1887

Abstract

Intracellular processing and trafficking of the baculovirus expressed cathepsin (V-CATH), which lacks canonical targeting signals, are poorly understood. The cathepsins of multiple nucleopolyhedrovirus (AcMNPV), multiple nucleopolyhedrovirus (CfMNPV) and most other alphabaculovirus group I nucleopolyhedroviruses have well-conserved N-termini containing overlapping chymotrypsin-cleavage (Y) and myristoylation (G) motifs, which are suggestive of proteolytic signal-peptide cleavage to generate proV-CATH and subsequent acylation. To determine proteolytic N-terminal processing of V-CATH, haemagglutinin epitope-coding tags were fused to the 5′ and/or 3′ ends of AcMNPV and CfMNPV . Immunoblot analysis suggested that a small N-terminal peptide is cleaved for both viruses, indicating that is expressed as a pre-proenzyme. The two viral homologues undergo similar proteolytic processing, but have different glycosylation or other post-translational modifications. An AcMNPV V-CATH–DsRED fusion protein co-localized to the endoplasmic reticulum with an HDEL motif-containing green fluorescent protein. Based on these findings, pre-proV-CATH processing and trafficking mechanisms are postulated.

Loading

Article metrics loading...

/content/journal/jgv/10.1099/vir.0.007740-0
2009-04-01
2020-09-26
Loading full text...

Full text loading...

/deliver/fulltext/jgv/90/4/995.html?itemId=/content/journal/jgv/10.1099/vir.0.007740-0&mimeType=html&fmt=ahah

References

  1. Daimon T., Katsuma S., Shimada T. 2007; Mutational analysis of active site residues of chitinase from Bombyx mori nucleopolyhedrovirus. Virus Res 124:168–175 [CrossRef]
    [Google Scholar]
  2. de Jong J. G., Lauzon H. A., Dominy C., Poloumienko A., Carstens E. B., Arif B. M., Krell P. J. 2005; Analysis of the Choristoneura fumiferana nucleopolyhedrovirus genome. J Gen Virol 86:929–943 [CrossRef]
    [Google Scholar]
  3. Haseloff J., Siemering K. R., Prasher D. C., Hodge S. 1997; Removal of a cryptic intron and subcellular localization of green fluorescent protein are required to mark transgenic Arabidopsis plants brightly. Proc Natl Acad Sci U S A 94:2122–2127 [CrossRef]
    [Google Scholar]
  4. Hawtin R. E., Zarkowska T., Arnold K., Thomas C. J., Gooday G. W., King L. A., Kuzio J. A., Possee R. D. 1997; Liquefaction of Autographa californica nucleopolyhedrovirus-infected insects is dependent on the integrity of virus-encoded chitinase and cathepsin genes. Virology 238:243–253 [CrossRef]
    [Google Scholar]
  5. Hodgson J. J., Arif B. M., Krell P. J. 2007; Reprogramming the chiA expression profile of Autographa californica multiple nucleopolyhedrovirus. J Gen Virol 88:2479–2487 [CrossRef]
    [Google Scholar]
  6. Hom L. G., Volkman L. E. 2000; Autographa californica M nucleopolyhedrovirus chiA is required for processing of V-CATH. Virology 277:178–183 [CrossRef]
    [Google Scholar]
  7. Hom L. G., Ohkawa T., Trudeau D., Volkman L. E. 2002; Autographa californica M nucleopolyhedrovirus ProV-CATH is activated during infected cell death. Virology 296:212–218 [CrossRef]
    [Google Scholar]
  8. Kaba S. A., Salcedo A. M., Wafula P. O., Vlak J. M., van Oers M. M. 2004; Development of a chitinase and v-cathepsin negative bacmid for improved integrity of secreted recombinant proteins. J Virol Methods 122:113–118 [CrossRef]
    [Google Scholar]
  9. Lanier L. M., Slack J. M., Volkman L. E. 1996; Actin binding and proteolysis by the baculovirus AcMNPV: the role of virion-associated V-CATH. Virology 216:380–388 [CrossRef]
    [Google Scholar]
  10. Maurer-Stroh S., Eisenhaber F. 2004; Myristoylation of viral and bacterial proteins. Trends Microbiol 12:178–185 [CrossRef]
    [Google Scholar]
  11. Resh M. D. 1999; Fatty acylation of proteins: new insights into membrane targeting of myristoylated and palmitoylated proteins. Biochim Biophys Acta 14511–16 [CrossRef]
    [Google Scholar]
  12. Saville G. P., Thomas C. J., Possee R. D., King L. A. 2002; Partial redistribution of the Autographa californica nucleopolyhedrovirus chitinase in virus-infected cells accompanies mutation of the carboxy-terminal KDEL ER-retention motif. J Gen Virol 83:685–694
    [Google Scholar]
  13. Shi X., Harrison R. L., Hollister J. R., Mohammed A., Fraser M. J., Jr & Jarvis D. L. 2007; Construction and characterization of new piggyBac vectors for constitutive or inducible expression of heterologous gene pairs and the identification of a previously unrecognized activator sequence in piggyBac. BMC Biotechnol 7:5 [CrossRef]
    [Google Scholar]
  14. Siemering K. R., Golbik R., Sever R., Haseloff J. 1996; Mutations that suppress the thermosensitivity of green fluorescent protein. Curr Biol 6:1653–1663 [CrossRef]
    [Google Scholar]
  15. Slack J. M., Kuzio J., Faulkner P. 1995; Characterization of v-cath, a cathepsin L-like proteinase expressed by the baculovirus Autographa californica multiple nuclear polyhedrosis virus. J Gen Virol 76:1091–1098 [CrossRef]
    [Google Scholar]
  16. Slack J. M., Ribeiro B. M., de Souza M. L. 2004; The gp64 locus of Anticarsia gemmatalis multicapsid nucleopolyhedrovirus contains a 3′ repair exonuclease homologue and lacks v-cath and ChiA genes. J Gen Virol 85:211–219 [CrossRef]
    [Google Scholar]
  17. Thomas C. J., Brown H. L., Hawes C. R., Lee B. Y., Min M. K., King L. A., Possee R. D. 1998; Localization of a baculovirus-induced chitinase in the insect cell endoplasmic reticulum. J Virol 72:10207–10212
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jgv/10.1099/vir.0.007740-0
Loading
/content/journal/jgv/10.1099/vir.0.007740-0
Loading

Data & Media loading...

Most cited this month Most Cited RSS feed

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error