IE1 and facilitate the localization of nucleopolyhedrovirus ORF8 to specific nuclear sites Free

Abstract

The nucleopolyhedrovirus (BmNPV) ORF8 protein has previously been reported to colocalize with IE1 to specific nuclear sites during infection. Transient expression of green fluorescent protein (GFP)-fused ORF8 showed the protein to have cytoplasmic localization, but following BmNPV infection the protein formed foci, suggesting that ORF8 requires some other viral factor(s) for this. Therefore, interacting factors were looked for using the yeast two-hybrid system and IE1 was identified. We mapped the interacting region of ORF8 using a yeast two-hybrid assay. An N-terminal region (residues 1–110) containing a predicted coiled-coil domain interacted with IE1, while a truncated N-terminal region (residues 1–78) that lacks this domain did not. In addition, a protein with a complete deletion of the N-terminal region failed to interact with IE1. These results suggest that the ORF8 N-terminal region containing the coiled-coil domain is required for the interaction with IE1. Next, whether IE1 plays a role in ORF8 localization was investigated. In the presence of IE1, GFP-ORF8 localized to the nucleus. In addition, cotransfection with a plasmid expressing IE1 and a plasmid containing the element resulted in nuclear foci formation. A GFP-fused ORF8 mutant protein containing the coiled-coil domain, previously shown to interact with IE1, also formed nuclear foci in the presence of IE1 and . However, ORF8 mutant proteins that did not interact with IE1 failed to form nuclear foci. In contrast to wild-type IE1, focus formation was not observed for an IE1 mutant protein that was deficient in binding. These results suggest that IE1 and facilitate the localization of BmNPV ORF8 to specific nuclear sites.

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2005-11-01
2024-03-28
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References

  1. Beniya H., Braunagel S. C., Summers M. D. 1998; Autographa californica nuclear polyhedrosis virus: subcellular localization and protein trafficking of BV/ODV-E26 to intranuclear membranes and viral envelopes. Virology 240:64–75 [CrossRef]
    [Google Scholar]
  2. Guarino L. A., Summers M. D. 1988; Functional mapping of Autographa californica nuclear polyhedrosis virus genes required for late gene expression. J Virol 62:463–471
    [Google Scholar]
  3. Higuchi R. 1990; Recombinant PCR. In PCR Protocol, a Guide to Methods and Application . pp  177–183 Edited by Gelfand M. A., Sninsky J. J., White T. J. San Diego, CA: Academic Press;
  4. Imai N., Kurihara M., Matsumoto S., Kang W. 2004; Bombyx mori nucleopolyhedrovirus orf8 encodes a nucleic acid binding protein that colocalizes with IE1 during infection. Arch Virol 149:1581–1594
    [Google Scholar]
  5. Imai N., Matsumoto S., Kang W. 2005; The formation of BmNPV IE2 nuclear foci is regulated by the functional domains for oligomerization and ubiquitin ligase activity. J Gen Virol 86:637–644 [CrossRef]
    [Google Scholar]
  6. Kang W., Suzuki M., Zemskov E., Okano K., Maeda S. 1999; Characterization of baculovirus repeated open reading frames ( bro ) in Bombyx mori nucleopolyhedrovirus. J Virol 73:10339–10345
    [Google Scholar]
  7. Kang W., Imai N., Suzuki M., Iwanaga M., Matsumoto S., Zemskov E. A. 2003; Interaction of Bombyx mori nucleopolyhedrovirus BRO-A and host cell protein LAMININ. Arch Virol 148:99–113 [CrossRef]
    [Google Scholar]
  8. Kawasaki Y., Matsumoto S., Nagamine T. 2004; Analysis of baculovirus IE1 in living cells: dynamics and spatial relationships to viral structural proteins. J Gen Virol 85:3575–3583 [CrossRef]
    [Google Scholar]
  9. Laemmli U. K. 1970; Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685 [CrossRef]
    [Google Scholar]
  10. Maeda S. 1989; Gene transfer vectors of a baculovirus, Bombyx mori , and their use for expression of foreign genes in insect cells. In Invertebrate Cell System Applications vol. I pp  167–181 Edited by Mitsuhashi J. Boca Raton, FL: CRC Press;
    [Google Scholar]
  11. Maeda S. 1995; Further development of recombinant baculovirus insecticides. Curr Opin Biotechnol 6:313–319 [CrossRef]
    [Google Scholar]
  12. Maeda S., Kawai T., Obinata M., Fujiwara H., Horiuchi T., Saeki Y., Sato Y., Furusawa M. 1985; Production of human α -interferon in silkworm using a baculovirus vector. Nature 315:592–594 [CrossRef]
    [Google Scholar]
  13. Maul G. G. 1998; Nuclear domain 10, the site of DNA virus transcription and replication. Bioessays 20:660–667 [CrossRef]
    [Google Scholar]
  14. Nagamine T., Kawasaki Y., Iizuka T., Matsumoto S. 2005; Focal distribution of baculovirus IE1 triggered by its binding to the hr DNA elements. J Virol 79:39–46 [CrossRef]
    [Google Scholar]
  15. Okano K., Mikhailov V. S., Maeda S. 1999; Colocalization of baculovirus IE-1 and two DNA-binding proteins, DBP and LEF-3, to viral replication factories. J Virol 73:110–119
    [Google Scholar]
  16. O'Reilly D. R., Passarelli A. L., Goldman I. F., Miller L. K. 1990; Characterization of the DA26 gene in a hyper variable region of the Autographa californica nuclear polyhedrosis virus genome. J Gen Virol 71:1029–1037 [CrossRef]
    [Google Scholar]
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