Summary
The fate of Semliki Forest virus (SFV) nucleocapsid, especially the capsid protein (C-protein), was investigated during the early stages of a productive infection in mosquitoAedes albopictus cells. Infection of the cells resulted in a time dependent accumulation of a C-protein derived fragment. This fragmentation of incoming viral nucleocapsid was prevented by NH4Cl, an agent generally used to elevate the pH in acidic intracellular compartments, suggesting that a low intravesicular pH is required for this process. Density gradient analysis of the postnuclear cell lysate demonstrated that the fragmentation was associated with a cellular compartment showing a density of 1.14±0.02 g/ml. This cellular compartment was devoid from a lysosomal marker enzyme and represented the timely preceding cellular fraction through which SFV passed before encountering a lysosomal fraction. Furthermore, the intracellular distribution of the viral,3H-uridine-labeled RNA suggested that the same fraction might represent a key cellular compartment in which the separation of the viral RNA from the viral structural proteins is primed. In conclusion, these data lead to the suggestion that the fragmentation of incoming SFV nucleocapsids inAedes albopictus cells might be the part of the mechanism leading to the release of viral RNA into the cytosol during early stages of productive infection.
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References
Coombs K, Brown B, Brown DT (1984) Evidence for a change in capsid morphology during Sindbis virus envelopment. Virus Res 1: 297–302
Froschauer S, Kartenbeck J, Helenius A (1988) Alphavirus RNA replicase is located on the cytoplasmic surface of endosomes and lysosomes. J Cell Biol 107: 2075–2086
Geigenmüller-Gnirke U, Nitschko H, Schlesinger S (1993) Deletion analysis of the capsid protein of Sindbis virus: identification of the RNA binding region. J Virol 67: 1620–1626
Hase T, Summers PT, Cohen WH (1989) A comparative study of entry modes into C6/36 cells by Semliki Forest and Japanese encephalitis viruses. Arch Virol 108: 101–114
Helenius A, Kartenbeck J, Simons K, Fries E (1980) On the entry of Semliki Forest virus into BHK-21 cells. J Cell Biol 84: 404–420
Helenius A, Marsh M, White J (1982) Inhibition of Semliki Forest virus penetration by lysosomotropic weak base. J Gen Virol 58: 47–61
Helenius A (1984) Semliki Forest virus penetration from endosomes. Biol Cell 51: 181–186
Igarashi A (1987) Isolation of a Singh's Aedes albopictus cell clone sensitive to Dengue and Chikungunya viruses. J Gen Virol 40: 531–544
Kielian MC, Helenius A (1985) pH-induced alterations in the fusogenic spike protein of Semliki Forest virus. J Cell Biol 101: 2284–2291
Lambert N (1989) Isolation of microsomes. In: Harris ELV, Angal S (eds) Protein purification methods: a practical approach, III. Series. IRL Press, Oxford, pp 101–108
Lanzrein M, Weingart R, Kempf C (1993) pH-dependent pore formation in Semliki Forest virus-infected Aedes albopictus cells. Virology 193: 296–302
Lanzrein M, Schlegel A, Kempf C (1994) Entry and uncoating of enveloped viruses. Biochem J 302: 313–320
Lowry OH, Rosebrough NJ, Farr AL, Randall RJ (1951) protein determination with the folin reagent. J Biol Chem 193: 265–275
Marsh M, Bolzau E, helenius A (1983) Penetration of Semliki Forest virus from acidic prelysosomal vacuoles. Cell 32: 931–940
Mauracher CA, Gillam S, Shukin R, Tingle AJ (1991) pH-dependent solubility shift of rubella virus capsid protein. Virology 181: 773–777
Owen KE, Kuhn RJ (1996) Identification of a region in the Sindbis virus nucleocapsid protein that is involved in specificity of RNA encapsidation. J Virol 70: 2757–2763
Pertoft H, Laurent TC, Seljelid R, Akerström G, Kagedal L, Hirstenstein M (1979) The use of density gradients of Percoll for the separation of biological particles. In: Peeters H (ed) Separation of cells and subcellular elements. Pergamon Press Ltd., Oxford, pp 67–72
Poole B, Ohkuma S (1981) Effect of week bases on the intralysosomal pH in mouse peritoneal macrophages. J Cell Biol 90: 665–669
Randolph VB, Stollar V (1990) Low pH-induced cell fusion in flavivirus infected Aedes albopictus cell cultures. J Gen Virol 71: 1845–1850
Rickwood D (1983) Iodinated density gradient media: a practical approach. IRL Press, Oxford
Schägger H, von Jagow G (1987) Tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis for the separation of proteins in the range from 1 to 100 kDa. Anal Biochem 166: 368–379
Schlegel A, Omar A, Jentsch P, Morell A, Kempf C (1991) Semliki Forest virus envelope proteins function as proton channels. Biosci Rep 11: 243–255
Schlegel A, Schaller J, Jentsch P, Kempf C (1993) Semliki Forest virus core protein fragmentation: its possible role in nucleocapsid disassembly. Biosci Rep 13: 333–347
Schmid S, Fuchs R, Male P, Mellman (1988) Two distinct populations of endosomes involved in membrane recycling and transport to lysosomes. Cell 52: 73–83
Schmid S, Fuchs R, Kielian M, Helenius A, Mellman I (1989) Acidification of endosome subpopulations in wild-type Chinese hamster ovary cells and temperature-sensitive acidification-defective mutants. J Cell Biol 108: 1291–1300
Singh I, Helenius A (1992) Role of ribosomes in Semliki Forest virus nucleocapsid uncoating. J Virol 66: 7049–7058
Söderlund H, von Bonsdorff CH, Ulmanen I (1979) Comparison of the structural properties of Sindbis and Semliki Forest virus nucleocapsids. J Gen Virol 45: 15–26
Strong RK, Harrison SC (1990) Proteolytic dissection of Sindbis virus core protein. J Virol 64: 3992–3994
Stubbs MJ, Miller A, Sizer PJH, Stephenson JR, Crooks AJ (1991) X-ray solution scattering of Sindbis virus: changes in conformation induced at low pH. J Mol Biol 221: 39–42
Ulmanen I, Soderlund H, Kääriäinen L (1979) Role of protein synthesis in the assembly of Semliki Forest virus nucleocapsid. Virology 99: 265–276
Wengler G, Wengler G (1984) Identification of a transfer of viral core protein to cellular ribosomes during the early stages of alphavirus infection. Virology 134: 435
Wengler G (1987) The mode of assembly of alphavirus cores implies a mechanism for the disassembly of the cores in the early stages of infection. Arch Virol 94: 1–14
Wengler G (1990) Structure and synthesis of the core protein: role in regulation of assembly and disassembly of alphavirus and flavivirus cores. In: Brinton MA, Heinz FX (ed) New aspects of positive-strand RNA viruses. American Society for Microbiology, Washington, pp 227–236
Wengler G, Würkner D, Wengler G (1992) Identification of a sequence element in the alphavirus core protein which mediates interaction of cores with ribosomes and the disassembly of cores. Virology 191: 880–888
Young JD-E, Young GPH, Cohn ZA, Lenard J (1983) Interaction of enveloped viruses with planar bilayer membranes: observations on Sendai, influenza, vesicular stomatitis, and Semliki Forest viruses. Virology 128: 186–194
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Mrkic, B., Kempf, C. The fragmentation of incoming Semliki Forest virus nucleocapsids in mosquito (Aedes Albopictus) cells might be coupled to virion uncoating. Archives of Virology 141, 1805–1821 (1996). https://doi.org/10.1007/BF01718196
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DOI: https://doi.org/10.1007/BF01718196