1887

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Volume 103, Issue 6

Phosphorylation of the multiple nucleopolyhedrovirus polyhedron envelope protein plays an important role in the formation of the occlusion body envelope No Access

Abstract

During the life cycle of a baculovirus, a crystallized protein matrix, formed by polyhedrin (POLH), is produced. The protein matrix is surrounded by a multilayered protein/carbohydrate envelope, and matrix and envelope together form a mature occlusion body (OB). The polyhedron envelope plays an important role in resistance against adverse external environments. The polyhedron envelope protein (PEP) is the main protein that forms the polyhedron envelope, but the mechanism of formation of the polyhedron envelope is unclear. Here, through immunofluorescence localization observations, we found that PEP interacted with both POLH and P10 during formation of the polyhedron envelope in the late stages of infection, and PEP was also required for P10 incorporation on the surface of OBs. In this process, the phosphorylation of PEP played an important role. PEP was determined to be a phosphorylated protein using the Phos-tag technique, and PK1 was determined to be the phosphokinase of PEP by co-immunoprecipitation and phosphorylation. Immunofluorescence localization revealed that PEP was continuously phosphorylated by PK1 after PEP entered the nucleus until PEP was correctly packaged on the OB surface. Multi-point mutations of PEP conservative potential phosphorylation sites showed that the simultaneous mutation of S85, T86 and Y92 caused changes in the location of PEP and P10 in the late stages of infection, and resulted in an OB surface that lacked the polyhedron envelope. These data suggested that the phosphorylation of PEP at particular sites, i.e. S85, T86 and Y92, plays an important role in the formation of the polyhedron envelope.

  • Received:
  • Accepted:
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Keyword(s): AcMNPV , P10 , PEP , phosphorylation and polyhedron envelope
Funding
This study was supported by the:
  • National Natural Science Foundation of China (Award 31572055)
    • Principle Award Recipient: XiulianSun
© 2022 The Authors
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/content/journal/jgv/10.1099/jgv.0.001759
2022-06-23
2024-04-20
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References

  1. Rohrmann GF. Baculovirus Molecular Biology MD, Bethesda: 2014
     [Google Scholar]
  2. Harrison RL, Herniou EA, Jehle JA, Theilmann DA, Burand JP et al. ICTV Virus Taxonomy Profile: Baculoviridae. J Gen Virol 2018; 99:1185–1186 [View Article] [PubMed]
     [Google Scholar]
  3. Jehle JA, Blissard GW, Bonning BC, Cory JS, Herniou EA et al. On the classification and nomenclature of baculoviruses: a proposal for revision. Arch Virol 2006; 151:1257–1266 [View Article] [PubMed]
     [Google Scholar]
  4. Slack J, Arif BM. The baculoviruses occlusion‐derived virus: virion structure and function. Adv Virus Res 200699–165
     [Google Scholar]
  5. Braunagel SC, Summers MD. Autographa californica nuclear polyhedrosis virus, PDV, and ECV viral envelopes and nucleocapsids: structural proteins, antigens, lipid and fatty acid profiles. Virology 1994; 202:315–328 [View Article] [PubMed]
     [Google Scholar]
  6. Blissard GW, Wenz JR. Baculovirus gp64 envelope glycoprotein is sufficient to mediate pH-dependent membrane fusion. J Virol 1992; 66:6829–6835 [View Article] [PubMed]
     [Google Scholar]
  7. Braunagel SC, Summers MD. Molecular biology of the baculovirus occlusion-derived virus envelope. Curr Drug Targets 2007; 8:1084–1095 [View Article] [PubMed]
     [Google Scholar]
  8. Miller LK. The Baculoviruses, Cytological Changes and Viral Morphogenesis during Baculovirus Infection Boston, MA: New York Springer Science+Business Media; 1997
     [Google Scholar]
  9. Blissard GW, Theilmann DA. Baculovirus entry and egress from insect cells. Annu Rev Virol 2018; 5:113–139 [View Article] [PubMed]
     [Google Scholar]
  10. Rohrmann GF. Polyhedrin structure. J Gen Virol 1986; 67 (Pt 8):1499–1513 [View Article] [PubMed]
     [Google Scholar]
  11. Gross CH, Russell RL, Rohrmann GF. Orgyia pseudotsugata baculovirus p10 and polyhedron envelope protein genes: analysis of their relative expression levels and role in polyhedron structure. J Gen Virol 1994; 75 (Pt 5):1115–1123 [View Article] [PubMed]
     [Google Scholar]
  12. Sajjan DB, Hinchigeri SB. Structural organization of baculovirus occlusion bodies and protective role of multilayered polyhedron envelope protein. Food Environ Virol 2016; 8:86–100 [View Article] [PubMed]
     [Google Scholar]
  13. Minion FC, Coons LB, Broome JR. Characterization of the polyhedral envelope of the nuclear polyhedrosis virus of Heliothis virescens. J Invertebr Pathol 1979; 34:303–307 [View Article]
     [Google Scholar]
  14. Whitt MA, Manning JS. A phosphorylated 34-kDa protein and A subpopulation of polyhedrin are thiol linked to the carbohydrate layer surrounding A baculovirus occlusion body. Virology 1988; 163:33–42 [View Article] [PubMed]
     [Google Scholar]
  15. Russell RL, Pearson MN, Rohrmann GF. Immunoelectron microscopic examination of Orgyia pseudotsugata multicapsid nuclear polyhedrosis virus-infected Lymantria dispar cells: time course and localization of major polyhedron-associated proteins. J Gen Virol 1991; 72:275–283 [View Article] [PubMed]
     [Google Scholar]
  16. Lee SY, Poloumienko A, Belfry S, Qu X, Chen W et al. A common pathway for p10 and calyx proteins in progressive stages of polyhedron envelope assembly in AcMNPV-infected Spodoptera frugiperda larvae. Arch Virol 1996; 141:1247–1258 [View Article] [PubMed]
     [Google Scholar]
  17. Wu X, Gong F, Cao D, Hu X, Wang W. Advances in crop proteomics: PTMs of proteins under abiotic stress. Proteomics 2016; 16:847–865 [View Article] [PubMed]
     [Google Scholar]
  18. Hunter T. Signaling--2000 and beyond. Cell 2000; 100:113–127 [View Article] [PubMed]
     [Google Scholar]
  19. Burnell JN, Hatch MD. Activation and inactivation of an enzyme catalyzed by a single, bifunctional protein: a new example and why. Arch Biochem Biophys 1986; 245:297–304 [View Article] [PubMed]
     [Google Scholar]
  20. Ubersax JA, Ferrell JE. Mechanisms of specificity in protein phosphorylation. Nat Rev Mol Cell Biol 2007; 8:530–541 [View Article] [PubMed]
     [Google Scholar]
  21. Olsen JV, Blagoev B, Gnad F, Macek B, Kumar C et al. Global, in vivo, and site-specific phosphorylation dynamics in signaling networks. Cell 2006; 127:635–648 [View Article] [PubMed]
     [Google Scholar]
  22. Nishi H, Fong JH, Chang C, Teichmann SA, Panchenko AR. Regulation of protein-protein binding by coupling between phosphorylation and intrinsic disorder: analysis of human protein complexes. Mol Biosyst 2013; 9:1620–1626 [View Article] [PubMed]
     [Google Scholar]
  23. Li A, Zhao H, Lai Q, Huang Z, Yuan M et al. Posttranslational modifications of baculovirus protamine-like protein P6.9 and the significance of its hyperphosphorylation for viral very late gene hyperexpression. J Virol 2015; 89:7646–7659 [View Article] [PubMed]
     [Google Scholar]
  24. Lai Q, Wu W, Li A, Wang W, Yuan M et al. The 38K-mediated specific dephosphorylation of the viral core protein p6.9 plays an important role in the nucleocapsid assembly of Autographa californica multiple nucleopolyhedrovirus. J Virol 2018; 92:e01989–01917 [View Article] [PubMed]
     [Google Scholar]
  25. Cheley S, Kosik KS, Paskevich P, Bakalis S, Bayley H. Phosphorylated baculovirus p10 is a heat-stable microtubule-associated protein associated with process formation in Sf9 cells. J Cell Sci 1992; 102 (Pt 4):739–752 [View Article] [PubMed]
     [Google Scholar]
  26. Raza F, McGouran JF, Kessler BM, Possee RD, King LA. Phosphorylation induces structural changes in the Autographa californica nucleopolyhedrovirus P10 protein. J Virol 2017; 91:13 [View Article] [PubMed]
     [Google Scholar]
  27. King LA, Possee RD. The Baculovirus Expression System Dordrecht: Springer; 1992
     [Google Scholar]
  28. Lei C, Yang J, Wang J, Hu J, Sun X. Molecular and biological characterization of Spodoptera frugiperda multiple nucleopolyhedrovirus field isolate and genotypes from China. Insects 2020; 11:E777 [View Article] [PubMed]
     [Google Scholar]
  29. Li J, Zhou Y, Lei C, Fang W, Sun X. Improvement in the UV resistance of baculoviruses by displaying nano-zinc oxide-binding peptides on the surfaces of their occlusion bodies. Appl Microbiol Biotechnol 2015; 99:6841–6853 [View Article] [PubMed]
     [Google Scholar]
  30. Lei C, Yang J, Hu J, Sun X. On the Calculation of TCID50 for Quantitation of Virus Infectivity. Virol Sin 2020; 36:141–144 [View Article]
     [Google Scholar]
  31. Kinoshita E, Kinoshita-Kikuta E, Takiyama K, Koike T. Phosphate-binding tag, a new tool to visualize phosphorylated proteins. Mol Cell Proteomics 2006; 5:749–757 [View Article] [PubMed]
     [Google Scholar]
  32. Hu J, Zhang G, Liang L, Lei C, Sun X. Increased excess intracellular cyclic di-AMP levels impair growth and virulence of Bacillus anthracis. J Bacteriol 2020; 202:e00653-19 [View Article] [PubMed]
     [Google Scholar]
  33. Criglar JM, Anish R, Hu L, Crawford SE, Sankaran B et al. Phosphorylation cascade regulates the formation and maturation of rotaviral replication factories. Proc Natl Acad Sci U S A 2018; 115:E12015–E12023 [View Article] [PubMed]
     [Google Scholar]
  34. Chun K-H, Cho S-J, Lee J-W, Seo JH, Kim K-W et al. Protein kinase C-δ interacts with and phosphorylates ARD1. J Cell Physiol 2021; 236:379–391 [View Article] [PubMed]
     [Google Scholar]
  35. Larkin MA, Blackshields G, Brown NP, Chenna R, McGettigan PA et al. Clustal W and Clustal X version 2.0. Bioinformatics 2007; 23:2947–2948 [View Article] [PubMed]
     [Google Scholar]
  36. Nicholas KN, Deerfield DW. GeneDoc: Analysis and visualization of genetic variation1997 EMBNEW News;
     [Google Scholar]
  37. Kuang W, Zhang H, Wang M, Zhou N-Y, Deng F et al. Three conserved regions in baculovirus sulfhydryl oxidase P33 are critical for enzymatic activity and function. J Virol 2017; 91:e01158–01117 [View Article] [PubMed]
     [Google Scholar]
  38. Graves LP, Hughes LC, Irons SL, Possee RD, King LA. In cultured cells the baculovirus P10 protein forms two independent intracellular structures that play separate roles in occlusion body maturation and their release by nuclear disintegration. PLoS Pathog 2019; 15:e1007827 [View Article] [PubMed]
     [Google Scholar]
  39. Carpentier DCJ, Griffiths CM, King LA. The baculovirus P10 protein of Autographa californica nucleopolyhedrovirus forms two distinct cytoskeletal-like structures and associates with polyhedral occlusion bodies during infection. Virology 2008; 371:278–291 [View Article] [PubMed]
     [Google Scholar]
  40. Liang C, Li M, Dai X, Zhao S, Hou Y et al. Autographa californica multiple nucleopolyhedrovirus PK-1 is essential for nucleocapsid assembly. Virology 2013; 443:349–357 [View Article] [PubMed]
     [Google Scholar]
  41. Li Y, Miller LK. Expression and functional analysis of a baculovirus gene encoding a truncated protein kinase homolog. Virology 1995; 206:314–323 [View Article] [PubMed]
     [Google Scholar]
  42. Zhang X, Xu K, Wei D, Wu W, Yang K et al. Baculovirus infection induces disruption of the nuclear lamina. Sci Rep 2017; 7:7823 [View Article] [PubMed]
     [Google Scholar]
  43. Russell RL, Rohrmann GF. A baculovirus polyhedron envelope protein: immunogold localization in infected cells and mature polyhedra. Virology 1990; 174:177–184 [View Article] [PubMed]
     [Google Scholar]
  44. van Lent JW, Groenen JT, Klinge-Roode EC, Rohrmann GF, Zuidema D et al. Localization of the 34 kDa polyhedron envelope protein in Spodoptera frugiperda cells infected with Autographa californica nuclear polyhedrosis virus. Arch Virol 1990; 111:103–114 [View Article] [PubMed]
     [Google Scholar]
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Phosphorylation of the Autographa californica multiple nucleopolyhedrovirus polyhedron envelope protein plays an important role in the formation of the occlusion body envelope
J. Gen. Virol. 103, 001759 (2022); https://doi.org/10.1099/jgv.0.001759
/content/journal/jgv/10.1099/jgv.0.001759
/content/journal/jgv/10.1099/jgv.0.001759
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