Virus Genes

, Volume 52, Issue 2, pp 303–307 | Cite as

ORF43 of maize rayado fino virus is dispensable for systemic infection of maize and transmission by leafhoppers

  • Michael C. Edwards
  • John J. Weiland
  • Jane Todd
  • Lucy R. Stewart
  • Shunwen Lu
Article
First Online:
Received:
Accepted:

Abstract

Maize rayado fino virus (MRFV) possesses an open reading frame (ORF43) predicted to encode a 43 kDa protein (p43) that has been postulated to be a viral movement protein. Using a clone of MRFV (pMRFV-US) from which infectious RNA can be produced, point mutations were introduced to either prevent initiation from three potential AUG initiation codons near the 5′-end of ORF43 or prematurely terminate translation of ORF43. Inoculation of maize seed via vascular puncture inoculation (VPI) resulted in plants exhibiting symptoms typical of MRFV infection for all mutants tested. Furthermore, corn leafhoppers (Dalbulus maidis) transmitted the virus mutants to healthy plants at a frequency similar to that for wild-type MRFV-US. Viral RNA recovered from plants infected with mutants both prior to and after leafhopper transmission retained mutations blocking ORF43 expression. The results indicate that ORF43 of MRFV is dispensable for both systemic infection of maize and transmission by leafhoppers.

Keywords

Tymovirus Marafivirus Virus movement Movement protein Overlapping protein 
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Notes

Acknowledgments

The authors would like to thank Andy Winter and Renee McClean for technical assistance. Funding for this work was provided by USDA-ARS CRIS project numbers 3060-22000-048-00D and 5082-22000-013-00D.

Supplementary material

11262_2016_1287_MOESM1_ESM.eps (24 mb)
Symptoms induced three weeks post inoculation by MRFV-US, initiation mutant MRFV Init1-4, and termination mutant MRFV Term3–3. Additional Fig. 1S (EPS 24595 kb)
11262_2016_1287_MOESM2_ESM.eps (1020 kb)
Accumulation of MRFV CP in maize infected with MRFV-US and ORF 43 mutants MRFV-Init1–4 and MRFV-Term 3–3. A partially purified preparation of MRFV (MRFV prep; ~150 ng) and a healthy plant extract served as checks. Maize seed (Zea mays cv ‘Silver Queen’) was inoculated by VPI using capped transcripts of MRFV-US and each of the ORF 43 mutants. Tissues from symptomatic plants and a healthy check plant were harvested at 12 days post-inoculation. Extracts were prepared from tissue using extraction buffer in a ratio of 3 ml buffer per gram tissue as previously described (Edwards et al. [3]) and then subjected to electrophoresis on a 12% denaturing polyacrylamide gel. After electroblotting to nitrocellulose, the blot was incubated with a 1:1000 dilution of rabbit anti-MRFV antiserum, followed by incubation in a 1:5000 dilution of goat anti-rabbit IgG conjugated to alkaline phosphatase. The chemiluminescent signal was recorded on a Fotodyne Luminary FX digital documentation system after incubation of the blot in CDP-Star. Bands representing the minor CP (min CP) and major CP (maj CP) of the virus are indicated. Coomassie staining of the same samples separated on a gel run in parallel provided a gel-loading standard via the large subunit of ribulose bisphosphate carboxylase (RBC). Additional Fig. 2S (EPS 1020 kb)

References

  1. 1.
    A.J. Gibbs, A.-L. Haenni, I. Jupin, G.P. Martelli, M.C. Edwards, R. Koenig, R.W. Hammond, S. Sabanadzovic, T.W. Dreher, in Virus Taxonomy, Ninth Report International Committee on the Taxonomy of Viruses, ed. by A. King, M. Adams, E. Carstens, E. Lefkowitz (Elsevier, Oxford, 2011), p. 944Google Scholar
  2. 2.
    T.W. Dreher, Mol. Plant Pathol. 5, 367–375 (2004)CrossRefPubMedGoogle Scholar
  3. 3.
    M.C. Edwards, J.J. Weiland, J. Todd, L.R. Stewart, Phytopathology 105, 833–839 (2015)CrossRefPubMedGoogle Scholar
  4. 4.
    R.W. Hammond, P. Ramirez, Virology 282, 338–347 (2001)CrossRefPubMedGoogle Scholar
  5. 5.
    C.S. Bozarth, J.J. Weiland, T.W. Dreher, Virology 187, 124–130 (1992)CrossRefPubMedGoogle Scholar
  6. 6.
    C.H. Tsai, T.W. Dreher, Mol. Plant Microbe Interact. 6, 268–273 (1993)CrossRefPubMedGoogle Scholar
  7. 7.
    P.K. Keese, A. Gibbs, Proc. Natl. Acad. Sci. U.S.A. 89, 9489–9493 (1992)CrossRefPubMedPubMedCentralGoogle Scholar
  8. 8.
    N. Sabath, A. Wagner, D. Karlin, Mol. Biol. Evol. 29, 3767–3780 (2012)CrossRefPubMedPubMedCentralGoogle Scholar
  9. 9.
    B.O. Agindotan, M.E. Gray, R.W. Hammond, C.A. Bradley, Arch. Virol. 157, 1825–1830 (2012)CrossRefPubMedGoogle Scholar
  10. 10.
    W. Maccheroni, M.C. Alegria, C.C. Greggio, J.P. Piazza, R.F. Kamla, P.R.A. Zacharias, M. Bar-Joseph, E.W. Kitajima, L.C. Assumpcão, G. Camarotte, J. Cardozo, E.C. Casagrande, F. Ferrari, S.F. Franco, P.F. Giachetto, A. Girasol, H. Jordão Jr, V.H.A. Silva, L.C.A. Souza, C.I. Aguilar-Vildoso, A.S. Zanca, P. Arruda, J.P. Kitajima, F.C. Reinach, J.A. Ferro, A.C.R. da Silva, J. Virol. 79, 3028–3037 (2005)CrossRefPubMedPubMedCentralGoogle Scholar
  11. 11.
    C.T. Ranjith-Kumar, K. Gopinath, A.N.K. Jacob, V. Srividhya, P. Elango, H.S. Savithri, Arch. Virol. 143, 1489–1500 (1998)CrossRefPubMedGoogle Scholar
  12. 12.
    A.T. Segwagwe, M.L. Putnam, K.L. Druffel, H.R. Pappu, K.C. Eastwell, Arch. Virol. 153, 1495–1503 (2008)CrossRefPubMedGoogle Scholar
  13. 13.
    M.C. Edwards, J.J. Weiland, Virology 450–451, 290–296 (2014)CrossRefPubMedGoogle Scholar
  14. 14.
    J.J. Weiland, M.C. Edwards, Eur. J. Plant Pathol. 131, 553–558 (2011)CrossRefGoogle Scholar
  15. 15.
    E.W. Kitajima, R. Gámez, Intervirology 19, 129–134 (1983)CrossRefPubMedGoogle Scholar
  16. 16.
    L.R. Nault, R.E. Gingery, D.T. Gordon, Phytopathology 70, 709–712 (1980)CrossRefGoogle Scholar
  17. 17.
    C. Rivera, R. Gámez, Intervirology 25, 76–82 (1986)CrossRefPubMedGoogle Scholar
  18. 18.
    W.J. Lucas, Virology 344, 169–184 (2006)CrossRefPubMedGoogle Scholar
  19. 19.
    H.B. Scholthof, Trends Plant Sci. 10, 376–382 (2005)CrossRefPubMedGoogle Scholar
  20. 20.
    J. Chen, W.X. Li, D. Xie, J.R. Peng, S.W. Ding, Plant Cell 16, 1302–1313 (2004)CrossRefPubMedPubMedCentralGoogle Scholar
  21. 21.
    J.M. O’Leary, C.M. Radcliffe, A.C. Willis, R.A. Dwek, P.M. Rudd, A.K. Downing, Protein Expr. Purif. 38, 217–227 (2004)CrossRefPubMedGoogle Scholar
  22. 22.
    S. Sabanadzovic, N. Abou Ghanem-Sabanadzovic, P. Saldarelli, G.P. Martelli, J. Gen. Virol. 82, 2009–2015 (2001)CrossRefPubMedGoogle Scholar
  23. 23.
    R.W. Fulton, J.L. Fulton, Phytopathology 70, 321–324 (1980)CrossRefGoogle Scholar
  24. 24.
    B.G. Bradel, W. Preil, H. Jeske, Virology 271, 289–297 (2000)CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media New York (Outside USA) 2016

Authors and Affiliations

  • Michael C. Edwards
    • 1
  • John J. Weiland
    • 1
  • Jane Todd
    • 2
  • Lucy R. Stewart
    • 2
  • Shunwen Lu
    • 1
  1. 1.United States Department of Agriculture – Agricultural Research Service, Cereal Crops Research UnitFargoUSA
  2. 2.United States Department of Agriculture – Agricultural Research Service Corn, Soybean, and Wheat Quality Research UnitWoosterUSA

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