1887

Abstract

Adenoviruses are non-enveloped DNA viruses that replicate in the nucleus of infected cells. One of the core proteins, named pVIII, is a minor capsid protein connecting the core with the inner surface of the capsid. Here, we report the characterization of minor capsid protein pVIII encoded by the L6 region of bovine adenovirus (BAdV)-3. Anti-pVIII serum detected a 24 kDa protein at 12–48 h post-infection and an additional 8 kDa protein at 24–48 h post-infection. While the 24 kDa protein was detected in empty capsids, only the C-terminal-cleaved 8 kDa protein was detected in the mature virion, suggesting that amino acids147–216 of the conserved C-terminus of BAdV-3 pVIII are incorporated in mature virions. Detection of hexon protein associated with both precursor (24 kDa) and cleaved (8 kDa) forms of pVIII suggest that the C-terminus of pVIII interacts with the hexon. The pVIII protein predominantly localizes to the nucleus of BAdV-3-infected cells utilizing the classical importin α/β dependent nuclear import pathway. Analysis of mutant pVIII demonstrated that amino acids 52–72 of the conserved N-terminus bind to importin α-3 with high affinity and are required for the nuclear localization.

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2014-08-01
2024-04-18
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References

  1. Adam S. A. 1999; Transport pathways of macromolecules between the nucleus and the cytoplasm. Curr Opin Cell Biol 11:402–406 [View Article][PubMed]
    [Google Scholar]
  2. Adam S. A., Marr R. S., Gerace L. 1990; Nuclear protein import in permeabilized mammalian cells requires soluble cytoplasmic factors. J Cell Biol 111:807–816 [View Article][PubMed]
    [Google Scholar]
  3. Anand S. K., Gaba A., Singh J., Tikoo S. K. 2014; Bovine adenovirus 3 core protein precursor pVII localizes to mitochondria, and modulates ATP synthesis, mitochondrial Ca2+ and mitochondrial membrane potential. J Gen Virol 95:442–452 [View Article][PubMed]
    [Google Scholar]
  4. Anderson C. W., Baum P. R., Gesteland R. F. 1973; Processing of adenovirus 2-induced proteins. J Virol 12:241–252[PubMed]
    [Google Scholar]
  5. Blanche F., Monegier B., Faucher D., Duchesne M., Audhuy F., Barbot A., Bouvier S., Daude G., Dubois H.other authors 2001; Polypeptide composition of an adenovirus type 5 used in cancer gene therapy. J Chromatogr A 921:39–48 [View Article][PubMed]
    [Google Scholar]
  6. Chelius D., Hühmer A. F., Shieh C. H., Lehmberg E., Traina J. A., Slattery T. K., Pungor E. Jr 2002; Analysis of the adenovirus type 5 proteome by liquid chromatography and tandem mass spectrometry methods. J Proteome Res 1:501–513 [View Article][PubMed]
    [Google Scholar]
  7. Christophe D., Christophe-Hobertus C., Pichon B. 2000; Nuclear targeting of proteins: how many different signals?. Cell Signal 12:337–341 [View Article][PubMed]
    [Google Scholar]
  8. Daikoku T., Kudoh A., Fujita M., Sugaya Y., Isomura H., Shirata N., Tsurumi T. 2005; Architecture of replication compartments formed during Epstein–Barr virus lytic replication. J Virol 79:3409–3418 [View Article][PubMed]
    [Google Scholar]
  9. Davis L. I. 1995; The nuclear pore complex. Annu Rev Biochem 64:865–896 [View Article][PubMed]
    [Google Scholar]
  10. de Bruyn Kops A., Uprichard S. L., Chen M., Knipe D. M. 1998; Comparison of the intranuclear distributions of herpes simplex virus proteins involved in various viral functions. Virology 252:162–178 [View Article][PubMed]
    [Google Scholar]
  11. Depping R., Steinhoff A., Schindler S. G., Friedrich B., Fagerlund R., Metzen E., Hartmann E., Köhler M. 2008; Nuclear translocation of hypoxia-inducible factors (HIFs): involvement of the classical importin α/β pathway. Biochim Biophys Acta 1783:394–404 [View Article][PubMed]
    [Google Scholar]
  12. Dingwall C., Laskey R. A. 1991; Nuclear targeting sequences – a consensus?. Trends Biochem Sci 16:478–481 [View Article][PubMed]
    [Google Scholar]
  13. Feldherr C. M., Kallenbach E., Schultz N. 1984; Movement of a karyophilic protein through the nuclear pores of oocytes. J Cell Biol 99:2216–2222 [View Article][PubMed]
    [Google Scholar]
  14. Gastaldelli M., Imelli N., Boucke K., Amstutz B., Meier O., Greber U. F. 2008; Infectious adenovirus type 2 transport through early but not late endosomes. Traffic 9:2265–2278 [View Article][PubMed]
    [Google Scholar]
  15. Goodwin D. J., Whitehouse A. 2001; A γ-2 herpesvirus nucleocytoplasmic shuttle protein interacts with importin α 1 and α 5. J Biol Chem 276:19905–19912 [View Article][PubMed]
    [Google Scholar]
  16. Görlich D., Mattaj I. W. 1996; Nucleocytoplasmic transport. Science 271:1513–1519 [View Article][PubMed]
    [Google Scholar]
  17. Görlich D., Kostka S., Kraft R., Dingwall C., Laskey R. A., Hartmann E., Prehn S. 1995; Two different subunits of importin cooperate to recognize nuclear localization signals and bind them to the nuclear envelope. Curr Biol 5:383–392 [View Article][PubMed]
    [Google Scholar]
  18. Görlich D., Henklein P., Laskey R. A., Hartmann E. 1996a; A 41 amino acid motif in importin-alpha confers binding to importin-beta and hence transit into the nucleus. EMBO J 15:1810–1817[PubMed]
    [Google Scholar]
  19. Görlich D., Kraft R., Kostka S., Vogel F., Hartmann E., Laskey R. A., Mattaj I. W., Izaurralde E. 1996b; Importin provides a link between nuclear protein import and U snRNA export. Cell 87:21–32 [View Article][PubMed]
    [Google Scholar]
  20. Imamoto N., Shimamoto T., Takao T., Tachibana T., Kose S., Matsubae M., Sekimoto T., Shimonishi Y., Yoneda Y. 1995; In vivo evidence for involvement of a 58 kDa component of nuclear pore-targeting complex in nuclear protein import. EMBO J 14:3617–3626[PubMed]
    [Google Scholar]
  21. Jäkel S., Görlich D. 1998; Importin β, transportin, RanBP5 and RanBP7 mediate nuclear import of ribosomal proteins in mammalian cells. EMBO J 17:4491–4502 [View Article][PubMed]
    [Google Scholar]
  22. Kieff E. 1996; Epstein–Barr virus and its replication. In Virology, 3rd edn. pp. 2343–2396 Edited by Fields B. N., Knipe D. M., Howley P. M., Chanock R. M., Melnick J. L., Monath T., Roizman B., Straus S. E. Philadelphia: Lippincott Williams and Wilkins;
    [Google Scholar]
  23. Köhler M., Speck C., Christiansen M., Bischoff F. R., Prehn S., Haller H., Görlich D., Hartmann E. 1999; Evidence for distinct substrate specificities of importin α family members in nuclear protein import. Mol Cell Biol 19:7782–7791[PubMed]
    [Google Scholar]
  24. Kulshreshtha V., Tikoo S. K. 2008; Interaction of bovine adenovirus-3 33K protein with other viral proteins. Virology 381:29–35 [View Article][PubMed]
    [Google Scholar]
  25. Kulshreshtha V., Babiuk L. A., Tikoo S. K. 2004; Role of bovine adenovirus-3 33K protein in viral replication. Virology 323:59–69 [View Article][PubMed]
    [Google Scholar]
  26. Lai M. C., Lin R. I., Tarn W. Y. 2001; Transportin-SR2 mediates nuclear import of phosphorylated SR proteins. Proc Natl Acad Sci U S A 98:10154–10159 [View Article][PubMed]
    [Google Scholar]
  27. Lim R. Y. H., Aebi U., Fahrenkrog B. 2008; Towards reconciling structure and function in the nuclear pore complex. Histochem Cell Biol 129:105–116 [View Article][PubMed]
    [Google Scholar]
  28. Liu G. Q., Babiss L. E., Volkert F. C., Young C. S., Ginsberg H. S. 1985; A thermolabile mutant of adenovirus 5 resulting from a substitution mutation in the protein VIII gene. J Virol 53:920–925[PubMed]
    [Google Scholar]
  29. Liu Y., Vellekamp G., Chen G., Mirza U. A., Wylie D., Twarowska B., Tang J. T., Porter F. W., Wang S.other authors 2003; Proteomic study of recombinant adenovirus 5 encoding human p53 by matrix-assisted laser desorption/ionization mass spectrometry in combination with database search. Int J Mass Spectrom 226:55–69 [View Article]
    [Google Scholar]
  30. Liu H., Jin L., Koh S. B., Atanasov I., Schein S., Wu L., Zhou Z. H. 2010; Atomic structure of human adenovirus by cryo-EM reveals interactions among protein networks. Science 329:1038–1043 [View Article][PubMed]
    [Google Scholar]
  31. Makadiya N. 2013; Functional characterization of 100K protein of bovine adenovirus type 3. PhD thesis, University of Saskatchewan.
    [Google Scholar]
  32. McConnell M. J., Imperiale M. J. 2004; Biology of adenovirus and its use as a vector for gene therapy. Hum Gene Ther 15:1022–1033 [View Article][PubMed]
    [Google Scholar]
  33. Moore M. S., Blobel G. 1993; The GTP-binding protein Ran/TC4 is required for protein import into the nucleus. Nature 365:661–663 [View Article][PubMed]
    [Google Scholar]
  34. Nemerow G. R., Pache L., Reddy V., Stewart P. L. 2009; Insights into adenovirus host cell interactions from structural studies. Virology 384:380–388 [View Article][PubMed]
    [Google Scholar]
  35. Newmeyer D. D., Forbes D. J. 1988; Nuclear import can be separated into distinct steps in vitro: nuclear pore binding and translocation. Cell 52:641–653 [View Article][PubMed]
    [Google Scholar]
  36. Paine P. L., Moore L. C., Horowitz S. B. 1975; Nuclear envelope permeability. Nature 254:109–114 [View Article][PubMed]
    [Google Scholar]
  37. Paterson C. P., Ayalew L. E., Tikoo S. K. 2012; Mapping of nuclear import signal and importin α3 binding regions of 52K protein of bovine adenovirus-3. Virology 432:63–72 [View Article][PubMed]
    [Google Scholar]
  38. Reddy P. S., Idamakanti N., Zakhartchouk A. N., Baxi M. K., Lee J. B., Pyne C., Babiuk L. A., Tikoo S. K. 1998; Nucleotide sequence, genome organization, and transcription map of bovine adenovirus type 3. J Virol 72:1394–1402[PubMed]
    [Google Scholar]
  39. Reddy V. S., Natchiar S. K., Stewart P. L., Nemerow G. R. 2010; Crystal structure of human adenovirus at 3.5 Å resolution. Science 329:1071–1075 [View Article][PubMed]
    [Google Scholar]
  40. Richardson W. D., Mills A. D., Dilworth S. M., Laskey R. A., Dingwall C. 1988; Nuclear protein migration involves two steps: rapid binding at the nuclear envelope followed by slower translocation through nuclear pores. Cell 52:655–664 [View Article][PubMed]
    [Google Scholar]
  41. Rohn K., Prusas C., Monreal G., Hess M. 1997; Identification and characterization of penton base and pVIII protein of egg drop syndrome virus. Virus Res 47:59–65 [View Article][PubMed]
    [Google Scholar]
  42. San Martín C. 2012; Latest insights on adenovirus structure and assembly. Viruses 4:847–877 [View Article][PubMed]
    [Google Scholar]
  43. Singh M., Shmulevitz M., Tikoo S. K. 2005; A newly identified interaction between IVa2 and pVIII proteins during porcine adenovirus type 3 infection. Virology 336:60–69 [View Article][PubMed]
    [Google Scholar]
  44. Tachibana T., Hieda M., Miyamoto Y., Kose S., Imamoto N., Yoneda Y. 2000; Recycling of importin α from the nucleus is suppressed by loss of RCC1 function in living mammalian cells. Cell Struct Funct 25:115–123 [View Article][PubMed]
    [Google Scholar]
  45. Takahashi E., Cohen S. L., Tsai P. K., Sweeney J. A. 2006; Quantitation of adenovirus type 5 empty capsids. Anal Biochem 349:208–217 [View Article][PubMed]
    [Google Scholar]
  46. Theerthagiri G., Eisenhardt N., Schwarz H., Antonin W. 2010; The nucleoporin Nup188 controls passage of membrane proteins across the nuclear pore complex. J Cell Biol 189:1129–1142 [View Article][PubMed]
    [Google Scholar]
  47. Tollefson A.E., Hermiston T.W., Wold W.S. 1998; Preparation and titration of CsCl2-banded adenovirus stock.. In Adenovirus Methods and Protocols pp. 1–9 Edited by Wold W. S. M. Totowa: Humana Press; [View Article]
    [Google Scholar]
  48. Tremblay M. L., Déry C. V., Talbot B. G., Weber J. 1983; In vitro cleavage specificity of the adenovirus type 2 proteinase. Biochim Biophys Acta 743:239–245 [View Article][PubMed]
    [Google Scholar]
  49. Vellekamp G., Porter F. W., Sutjipto S., Cutler C., Bondoc L., Liu Y.-H., Wylie D., Cannon-Carlson S., Tang J. T.other authors 2001; Empty capsids in column-purified recombinant adenovirus preparations. Hum Gene Ther 12:1923–1936 [View Article][PubMed]
    [Google Scholar]
  50. Wang P., Palese P., O’Neill R. E. 1997; The NPI-1/NPI-3 (karyopherin alpha) binding site on the influenza a virus nucleoprotein NP is a nonconventional nuclear localization signal. J Virol 71:1850–1856[PubMed]
    [Google Scholar]
  51. Webster A., Russell W. C., Kemp G. D. 1989; Characterization of the adenovirus proteinase: development and use of a specific peptide assay. J Gen Virol 70:3215–3223 [View Article][PubMed]
    [Google Scholar]
  52. Welch K., Franke J., Köhler M., Macara I. G. 1999; RanBP3 contains an unusual nuclear localization signal that is imported preferentially by importin-α3. Mol Cell Biol 19:8400–8411[PubMed]
    [Google Scholar]
  53. Wu Q., Tikoo S. K. 2004; Altered tropism of recombinant bovine adenovirus type-3 expressing chimeric fiber. Virus Res 99:9–15 [View Article][PubMed]
    [Google Scholar]
  54. Yoneda Y., Imamoto-Sonobe N., Yamaizumi M., Uchida T. 1987; Reversible inhibition of protein import into the nucleus by wheat germ agglutinin injected into cultured cells. Exp Cell Res 173:586–595 [View Article][PubMed]
    [Google Scholar]
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