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Genome segment reassortment identifies non-structural protein NS3 as a key protein in African horsesickness virus release and alteration of membrane permeability

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Abstract

The role of African horsesickness virus (AHSV) nonstructural membrane protein NS3 in determining the effects of AHSV infection on Vero cells was examined. NS3 protein sequences are highly variable and cluster into three phylogenetic groups, α, β, and γ. Three AHSV strains, with NS3 from α, β, or γ, were shown to have quantitatively different phenotypes in Vero cells. Reassortants between these strains, in which the S10 genome segment encoding NS3 was exchanged alone or with other segments, were generated and compared to parental strains. Exchange of the NS3 gene resulted in changes in virus release, membrane permeability and total virus yield, indicating an important role for NS3 in these viral properties. Differences in the cytopathicity and the effect on cell viability between the parental strains could not be associated with NS3 alone, and it is likely that a number of viral and host factors play a role.

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References

  1. Balasuriya UB, Nadler SA, Wilson WC, Pritchard LI, Smythe AB, Savini G, Monaco F, De Santis P, Zhang N, Tabachnick WJ, Maclachlan NJ (2008) The NS3 proteins of global strains of bluetongue virus evolve into regional topotypes through negative (purifying) selection. Vet Microbiol 126:91–100

    Article  PubMed  CAS  Google Scholar 

  2. Boyce M, Celma C, Roy P (2008) Development of reverse genetics systems for bluetongue virus: recovery of infectious virus from synthetic RNA transcripts. J Virol 82:8339–8348

    Article  PubMed  CAS  Google Scholar 

  3. Carrasco L (1995) Modification of membrane permeability by animal viruses. Adv Virus Res 45:61–112

    Article  PubMed  CAS  Google Scholar 

  4. Coetzer JAW, Guthrie AJ (2004) African horsesickness. In: Coetzer JAW, Tustin RC (eds) Infectious diseases of livestock. Oxford University Press, Cape Town, pp 1231–1246

    Google Scholar 

  5. Cowley JA, Gorman BM (1987) Genetic reassortants for identification of the genome segment coding for the bluetongue virus hemagglutinin. J Virol 61:2304–2306

    PubMed  CAS  Google Scholar 

  6. DeMaula CD, Jutila MA, Wilson DW, MacLachlan NJ (2001) Infection kinetics, prostacyclin release and cytokine-mediated modulation of the mechanism of cell death during bluetongue virus infection of cultured ovine and bovine pulmonary artery and lung microvascular endothelial cells. J Gen Virol 82:787–794

    PubMed  CAS  Google Scholar 

  7. DeMaula CD, Leutenegger CM, Jutila MA, MacLachlan NJ (2002) Bluetongue virus-induced activation of primary bovine lung microvascular endothelial cells. Vet Immunol Immunopathol 86:147–157

    Article  PubMed  CAS  Google Scholar 

  8. Eaton BT, Hyatt AD, Brookes SM (1990) The replication of bluetongue virus. Curr Top Microbiol Immunol 162:89–118

    PubMed  CAS  Google Scholar 

  9. Gomez-Villamandos JC, Sanchez C, Carrasco L, Laviada MM, Bautista MJ, Martinez-Torrecuadrada J, Sanchez-Vizcaino JM, Sierra MA (1999) Pathogenesis of African horse sickness: ultrastructural study of the capillaries in experimental infection. J Comp Pathol 121:101–116

    Article  PubMed  CAS  Google Scholar 

  10. Gonzalez ME, Carrasco L (2003) Viroporins. FEBS Lett 552:28–34

    Article  PubMed  CAS  Google Scholar 

  11. Han Z, Harty RN (2004) The NS3 protein of bluetongue virus exhibits viroporin-like properties. J Biol Chem 279:43092–43097

    Article  PubMed  CAS  Google Scholar 

  12. Huismans H, Els HJ (1979) Characterization of the tubules associated with the replication of three different orbiviruses. Virology 92:397–406

    Article  PubMed  CAS  Google Scholar 

  13. Laegreid WW, Burrage TG, Stone-Marschat M, Skowronek A (1992) Electron microscopic evidence for endothelial infection by African horsesickness virus. Vet Pathol 29:554–556

    Article  PubMed  CAS  Google Scholar 

  14. Laegreid WW, Stone-Marschat M, Skowronek A, Burrage T (1992) Infection of endothelial cells by African horse sickness viruses. In: Walton TE, Osburn BI (eds) Bluetongue, African Horse Sickness, and Related Orbiviruses. CRC Press, Florida, pp 807–814

    Google Scholar 

  15. Laegreid WW, Skowronek A, Stone-Marschat M, Burrage T (1993) Characterization of virulence variants of African horsesickness virus. Virology 195:836–839

    Article  PubMed  CAS  Google Scholar 

  16. Madan V, Castello A, Carrasco L (2008) Viroporins from RNA viruses induce caspase-dependent apoptosis. Cell Microbiol 10:437–451

    PubMed  CAS  Google Scholar 

  17. Martin LA, Meyer AJ, O’Hara RS, Fu H, Mellor PS, Knowles NJ, Mertens PP (1998) Phylogenetic analysis of African horse sickness virus segment 10: sequence variation, virulence characteristics and cell exit. Arch Virol (Suppl) 14:281–293

    CAS  Google Scholar 

  18. Mellor PS, Hamblin C (2004) African horse sickness. Vet Res 35:445–466

    Article  PubMed  Google Scholar 

  19. Moody MD, Joklik WK (1989) The function of reovirus proteins during the reovirus multiplication cycle: analysis using monoreassortants. Virology 173:437–446

    Article  PubMed  CAS  Google Scholar 

  20. Mortola E, Noad R, Roy P (2004) Bluetongue virus outer capsid proteins are sufficient to trigger apoptosis in mammalian cells. J Virol 78:2875–2883

    Article  PubMed  CAS  Google Scholar 

  21. O’Hara RS, Meyer AJ, Burroughs JN, Pullen L, Martin LA, Mertens PP (1998) Development of a mouse model system, coding assignments and identification of the genome segments controlling virulence of African horse sickness virus serotypes 3 and 8. Arch Virol (Suppl) 14:259–279

    CAS  Google Scholar 

  22. Owens RJ, Limn C, Roy P (2004) Role of an arbovirus nonstructural protein in cellular pathogenesis and virus release. J Virol 78:6649–6656

    Article  PubMed  CAS  Google Scholar 

  23. Pappas C, Aguilar PV, Basler CF, Solorzano A, Zeng H, Perrone LA, Palese P, Garcia-Sastre A, Katz JM, Tumpey TM (2008) Single gene reassortants identify a critical role for PB1, HA, and NA in the high virulence of the 1918 pandemic influenza virus. Proc Natl Acad Sci USA 105:3064–3069

    Article  PubMed  CAS  Google Scholar 

  24. Parks CL, Latham T, Cahill A, O’Neill RE, Passarotti CJ, Buonagurio DA, Bechert TM, D’Arco GA, Neumann G, Destefano J, Arendt HE, Obregon J, Shutyak L, Hamm S, Sidhu MS, Zamb TJ, Udem SA (2007) Phenotypic properties resulting from directed gene segment reassortment between wild-type A/Sydney/5/97 influenza virus and the live attenuated vaccine strain. Virology 367:275–287

    Article  PubMed  CAS  Google Scholar 

  25. Potgieter AC, Steele AD, van Dijk AA (2002) Cloning of complete genome sets of six dsRNA viruses using an improved cloning method for large dsRNA genes. J Gen Virol 83:2215–2223

    PubMed  CAS  Google Scholar 

  26. Quan M, Van Vuuren M, Howell PG, Groenewald D, Guthrie AJ (2008) Molecular epidemiology of the African horse sickness virus S10 gene. J Gen Virol 89:1159–1168

    Article  PubMed  CAS  Google Scholar 

  27. Ramig RF, Garrison C, Chen D, Bell-Robinson D (1989) Analysis of reassortment and superinfection during mixed infection of Vero cells with bluetongue virus serotypes 10 and 17. J Gen Virol 70:2595–2603

    Article  PubMed  Google Scholar 

  28. Roner MR, Mutsoli C (2007) The use of monoreassortants and reverse genetics to map reovirus lysis of a ras-transformed cell line. J Virol Methods 139:132–142

    Article  PubMed  CAS  Google Scholar 

  29. Roy P, Mertens PP, Casal I (1994) African horse sickness virus structure. Comp Immunol Microbiol Infect Dis 17:243–273

    Article  PubMed  CAS  Google Scholar 

  30. Sailleau C, Moulay S, Zientara S (1997) Nucleotide sequence comparison of the segments S10 of the nine African horsesickness virus serotypes. Arch Virol 142:965–978

    Article  PubMed  CAS  Google Scholar 

  31. Skowronek AJ, LaFranco L, Stone-Marschat MA, Burrage TG, Rebar AH, Laegreid WW (1995) Clinical pathology and hemostatic abnormalities in experimental African horsesickness. Vet Pathol 32:112–121

    Article  PubMed  CAS  Google Scholar 

  32. Stoltz MA, Van der Merwe CF, Coetzee J, Huismans H (1996) Subcellular localization of the nonstructural protein NS3 of African horsesickness virus. Onderstepoort J Vet Res 63:57–61

    PubMed  CAS  Google Scholar 

  33. Tauscher GI, Desselberger U (1997) Viral determinants of rotavirus pathogenicity in pigs: production of reassortants by asynchronous coinfection. J Virol 71:853–857

    PubMed  CAS  Google Scholar 

  34. Van Niekerk M, Smit CC, Fick WC, Van Staden V, Huismans H (2001) Membrane association of African horsesickness virus nonstructural protein NS3 determines its cytotoxicity. Virology 279:499–508

    Article  PubMed  CAS  Google Scholar 

  35. Van Niekerk M, Van Staden V, Van Dijk AA, Huismans H (2001) Variation of African horsesickness virus nonstructural protein NS3 in southern Africa. J Gen Virol 82:149–158

    PubMed  Google Scholar 

  36. Van Niekerk M, Freeman M, Paweska JT, Howell PG, Guthrie AJ, Potgieter AC, Van Staden V, Huismans H (2003) Variation in the NS3 gene and protein in South African isolates of bluetongue and equine encephalosis viruses. J Gen Virol 84:581–590

    Article  PubMed  CAS  Google Scholar 

  37. Van Staden V, Stoltz MA, Huismans H (1995) Expression of nonstructural protein NS3 of African horsesickness virus (AHSV): evidence for a cytotoxic effect of NS3 in insect cells, and characterization of the gene products in AHSV infected Vero cells. Arch Virol 140:289–306

    Article  PubMed  Google Scholar 

  38. Wirblich C, Bhattacharya B, Roy P (2006) Nonstructural protein 3 of bluetongue virus assists virus release by recruiting ESCRT-I protein Tsg101. J Virol 80:460–473

    Article  PubMed  CAS  Google Scholar 

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Acknowledgments

The authors gratefully acknowledge Flip Wege for technical assistance, Carel Smit and Eugene Makgopa for their help in generating reassortants and Dr A. C. Potgieter of the Onderstepoort Veterinary Institute for the cDNA amplification of the viral dsRNA genomes. Financial assistance was provided by the National Research Foundation.

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  1. University of Pretoria, Pretoria, South Africa

    Tracy L. Meiring, Henk Huismans & Vida van Staden

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  1. Tracy L. Meiring
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  2. Henk Huismans
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  3. Vida van Staden
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Correspondence to Vida van Staden.

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Meiring, T.L., Huismans, H. & van Staden, V. Genome segment reassortment identifies non-structural protein NS3 as a key protein in African horsesickness virus release and alteration of membrane permeability. Arch Virol 154, 263–271 (2009). https://doi.org/10.1007/s00705-008-0302-8

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  • DOI: https://doi.org/10.1007/s00705-008-0302-8

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