1887

Abstract

In previous studies, protein interaction maps of different potyviruses have been generated using yeast two-hybrid (YTH) systems, and these maps have demonstrated a high diversity of interactions of potyviral proteins. Using an optimized bimolecular fluorescence complementation (BiFC) system, a complete interaction matrix for proteins of a potyvirus was developed for the first time under conditions with ten proteins from plum pox virus (PPV). In total, 52 of 100 possible interactions were detected, including the self-interactions of CI, 6K2, VPg, NIa-Pro, NIb and CP, which is more interactions than have ever been detected for any other potyvirus in a YTH approach. Moreover, the BiFC system was shown to be able to localize the protein interactions, which was typified for the protein self-interactions indicated above. Additionally, experiments were carried out with the P3N-PIPO protein, revealing an interaction with CI but not with CP and supporting the involvement of P3N-PIPO in the cell-to-cell movement of potyviruses. No self-interaction of the PPV helper component–proteinase (HC-Pro) was detected using BiFC . Therefore, additional experiments with turnip mosaic virus (TuMV) HC-Pro, PPV_HC-Pro and their mutants were conducted. The self-interaction of TuMV_HCpro, as recently demonstrated, and the self-interaction of the TuMV_ and PPV_HC-Pro mutants were shown by BiFC , indicating that HC-Pro self-interactions may be species-specific. BiFC is a very useful and reliable method for the detection and localization of protein interactions , thus enabling investigations under more natural conditions than studies in yeast cells.

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2011-12-01
2019-12-06
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References

  1. Arbatova J., Lehto K., Pehu E., Pehu T.. ( 1998; ). Localization of the P1 protein of potato Y potyvirus in association with cytoplasmic inclusion bodies and in the cytoplasm of infected cells. . J Gen Virol 79:, 2319–2323.[PubMed]
    [Google Scholar]
  2. Atmakuri K., Ding Z., Christie P. J.. ( 2003; ). VirE2, a type IV secretion substrate, interacts with the VirD4 transfer protein at cell poles of Agrobacterium tumefaciens . . Mol Microbiol 49:, 1699–1713. [CrossRef] [PubMed]
    [Google Scholar]
  3. Atreya C. D., Pirone T. P.. ( 1993; ). Mutational analysis of the helper component-proteinase gene of a potyvirus: effects of amino acid substitutions, deletions, and gene replacement on virulence and aphid transmissibility. . Proc Natl Acad Sci U S A 90:, 11919–11923. [CrossRef] [PubMed]
    [Google Scholar]
  4. Beauchemin C., Boutet N., Laliberté J.-F.. ( 2007; ). Visualization of the interaction between the precursors of VPg, the viral protein linked to the genome of Turnip mosaic virus, and the translation eukaryotic initiation factor iso 4E in planta . . J Virol 81:, 775–782. [CrossRef] [PubMed]
    [Google Scholar]
  5. Berger P. H., Hunt A. G., Domier L. L., Hellmann G. M., Stram Y., Thornbury D. W., Pirone T. P.. ( 1989; ). Expression in transgenic plants of a viral gene product that mediates insect transmission of potyviruses. . Proc Natl Acad Sci U S A 86:, 8402–8406. [CrossRef] [PubMed]
    [Google Scholar]
  6. Bevan M.. ( 1984; ). Binary Agrobacterium vectors for plant transformation. . Nucleic Acids Res 12:, 8711–8721. [CrossRef] [PubMed]
    [Google Scholar]
  7. Blanc S., López-Moya J. J., Wang R., García-Lampasona S., Thornbury D. W., Pirone T. P.. ( 1997; ). A specific interaction between coat protein and helper component correlates with aphid transmission of a potyvirus. . Virology 231:, 141–147. [CrossRef] [PubMed]
    [Google Scholar]
  8. Blanc S., Ammar E. D., García-Lampasona S., Dolja V. V., Llave C., Baker J., Pirone T. P.. ( 1998; ). Mutations in the potyvirus helper component protein: effects on interactions with virions and aphid stylets. . J Gen Virol 79:, 3119–3122.[PubMed]
    [Google Scholar]
  9. Blondel M., Bach S., Bamps S., Dobbelaere J., Wiget P., Longaretti C., Barral Y., Meijer L., Peter M.. ( 2005; ). Degradation of Hof1 by SCFGrr1 is important for actomyosin contraction during cytokinesis in yeast. . EMBO J 24:, 1440–1452. [CrossRef] [PubMed]
    [Google Scholar]
  10. Bracha-Drori K., Shichrur K., Katz A., Oliva M., Angelovici R., Yalovsky S., Ohad N.. ( 2004; ). Detection of protein–protein interactions in plants using bimolecular fluorescence complementation. . Plant J 40:, 419–427. [CrossRef] [PubMed]
    [Google Scholar]
  11. Carrington J. C., Jensen P. E., Schaad M. C.. ( 1998; ). Genetic evidence for an essential role for potyvirus CI protein in cell-to-cell movement. . Plant J 14:, 393–400. [CrossRef] [PubMed]
    [Google Scholar]
  12. Choi I. R., Stenger D. C., French R.. ( 2000; ). Multiple interactions among proteins encoded by the mite-transmitted wheat streak mosaic tritimovirus. . Virology 267:, 185–198. [CrossRef] [PubMed]
    [Google Scholar]
  13. Chung B. Y.-W., Miller W. A., Atkins J. F., Firth A. E.. ( 2008; ). An overlapping essential gene in the Potyviridae . . Proc Natl Acad Sci U S A 105:, 5897–5902. [CrossRef] [PubMed]
    [Google Scholar]
  14. Citovsky V., Lee L.-Y., Vyas S., Glick E., Chen M.-H., Vainstein A., Gafni Y., Gelvin S. B., Tzfira T.. ( 2006; ). Subcellular localization of interacting proteins by bimolecular fluorescence complementation in planta . . J Mol Biol 362:, 1120–1131. [CrossRef] [PubMed]
    [Google Scholar]
  15. Citovsky V., Gafni Y., Tzfira T.. ( 2008; ). Localizing protein–protein interactions by bimolecular fluorescence complementation in planta . . Methods 45:, 196–206. [CrossRef] [PubMed]
    [Google Scholar]
  16. Cole K. C., McLaughlin H. W., Johnson D. I.. ( 2007; ). Use of bimolecular fluorescence complementation to study in vivo interactions between Cdc42p and Rdi1p of Saccharomyces cerevisiae . . Eukaryot Cell 6:, 378–387. [CrossRef] [PubMed]
    [Google Scholar]
  17. Cronin S., Verchot J., Haldeman-Cahill R., Schaad M. C., Carrington J. C.. ( 1995; ). Long-distance movement factor: a transport function of the potyvirus helper component proteinase. . Plant Cell 7:, 549–559. [CrossRef] [PubMed]
    [Google Scholar]
  18. Deblaere R., Bytebier B., De Greve H., Deboeck F., Schell J., Van Montagu M., Leemans J.. ( 1985; ). Efficient octopine Ti plasmid-derived vectors for Agrobacterium-mediated gene transfer to plants. . Nucleic Acids Res 13:, 4777–4788. [CrossRef] [PubMed]
    [Google Scholar]
  19. Dolja V. V., Boyko V. P., Agranovsky A. A., Koonin E. V.. ( 1991; ). Phylogeny of capsid proteins of rod-shaped and filamentous RNA plant viruses: two families with distinct patterns of sequence and probably structure conservation. . Virology 184:, 79–86. [CrossRef] [PubMed]
    [Google Scholar]
  20. Dolja V. V., Haldeman R., Robertson N. L., Dougherty W. G., Carrington J. C.. ( 1994; ). Distinct functions of capsid protein in assembly and movement of tobacco etch potyvirus in plants. . EMBO J 13:, 1482–1491.[PubMed]
    [Google Scholar]
  21. Fields S., Song O.. ( 1989; ). A novel genetic system to detect protein–protein interactions. . Nature 340:, 245–246. [CrossRef] [PubMed]
    [Google Scholar]
  22. Golemis E. A., Serebriiskii I., Law S. F.. ( 1999; ). The yeast two-hybrid system: criteria for detecting physiologically significant protein–protein interactions. . Curr Issues Mol Biol 1:, 31–45.[PubMed]
    [Google Scholar]
  23. Guo D., Merits A., Saarma M.. ( 1999; ). Self-association and mapping of interaction domains of helper component-proteinase of potato A potyvirus. . J Gen Virol 80:, 1127–1131.[PubMed]
    [Google Scholar]
  24. Guo D., Rajamäki M. L., Saarma M., Valkonen J. P.. ( 2001; ). Towards a protein interaction map of potyviruses: protein interaction matrixes of two potyviruses based on the yeast two-hybrid system. . J Gen Virol 82:, 935–939.[PubMed]
    [Google Scholar]
  25. Hong Y., Levay K., Murphy J. F., Klein P. G., Shaw J. G., Hunt A. G.. ( 1995; ). A potyvirus polymerase interacts with the viral coat protein and VPg in yeast cells. . Virology 214:, 159–166. [CrossRef] [PubMed]
    [Google Scholar]
  26. Hu C.-D., Kerppola T. K.. ( 2003; ). Simultaneous visualization of multiple protein interactions in living cells using multicolor fluorescence complementation analysis. . Nat Biotechnol 21:, 539–545. [CrossRef] [PubMed]
    [Google Scholar]
  27. Hu C.-D., Chinenov Y., Kerppola T. K.. ( 2002; ). Visualization of interactions among bZIP and Rel family proteins in living cells using bimolecular fluorescence complementation. . Mol Cell 9:, 789–798. [CrossRef] [PubMed]
    [Google Scholar]
  28. Jacquet C., Delecolle B., Raccah B., Lecoq H., Dunez J., Ravelonandro M.. ( 1998; ). Use of modified plum pox virus coat protein genes developed to limit heteroencapsidation-associated risks in transgenic plants. . J Gen Virol 79:, 1509–1517.[PubMed]
    [Google Scholar]
  29. Jagadish M. N., Huang D., Ward C. W.. ( 1993; ). Site-directed mutagenesis of a potyvirus coat protein and its assembly in Escherichia coli . . J Gen Virol 74:, 893–896. [CrossRef] [PubMed]
    [Google Scholar]
  30. Jakab G., Droz E., Brigneti G., Baulcombe D., Malnoë P.. ( 1997; ). Infectious in vivo and in vitro transcripts from a full-length cDNA clone of PVY-N605, a Swiss necrotic isolate of potato virus Y. . J Gen Virol 78:, 3141–3145.[PubMed]
    [Google Scholar]
  31. Kang S.-H., Lim W.-S., Kim K.-H.. ( 2004; ). A protein interaction map of Soybean mosaic virus strain G7H based on the yeast two-hybrid system. . Mol Cells 18:, 122–126.[PubMed]
    [Google Scholar]
  32. Kang S.-H., Lim W.-S., Hwang S.-H., Park J.-W., Choi H.-S., Kim K.-H.. ( 2006; ). Importance of the C-terminal domain of Soybean mosaic virus coat protein for subunit interactions. . J Gen Virol 87:, 225–229. [CrossRef] [PubMed]
    [Google Scholar]
  33. Kasschau K. D., Carrington J. C.. ( 2001; ). Long-distance movement and replication maintenance functions correlate with silencing suppression activity of potyviral HC-Pro. . Virology 285:, 71–81. [CrossRef] [PubMed]
    [Google Scholar]
  34. Li X. H., Valdez P., Olvera R. E., Carrington J. C.. ( 1997; ). Functions of the tobacco etch virus RNA polymerase (NIb): subcellular transport and proteinĄprotein interaction with VPg/proteinase (NIa). . J Virol 71:, 1598–1607.[PubMed]
    [Google Scholar]
  35. Lin L., Shi Y., Luo Z., Lu Y., Zheng H., Yan F., Chen J., Chen J., Adams M. J., Wu Y.. ( 2009; ). Protein–protein interactions in two potyviruses using the yeast two-hybrid system. . Virus Res 142:, 36–40. [CrossRef] [PubMed]
    [Google Scholar]
  36. López L., Urzainqui A., Domínguez E., García J. A.. ( 2001; ). Identification of an N-terminal domain of the plum pox potyvirus CI RNA helicase involved in self-interaction in a yeast two-hybrid system. . J Gen Virol 82:, 677–686.[PubMed]
    [Google Scholar]
  37. López-Moya J. J., Fernández-Fernández M. R., Cambra M., García J. A.. ( 2000; ). Biotechnological aspects of plum pox virus. . J Biotechnol 76:, 121–136. [CrossRef] [PubMed]
    [Google Scholar]
  38. Maiss E., Timpe U., Brisske-Rode A., Lesemann D.-E., Casper R.. ( 1992; ). Infectious in vivo transcripts of a plum pox potyvirus full-length cDNA clone containing the cauliflower mosaic virus 35S RNA promoter. . J Gen Virol 73:, 709–713. [CrossRef] [PubMed]
    [Google Scholar]
  39. Martin K., Kopperud K., Chakrabarty R., Banerjee R., Brooks R., Goodin M. M.. ( 2009; ). Transient expression in Nicotiana benthamiana fluorescent marker lines provides enhanced definition of protein localization, movement and interactions in planta . . Plant J 59:, 150–162. [CrossRef] [PubMed]
    [Google Scholar]
  40. Merits A., Guo D., Järvekülg L., Saarma M.. ( 1999; ). Biochemical and genetic evidence for interactions between potato A potyvirus-encoded proteins P1 and P3 and proteins of the putative replication complex. . Virology 263:, 15–22. [CrossRef] [PubMed]
    [Google Scholar]
  41. Peng Y. H., Kadoury D., Gal-On A., Huet H., Wang Y., Raccah B.. ( 1998; ). Mutations in the HC-Pro gene of zucchini yellow mosaic potyvirus: effects on aphid transmission and binding to purified virions. . J Gen Virol 79:, 897–904.[PubMed]
    [Google Scholar]
  42. Phizicky E. M., Fields S.. ( 1995; ). Protein–protein interactions: methods for detection and analysis. . Microbiol Rev 59:, 94–123.[PubMed]
    [Google Scholar]
  43. Plisson C., Drucker M., Blanc S., German-Retana S., Le Gall O., Thomas D., Bron P.. ( 2003; ). Structural characterization of HC-Pro, a plant virus multifunctional protein. . J Biol Chem 278:, 23753–23761. [CrossRef] [PubMed]
    [Google Scholar]
  44. Riechmann J. L., Laín S., García J. A.. ( 1992; ). Highlights and prospects of potyvirus molecular biology. . J Gen Virol 73:, 1–16. [CrossRef] [PubMed]
    [Google Scholar]
  45. Roudet-Tavert G., German-Retana S., Delaunay T., Delécolle B., Candresse T., Le Gall O.. ( 2002; ). Interaction between potyvirus helper component-proteinase and capsid protein in infected plants. . J Gen Virol 83:, 1765–1770.[PubMed]
    [Google Scholar]
  46. Ruiz-Ferrer V., Boskovic J., Alfonso C., Rivas G., Llorca O., López-Abella D., López-Moya J. J.. ( 2005; ). Structural analysis of tobacco etch potyvirus HC-Pro oligomers involved in aphid transmission. . J Virol 79:, 3758–3765. [CrossRef] [PubMed]
    [Google Scholar]
  47. Schaad M. C., Haldeman-Cahill R., Cronin S., Carrington J. C.. ( 1996; ). Analysis of the VPg-proteinase (NIa) encoded by tobacco etch potyvirus: effects of mutations on subcellular transport, proteolytic processing, and genome amplification. . J Virol 70:, 7039–7048.[PubMed]
    [Google Scholar]
  48. 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. [CrossRef] [PubMed]
    [Google Scholar]
  49. Seo J.-K., Kang S.-H., Seo B. Y., Jung J. K., Kim K.-H.. ( 2010; ). Mutational analysis of interaction between coat protein and helper component-proteinase of Soybean mosaic virus involved in aphid transmission. . Mol Plant Pathol 11:, 265–276. [CrossRef] [PubMed]
    [Google Scholar]
  50. Shen W. T., Wang M. Q., Yan P., Gao L., Zhou P.. ( 2010; ). Protein interaction matrix of Papaya ringspot virus type P based on a yeast two-hybrid system. . Acta Virol 54:, 49–54. [CrossRef] [PubMed]
    [Google Scholar]
  51. Shukla D. D., Frenkel M. J., Ward C. W.. ( 1991; ). Structure and function of the potyvirus genome with special references to the coat protein coding region. . Can J Plant Pathol 13:, 178–191. [CrossRef]
    [Google Scholar]
  52. Shukla D. D., Ward C. W., Brunt A. A.. ( 1994; ). The Potyviridae, p. 516. Wallingford, UK:: CAB International;.
    [Google Scholar]
  53. Stolpe T., Süsslin C., Marrocco K., Nick P., Kretsch T., Kircher S.. ( 2005; ). In planta analysis of protein–protein interactions related to light signaling by bimolecular fluorescence complementation. . Protoplasma 226:, 137–146. [CrossRef] [PubMed]
    [Google Scholar]
  54. Sung M.-K., Huh W.-K.. ( 2007; ). Bimolecular fluorescence complementation analysis system for in vivo detection of protein–protein interaction in Saccharomyces cerevisiae . . Yeast 24:, 767–775. [CrossRef] [PubMed]
    [Google Scholar]
  55. Thornbury D. W., Hellmann G. M., Rhoads R. E., Pirone T. P.. ( 1985; ). Purification and characterization of potyvirus helper component. . Virology 144:, 260–267. [CrossRef] [PubMed]
    [Google Scholar]
  56. Towbin H., Staehelin T., Gordon J.. ( 1979; ). Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. . Proc Natl Acad Sci U S A 76:, 4350–4354. [CrossRef] [PubMed]
    [Google Scholar]
  57. Urcuqui-Inchima S., Walter J., Drugeon G., German-Retana S., Haenni A.-L., Candresse T., Bernardi F., Le Gall O.. ( 1999; ). Potyvirus helper component-proteinase self-interaction in the yeast two-hybrid system and delineation of the interaction domain involved. . Virology 258:, 95–99. [CrossRef] [PubMed]
    [Google Scholar]
  58. Urcuqui-Inchima S., Haenni A.-L., Bernardi F.. ( 2001; ). Potyvirus proteins: a wealth of functions. . Virus Res 74:, 157–175. [CrossRef] [PubMed]
    [Google Scholar]
  59. Varrelmann M., Maiss E.. ( 2000; ). Mutations in the coat protein gene of Plum pox virus suppress particle assembly, heterologous encapsidation and complementation in transgenic plants of Nicotiana benthamiana . . J Gen Virol 81:, 567–576.[PubMed]
    [Google Scholar]
  60. Voinnet O., Pinto Y. M., Baulcombe D. C.. ( 1999; ). Suppression of gene silencing: a general strategy used by diverse DNA and RNA viruses of plants. . Proc Natl Acad Sci U S A 96:, 14147–14152. [CrossRef] [PubMed]
    [Google Scholar]
  61. Voinnet O., Rivas S., Mestre P., Baulcombe D.. ( 2003; ). An enhanced transient expression system in plants based on suppression of gene silencing by the p19 protein of tomato bushy stunt virus. . Plant J 33:, 949–956. [CrossRef] [PubMed]
    [Google Scholar]
  62. Voloudakis A. E., Malpica C. A., Aleman-Verdaguer M.-E., Stark D. M., Fauquet C. M., Beachy R. N.. ( 2004; ). Structural characterization of Tobacco etch virus coat protein mutants. . Arch Virol 149:, 699–712. [CrossRef] [PubMed]
    [Google Scholar]
  63. Walter M., Chaban C., Schütze K., Batistic O., Weckermann K., Näke C., Blazevic D., Grefen C., Schumacher K. et al. & other authors ( 2004; ). Visualization of protein interactions in living plant cells using bimolecular fluorescence complementation. . Plant J 40:, 428–438. [CrossRef] [PubMed]
    [Google Scholar]
  64. Wang R. Y., Pirone T. P.. ( 1999; ). Purification and characterization of turnip mosaic virus helper component protein. . Phytopathology 89:, 564–567. [CrossRef] [PubMed]
    [Google Scholar]
  65. Wei T., Zhang C., Hong J., Xiong R., Kasschau K. D., Zhou X., Carrington J. C., Wang A.. ( 2010a; ). Formation of complexes at plasmodesmata for potyvirus intercellular movement is mediated by the viral protein P3N-PIPO. . PLoS Pathog 6:, e1000962. [CrossRef] [PubMed]
    [Google Scholar]
  66. Wei T., Huang T.-S., McNeil J., Laliberté J.-F., Hong J., Nelson R. S., Wang A.. ( 2010b; ). Sequential recruitment of the endoplasmic reticulum and chloroplasts for plant potyvirus replication. . J Virol 84:, 799–809. [CrossRef] [PubMed]
    [Google Scholar]
  67. Wen R.-H., Hajimorad M. R.. ( 2010; ). Mutational analysis of the putative pipo of soybean mosaic virus suggests disruption of PIPO protein impedes movement. . Virology 400:, 1–7. [CrossRef] [PubMed]
    [Google Scholar]
  68. Xiang C., Han P., Lutziger I., Wang K., Oliver D. J.. ( 1999; ). A mini binary vector series for plant transformation. . Plant Mol Biol 40:, 711–717. [CrossRef] [PubMed]
    [Google Scholar]
  69. Yambao M. L., Masuta C., Nakahara K., Uyeda I.. ( 2003; ). The central and C-terminal domains of VPg of Clover yellow vein virus are important for VPg–HCPro and VPg–VPg interactions. . J Gen Virol 84:, 2861–2869. [CrossRef] [PubMed]
    [Google Scholar]
  70. Zheng H., Yan F., Lu Y., Sun L., Lin L., Cai L., Hou M., Chen J.. ( 2011; ). Mapping the self-interacting domains of TuMV HC-Pro and the subcellular localization of the protein. . Virus Genes 42:, 110–116. [CrossRef] [PubMed]
    [Google Scholar]
  71. Zilian E., Maiss E.. ( 2011; ). An optimized mRFP-based bimolecular fluorescence complementation system for the detection of protein–protein interactions in planta . . J Virol Methods 174:, 158–165. [CrossRef] [PubMed]
    [Google Scholar]
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