1887

Journal of General Virology header logo

Volume 103, Issue 11

Orchid fleck dichorhavirus movement protein shows RNA silencing suppressor activity No Access

Abstract

To counteract RNA interference-mediated antiviral defence, virus genomes evolved to express proteins that inhibit this plant defence mechanism. Using six independent biological approaches, we show that orchid fleck dichorhavirus citrus strain (OFV-citrus) movement protein (MP) may act as a viral suppressor of RNA silencing (VSR). By using the alfalfa mosaic virus (AMV) RNA 3 expression vector, it was observed that the MP triggered necrosis response in transgenic tobacco leaves and increased the viral RNA (vRNA) accumulation. The use of the potato virus X (PVX) expression system revealed that the expression of MP increased both the severity of the PVX infection and the accumulation of PVX RNAs, further supporting that MP could act as an RNA silencing suppressor (RSS). From the analysis of the RSS-defective turnip crinkle virus (TCV), we do not find local RSS activity for MP, suggesting a link between MP suppressor activity and the prevention of systemic silencing. In the analysis of local suppressive activity using the GFP-based agroinfiltration assay in (16 c line), we do not identify local RSS activity for the five OFV RNA1-encoded proteins. However, when evaluating the small interfering RNA (siRNA) accumulation, we find that the expression of MP significantly reduces the accumulation of GFP-derived siRNA. Finally, we examine whether the MP can prevent systemic silencing in 16c plants. Our findings show that MP inhibits the long-distance spread of RNA silencing, but does not affect the short-distance spread. Together, our findings indicate that MP is part of OFV’s counter-defence mechanism, acting mainly in the prevention of systemic long-distance silencing. This work presents the first report of a VSR for a member of the genus .

  • Received:
  • Accepted:
  • Published Online:
Keyword(s): dichorhavirus , movement protein , OFV , RNA silencing and viral suppressors of RNA silencing
Funding
This study was supported by the:
  • INVESTIR IMÓVEIS LDTA
    • Principle Award Recipient: MikhailOliveira Leatro
  • Ministerio de Ciencia e Innovación (Award MCIN/AEI/10.13039/501100011033)
    • Principle Award Recipient: VicentePallás
  • Fondo Europeo de Desarrollo Regional (FEDER)
    • Principle Award Recipient: VicentePallás
© 2022 The Authors
Loading

Article metrics loading...

/content/journal/jgv/10.1099/jgv.0.001805
2022-11-18
2024-05-13
Loading full text...

Full text loading...

References

  1. Molnar A, Melnyk C, Baulcombe DC. Silencing signals in plants: a long journey for small RNAs. Genome Biol 2011; 12:215 [View Article]
     [Google Scholar]
  2. Pumplin N, Voinnet O. RNA silencing suppression by plant pathogens: defence, counter-defence and counter-counter-defence. Nat Rev Microbiol 2013; 11:745–760 [View Article] [PubMed]
     [Google Scholar]
  3. Hamilton AJ, Baulcombe DC. A species of small antisense RNA in posttranscriptional gene silencing in plants. Science 1999; 286:950–952 [View Article] [PubMed]
     [Google Scholar]
  4. Hammond SM, Bernstein E, Beach D, Hannon GJ. An RNA-directed nuclease mediates post-transcriptional gene silencing in Drosophila cells. Nature 2000; 404:293–296 [View Article] [PubMed]
     [Google Scholar]
  5. Borges F, Martienssen RA. The expanding world of small RNAs in plants. Nat Rev Mol Cell Biol 2015; 16:727–741 [View Article] [PubMed]
     [Google Scholar]
  6. Takeda A, Iwasaki S, Watanabe T, Utsumi M, Watanabe Y. The mechanism selecting the guide strand from small RNA duplexes is different among argonaute proteins. Plant Cell Physiol 2008; 49:493–500 [View Article]
     [Google Scholar]
  7. Ding SW, Voinnet O. Antiviral immunity directed by small RNAs. Cell 2007; 130:413–426 [View Article] [PubMed]
     [Google Scholar]
  8. Nakanishi K. Anatomy of RISC: how do small RNAs and chaperones activate Argonaute proteins?. Wiley Interdiscip Rev RNA 2016; 7:637–660 [View Article] [PubMed]
     [Google Scholar]
  9. Voinnet O, Pinto YM, Baulcombe DC. Suppression of gene silencing: a general strategy used by diverse DNA and RNA viruses of plants. Proc Natl Acad Sci U S A 1999; 96:14147–14152 [View Article] [PubMed]
     [Google Scholar]
  10. Moon JY, Park JM. Cross-talk in viral defense signaling in plants. Front Microbiol 2016; 7:2068 [View Article]
     [Google Scholar]
  11. Burgyán J, Havelda Z. Viral suppressors of RNA silencing. Trends Plant Sci 2011; 16:265–272 [View Article] [PubMed]
     [Google Scholar]
  12. Li WX, Ding SW. Viral suppressors of RNA silencing. Curr Opin Biotechnol 2001; 12:150–154 [View Article] [PubMed]
     [Google Scholar]
  13. Lu R, Folimonov A, Shintaku M, Li W-X, Falk BW et al. Three distinct suppressors of RNA silencing encoded by a 20-kb viral RNA genome. Proc Natl Acad Sci U S A 2004; 101:15742–15747 [View Article] [PubMed]
     [Google Scholar]
  14. Mérai Z, Kerényi Z, Kertész S, Magna M, Lakatos L et al. Double-stranded RNA binding may be a general plant RNA viral strategy to suppress RNA silencing. J Virol 2006; 80:5747–5756 [View Article] [PubMed]
     [Google Scholar]
  15. Voinnet O, Lederer C, Baulcombe DC. A viral movement protein prevents spread of the gene silencing signal in Nicotiana benthamiana. Cell 2000; 103:157–167 [View Article] [PubMed]
     [Google Scholar]
  16. Hamilton A, Voinnet O, Chappell L, Baulcombe D. Two classes of short interfering RNA in RNA silencing. EMBO J 2002; 21:4671–4679 [View Article] [PubMed]
     [Google Scholar]
  17. Powers JG, Sit TL, Qu F, Morris TJ, Kim K-H et al. A versatile assay for the identification of RNA silencing suppressors based on complementation of viral movement. Mol Plant Microbe Interact 2008; 21:879–890 [View Article] [PubMed]
     [Google Scholar]
  18. Martínez-Pérez M, Navarro JA, Pallás V, Sánchez-Navarro JA. A sensitive and rapid RNA silencing suppressor activity assay based on alfalfa mosaic virus expression vector. Virus Res 2019; 272:197733 [View Article] [PubMed]
     [Google Scholar]
  19. Leastro MO, Castro DYO, Freitas-Astúa J, Kitajima EW, Pallás V et al. Citrus Leprosis Virus C Encodes Three Proteins With Gene Silencing Suppression Activity. Front Microbiol 2020; 11:1231 [View Article]
     [Google Scholar]
  20. Moissiard G, Voinnet O. Viral suppression of RNA silencing in plants. Mol Plant Pathol 2004; 5:71–82 [View Article] [PubMed]
     [Google Scholar]
  21. Voinnet O, Vain P, Angell S, Baulcombe DC. Systemic spread of sequence-specific transgene RNA degradation in plants is initiated by localized introduction of ectopic promoterless DNA. Cell 1998; 95:177–187 [View Article] [PubMed]
     [Google Scholar]
  22. Cañizares MC, Navas-Castillo J, Moriones E. Multiple suppressors of RNA silencing encoded by both genomic RNAs of the crinivirus, Tomato chlorosis virus. Virology 2008; 379:168–174 [View Article] [PubMed]
     [Google Scholar]
  23. Renovell Á, Vives MC, Ruiz-Ruiz S, Navarro L, Moreno P et al. The Citrus leaf blotch virus movement protein acts as silencing suppressor. Virus Genes 2012; 44:131–140 [View Article] [PubMed]
     [Google Scholar]
  24. Gupta AK, Hein GL, Graybosch RA, Tatineni S. Octapartite negative-sense RNA genome of High Plains wheat mosaic virus encodes two suppressors of RNA silencing. Virology 2018; 518:152–162 [View Article] [PubMed]
     [Google Scholar]
  25. Samuel GH, Wiley MR, Badawi A, Adelman ZN, Myles KM. Yellow fever virus capsid protein is a potent suppressor of RNA silencing that binds double-stranded RNA. Proc Natl Acad Sci U S A 2016; 113:13863–13868 [View Article] [PubMed]
     [Google Scholar]
  26. Freitas-Astúa J, Ramos-González PL, Arena GD, Tassi AD, Kitajima EW. Brevipalpus-transmitted viruses: parallelism beyond a common vector or convergent evolution of distantly related pathogens?. Curr Opin Virol 2018; 33:66–73 [View Article] [PubMed]
     [Google Scholar]
  27. Dietzgen RG, Freitas-Astúa J, Chabi-Jesus C, Ramos-González PL, Goodin MM et al. Dichorhaviruses in their Host Plants and Mite Vectors. Adv Virus Res 2018; 102:119–148 [View Article] [PubMed]
     [Google Scholar]
  28. Roy A, Stone AL, Shao J, Otero-Colina G, Wei G et al. Identification and Molecular Characterization of Nuclear Citrus leprosis virus, a Member of the Proposed Dichorhavirus Genus Infecting Multiple Citrus Species in Mexico. Phytopathology 2015; 105:564–575 [View Article] [PubMed]
     [Google Scholar]
  29. Cruz-Jaramillo JL, Ruiz-Medrano R, Rojas-Morales L, López-Buenfil JA, Morales-Galván O et al. Characterization of a proposed dichorhavirus associated with the citrus leprosis disease and analysis of the host response. Viruses 2014; 6:2602–2622 [View Article] [PubMed]
     [Google Scholar]
  30. Cook G, Kirkman W, Clase R, Steyn C, Basson E et al. Orchid fleck virus associated with the first case of citrus leprosis-N in South Africa. Eur J Plant Pathol 2019; 155:1373–1379 [View Article]
     [Google Scholar]
  31. Kondo H, Maruyama K, Chiba S, Andika IB, Suzuki N. Transcriptional mapping of the messenger and leader RNAs of orchid fleck virus, a bisegmented negative-strand RNA virus. Virology 2014; 452–453:166–174 [View Article]
     [Google Scholar]
  32. Kondo H, Chiba S, Andika IB, Maruyama K, Tamada T et al. Orchid fleck virus structural proteins N and P form intranuclear viroplasm-like structures in the absence of viral infection. J Virol 2013; 87:7423–7434 [View Article] [PubMed]
     [Google Scholar]
  33. Leastro MO, Freitas-Astúa J, Kitajima EW, Pallás V, Sánchez-Navarro JA. Dichorhaviruses movement protein and nucleoprotein form a protein complex that may be required for virus spread and interacts in vivo with viral movement-related cilevirus proteins. Front Microbiol 2020; 11:571807 [View Article]
     [Google Scholar]
  34. Dietzgen RG, Kuhn JH, Clawson AN, Freitas-Astúa J, Goodin MM et al. Dichorhavirus: a proposed new genus for Brevipalpus mite-transmitted, nuclear, bacilliform, bipartite, negative-strand RNA plant viruses. Arch Virol 2014; 159:607–619 [View Article] [PubMed]
     [Google Scholar]
  35. Bejerman N, Mann KS, Dietzgen RG. Alfalfa dwarf cytorhabdovirus P protein is a local and systemic RNA silencing supressor which inhibits programmed RISC activity and prevents transitive amplification of RNA silencing. Virus Res 2016; 224:19–28 [View Article] [PubMed]
     [Google Scholar]
  36. Mann KS, Johnson KN, Dietzgen RG. Cytorhabdovirus phosphoprotein shows RNA silencing suppressor activity in plants, but not in insect cells. Virology 2015; 476:413–418 [View Article] [PubMed]
     [Google Scholar]
  37. Guo H, Song X, Xie C, Huo Y, Zhang F et al. Rice yellow stunt rhabdovirus protein 6 suppresses systemic RNA silencing by blocking RDR6-mediated secondary siRNA synthesis. Mol Plant Microbe Interact 2013; 26:927–936 [View Article] [PubMed]
     [Google Scholar]
  38. Rabieifaradonbeh S, Afsharifar A, Finetti-Sialer MM. Molecular and functional characterization of the barley yellow striate mosaic virus genes encoding phosphoprotein, P3, P6 and P9. Eur J Plant Pathol 2021; 161:107–121 [View Article]
     [Google Scholar]
  39. Mann KS, Johnson KN, Carroll BJ, Dietzgen RG. Cytorhabdovirus P protein suppresses RISC-mediated cleavage and RNA silencing amplification in planta. Virology 2016; 490:27–40 [View Article] [PubMed]
     [Google Scholar]
  40. Ruiz MT, Voinnet O, Baulcombe DC. Initiation and maintenance of virus-induced gene silencing. Plant Cell 1998; 10:937–946 [View Article] [PubMed]
     [Google Scholar]
  41. Patil BL, Fauquet CM. Light intensity and temperature affect systemic spread of silencing signal in transient agroinfiltration studies. Mol Plant Pathol 2015; 16:484–494 [View Article] [PubMed]
     [Google Scholar]
  42. van Dun CM, van Vloten-Doting L, Bol JF. Expression of alfalfa mosaic virus cDNA1 and 2 in transgenic tobacco plants. Virology 1988; 163:572–578 [View Article]
     [Google Scholar]
  43. Leastro MO, Freitas-Astúa J, Kitajima EW, Pallás V, Sánchez-Navarro JA. Unravelling the involvement of cilevirus p32 protein in the viral transport. Sci Rep 2021; 11:2943 [View Article]
     [Google Scholar]
  44. Lu R, Malcuit I, Moffett P, Ruiz MT, Peart J et al. High throughput virus-induced gene silencing implicates heat shock protein 90 in plant disease resistance. EMBO J 2003; 22:5690–5699 [View Article] [PubMed]
     [Google Scholar]
  45. Sanchez-Navarro J, Miglino R, Ragozzino A, Bol JF. Engineering of alfalfa mosaic virus RNA 3 into an expression vector. Arch Virol 2001; 146:923–939 [View Article]
     [Google Scholar]
  46. Leastro MO, Villar-Álvarez D, Freitas-Astúa J, Kitajima EW, Pallás V et al. Spontaneous Mutation in the Movement Protein of Citrus Leprosis Virus C2, in a Heterologous Virus Infection Context, Increases Cell-to-Cell Transport and Generates Fitness Advantage. Viruses 2021; 13:2498 [View Article] [PubMed]
     [Google Scholar]
  47. Taschner PE, van der Kuyl AC, Neeleman L, Bol JF. Replication of an incomplete alfalfa mosaic virus genome in plants transformed with viral replicase genes. Virology 1991; 181:445–450 [View Article] [PubMed]
     [Google Scholar]
  48. Loesch-Fries LS, Jarvis NP, Krahn KJ, Nelson SE, Hall TC. Expression of alfalfa mosaic virus RNA 4 cDNA transcripts in vitro and in vivo. Virology 1985; 146:177–187 [View Article] [PubMed]
     [Google Scholar]
  49. Jones L, Hamilton AJ, Voinnet O, Thomas CL, Maule AJ et al. RNA-DNA interactions and DNA methylation in post-transcriptional gene silencing. Plant Cell 1999; 11:2291–2301 [View Article] [PubMed]
     [Google Scholar]
  50. Hellens RP, Edwards EA, Leyland NR, Bean S, Mullineaux PM. pGreen: a versatile and flexible binary Ti vector for Agrobacterium-mediated plant transformation. Plant Mol Biol 2000; 42:819–832 [View Article] [PubMed]
     [Google Scholar]
  51. Leastro MO, Pallás V, Resende RO, Sánchez-Navarro JA. The movement proteins (NSm) of distinct tospoviruses peripherally associate with cellular membranes and interact with homologous and heterologous NSm and nucleocapsid proteins. Virology 2015; 478:39–49 [View Article] [PubMed]
     [Google Scholar]
  52. Pallás V, Más P, Sánchez-Navarro JA. Detection of plant RNA viruses by nonisotopic dot-blot hybridization. Methods Mol Biol 1998; 81:461–468 [View Article] [PubMed]
     [Google Scholar]
  53. Anandalakshmi R, Pruss GJ, Ge X, Marathe R, Mallory AC et al. A viral suppressor of gene silencing in plants. Proc Natl Acad Sci U S A 1998; 95:13079–13084 [View Article] [PubMed]
     [Google Scholar]
  54. Mallory AC, Reinhart BJ, Bartel D, Vance VB, Bowman LH. A viral suppressor of RNA silencing differentially regulates the accumulation of short interfering RNAs and micro-RNAs in tobacco. Proc Natl Acad Sci U S A 2002; 99:15228–15233 [View Article]
     [Google Scholar]
  55. Yang X, Ren Y, Sun S, Wang D, Zhang F et al. Identification of the Potential Virulence Factors and RNA Silencing Suppressors of Mulberry Mosaic Dwarf-Associated Geminivirus. Viruses 2018; 10:E472 [View Article]
     [Google Scholar]
  56. Himber C, Dunoyer P, Moissiard G, Ritzenthaler C, Voinnet O. Transitivity-dependent and -independent cell-to-cell movement of RNA silencing. EMBO J 2003; 22:4523–4533 [View Article] [PubMed]
     [Google Scholar]
  57. Melnyk CW, Molnar A, Baulcombe DC. Intercellular and systemic movement of RNA silencing signals. EMBO J 2011; 30:3553–3563 [View Article] [PubMed]
     [Google Scholar]
  58. Lucas WJ. Plant viral movement proteins: agents for cell-to-cell trafficking of viral genomes. Virology 2006; 344:169–184 [View Article] [PubMed]
     [Google Scholar]
  59. Navarro JA, Sanchez-Navarro JA, Pallas V. Key checkpoints in the movement of plant viruses through the host. Adv Virus Res 2019; 104:1–64 [View Article] [PubMed]
     [Google Scholar]
  60. Amari K, Vazquez F, Heinlein M. Manipulation of plant host susceptibility: an emerging role for viral movement proteins?. Front Plant Sci 2012; 3:10 [View Article] [PubMed]
     [Google Scholar]
  61. Fusaro AF, Barton DA, Nakasugi K, Jackson C, Kalischuk ML et al. The Luteovirus P4 Movement Protein Is a Suppressor of Systemic RNA Silencing. Viruses 2017; 9:E294 [View Article] [PubMed]
     [Google Scholar]
  62. Zhang C, Chen D, Yang G, Yu X, Wu J. Rice Stripe Mosaic Virus-Encoded P4 Is a Weak Suppressor of Viral RNA Silencing and Is Required for Disease Symptom Development. Mol Plant Microbe Interact 2020; 33:412–422 [View Article] [PubMed]
     [Google Scholar]
  63. Qu F, Morris TJ. Suppressors of RNA silencing encoded by plant viruses and their role in viral infections. FEBS Lett 2005; 579:5958–5964 [View Article] [PubMed]
     [Google Scholar]
  64. Carpino C, Ferriol Safont I, Elvira-González L, Medina V, Rubio L et al. RNA2-encoded VP37 protein of Broad bean wilt virus 1 is a determinant of pathogenicity, host susceptibility, and a suppressor of post-transcriptional gene silencing. Mol Plant Pathol 2020; 21:1421–1435 [View Article] [PubMed]
     [Google Scholar]
  65. Curaba J, Chen X. Biochemical activities of Arabidopsis RNA-dependent RNA polymerase 6. J Biol Chem 2008; 283:3059–3066 [View Article] [PubMed]
     [Google Scholar]
  66. Peragine A, Yoshikawa M, Wu G, Albrecht HL, Poethig RS. SGS3 and SGS2/SDE1/RDR6 are required for juvenile development and the production of trans-acting siRNAs in Arabidopsis. Genes Dev 2004; 18:2368–2379 [View Article]
     [Google Scholar]
  67. Mourrain P, Béclin C, Elmayan T, Feuerbach F, Godon C et al. Arabidopsis SGS2 and SGS3 genes are required for posttranscriptional gene silencing and natural virus resistance. Cell 2000; 101:533–542 [View Article] [PubMed]
     [Google Scholar]
  68. Béclin C, Boutet S, Waterhouse P, Vaucheret H. A branched pathway for transgene-induced RNA silencing in plants. Curr Biol 2002; 12:684–688 [View Article]
     [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jgv/10.1099/jgv.0.001805
Loading
Orchid fleck dichorhavirus movement protein shows RNA silencing suppressor activity
J. Gen. Virol. 103, 001805 (2022); https://doi.org/10.1099/jgv.0.001805
/content/journal/jgv/10.1099/jgv.0.001805
/content/journal/jgv/10.1099/jgv.0.001805
Loading

Data & Media loading...

Supplements

Supplementary material 1

PDF

Most cited 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