 is encoded by the 5'-noncoding region and the E2 envelope glycoprotein. J Virol. 1993 Mar;67(3):1269–1277. [PMC free article] [PubMed]
  • Kuhn RJ, Griffin DE, Zhang H, Niesters HG, Strauss JH. Attenuation of Sindbis virus neurovirulence by using defined mutations in nontranslated regions of the genome RNA. J Virol. 1992 Dec;66(12):7121–7127. [PMC free article] [PubMed]
  • Kuhn RJ, Niesters HG, Hong Z, Strauss JH. Infectious RNA transcripts from Ross River virus cDNA clones and the construction and characterization of defined chimeras with Sindbis virus. Virology. 1991 Jun;182(2):430–441. [PubMed]
  • Kunkel TA, Roberts JD, Zakour RA. Rapid and efficient site-specific mutagenesis without phenotypic selection. Methods Enzymol. 1987;154:367–382. [PubMed]
  • Liljeström P, Lusa S, Huylebroeck D, Garoff H. In vitro mutagenesis of a full-length cDNA clone of Semliki Forest virus: the small 6,000-molecular-weight membrane protein modulates virus release. J Virol. 1991 Aug;65(8):4107–4113. [PMC free article] [PubMed]
  • Lustig S, Jackson AC, Hahn CS, Griffin DE, Strauss EG, Strauss JH. Molecular basis of Sindbis virus neurovirulence in mice. J Virol. 1988 Jul;62(7):2329–2336. [PMC free article] [PubMed]
  • Meyer WJ, Johnston RE. Structural rearrangement of infecting Sindbis virions at the cell surface: mapping of newly accessible epitopes. J Virol. 1993 Sep;67(9):5117–5125. [PMC free article] [PubMed]
  • Olmsted RA, Baric RS, Sawyer BA, Johnston RE. Sindbis virus mutants selected for rapid growth in cell culture display attenuated virulence in animals. Science. 1984 Jul 27;225(4660):424–427. [PubMed]
  • Olmsted RA, Meyer WJ, Johnston RE. Characterization of Sindbis virus epitopes important for penetration in cell culture and pathogenesis in animals. Virology. 1986 Jan 30;148(2):245–254. [PubMed]
  • Paredes AM, Brown DT, Rothnagel R, Chiu W, Schoepp RJ, Johnston RE, Prasad BV. Three-dimensional structure of a membrane-containing virus. Proc Natl Acad Sci U S A. 1993 Oct 1;90(19):9095–9099. [PMC free article] [PubMed]
  • Pence DF, Davis NL, Johnston RE. Antigenic and genetic characterization of Sindbis virus monoclonal antibody escape mutants which define a pathogenesis domain on glycoprotein E2. Virology. 1990 Mar;175(1):41–49. [PubMed]
  • PFEFFERKORN ER, HUNTER HS. PURIFICATION AND PARTIAL CHEMICAL ANALYSIS OF SINDBIS VIRUS. Virology. 1963 Jul;20:433–445. [PubMed]
  • Polo JM, Davis NL, Rice CM, Huang HV, Johnston RE. Molecular analysis of Sindbis virus pathogenesis in neonatal mice by using virus recombinants constructed in vitro. J Virol. 1988 Jun;62(6):2124–2133. [PMC free article] [PubMed]
  • Polo JM, Johnston RE. Attenuating mutations in glycoproteins E1 and E2 of Sindbis virus produce a highly attenuated strain when combined in vitro. J Virol. 1990 Sep;64(9):4438–4444. [PMC free article] [PubMed]
  • Polo JM, Johnston RE. Mutational analysis of a virulence locus in the E2 glycoprotein gene of Sindbis virus. J Virol. 1991 Nov;65(11):6358–6361. [PMC free article] [PubMed]
  • Racaniello VR, Baltimore D. Cloned poliovirus complementary DNA is infectious in mammalian cells. Science. 1981 Nov 20;214(4523):916–919. [PubMed]
  • Rezapkin GV, Norwood LP, Taffs RE, Dragunsky EM, Levenbook IS, Chumakov KM. Microevolution of type 3 Sabin strain of poliovirus in cell cultures and its implications for oral poliovirus vaccine quality control. Virology. 1995 Aug 20;211(2):377–384. [PubMed]
  • Rice CM, Levis R, Strauss JH, Huang HV. Production of infectious RNA transcripts from Sindbis virus cDNA clones: mapping of lethal mutations, rescue of a temperature-sensitive marker, and in vitro mutagenesis to generate defined mutants. J Virol. 1987 Dec;61(12):3809–3819. [PMC free article] [PubMed]
  • Rice CM, Strauss JH. Nucleotide sequence of the 26S mRNA of Sindbis virus and deduced sequence of the encoded virus structural proteins. Proc Natl Acad Sci U S A. 1981 Apr;78(4):2062–2066. [PMC free article] [PubMed]
  • Riviere Y, Ahmed R, Southern PJ, Buchmeier MJ, Oldstone MB. Genetic mapping of lymphocytic choriomeningitis virus pathogenicity: virulence in guinea pigs is associated with the L RNA segment. J Virol. 1985 Sep;55(3):704–709. [PMC free article] [PubMed]
  • Riviere Y, Ahmed R, Southern P, Oldstone MB. Perturbation of differentiated functions during viral infection in vivo. II. Viral reassortants map growth hormone defect to the S RNA of the lymphocytic choriomeningitis virus genome. Virology. 1985 Apr 15;142(1):175–182. [PubMed]
  • Rümenapf T, Strauss EG, Strauss JH. Aura virus is a New World representative of Sindbis-like viruses. Virology. 1995 Apr 20;208(2):621–633. [PubMed]
  • Russell DL, Dalrymple JM, Johnston RE. Sindbis virus mutations which coordinately affect glycoprotein processing, penetration, and virulence in mice. J Virol. 1989 Apr;63(4):1619–1629. [PMC free article] [PubMed]
  • SABIN AB, HENNESSEN WA, WINSSER J. Studies on variants of poliomyelitis virus. I. Experimental segregation and properties of avirulent variants of three immunologic types. J Exp Med. 1954 Jun 1;99(6):551–576. [PMC free article] [PubMed]
  • Sanger F, Nicklen S, Coulson AR. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. [PMC free article] [PubMed]
  • Schild GC, Oxford JS, de Jong JC, Webster RG. Evidence for host-cell selection of influenza virus antigenic variants. Nature. 1983 Jun 23;303(5919):706–709. [PubMed]
  • Schoepp RJ, Johnston RE. Directed mutagenesis of a Sindbis virus pathogenesis site. Virology. 1993 Mar;193(1):149–159. [PubMed]
  • Sherman LA, Griffin DE. Pathogenesis of encephalitis induced in newborn mice by virulent and avirulent strains of Sindbis virus. J Virol. 1990 May;64(5):2041–2046. [PMC free article] [PubMed]
  • Shirako Y, Niklasson B, Dalrymple JM, Strauss EG, Strauss JH. Structure of the Ockelbo virus genome and its relationship to other Sindbis viruses. Virology. 1991 Jun;182(2):753–764. [PubMed]
  • Spriggs DR, Bronson RT, Fields BN. Hemagglutinin variants of reovirus type 3 have altered central nervous system tropism. Science. 1983 Apr 29;220(4596):505–507. [PubMed]
  • Strauss EG, Lenches EM, Strauss JH. Mutants of sindbis virus. I. Isolation and partial characterization of 89 new temperature-sensitive mutants. Virology. 1976 Oct 1;74(1):154–168. [PubMed]
  • Strauss EG, Rice CM, Strauss JH. Complete nucleotide sequence of the genomic RNA of Sindbis virus. Virology. 1984 Feb;133(1):92–110. [PubMed]
  • Strauss EG, Stec DS, Schmaljohn AL, Strauss JH. Identification of antigenically important domains in the glycoproteins of Sindbis virus by analysis of antibody escape variants. J Virol. 1991 Sep;65(9):4654–4664. [PMC free article] [PubMed]
  • Strauss JH, Strauss EG. The alphaviruses: gene expression, replication, and evolution. Microbiol Rev. 1994 Sep;58(3):491–562. [PMC free article] [PubMed]
  • TAYLOR RM, HURLBUT HS, WORK TH, KINGSTON JR, FROTHINGHAM TE. Sindbis virus: a newly recognized arthropodtransmitted virus. Am J Trop Med Hyg. 1955 Sep;4(5):844–862. [PubMed]
  • Tucker PC, Griffin DE. Mechanism of altered Sindbis virus neurovirulence associated with a single-amino-acid change in the E2 Glycoprotein. J Virol. 1991 Mar;65(3):1551–1557. [PMC free article] [PubMed]
  • Tucker PC, Strauss EG, Kuhn RJ, Strauss JH, Griffin DE. Viral determinants of age-dependent virulence of Sindbis virus for mice. J Virol. 1993 Aug;67(8):4605–4610. [PMC free article] [PubMed]
  • Ward CD, Flanegan JB. Determination of the poliovirus RNA polymerase error frequency at eight sites in the viral genome. J Virol. 1992 Jun;66(6):3784–3793. [PMC free article] [PubMed]

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