Virus Genes

, Volume 54, Issue 4, pp 550–560 | Cite as

Functional characterization of the bovine foamy virus miRNA expression cassette and its dumbbell-shaped pri-miRNA

  • Wenhu Cao
  • Anke Heit
  • Agnes Hotz-Wagenblatt
  • Martin Löchelt


Foamy viruses are unconventional and complex retroviruses distinct from the other members of the Retroviridae family. Currently, no disease has been firmly linked to persistent foamy virus infection of their cognate host including simians, bovines, felines, and equines or upon zoonotic transmission of different simian foamy viruses to humans. Bovine and simian foamy viruses have been recently shown to encode a RNA polymerase-III-driven micro RNA cluster which likely modulates and regulates host–virus interactions at different levels. Using sub-genomic bovine foamy virus micro RNA expression plasmids and dual luciferase reporter assays as readout, the requirements for expression and processing of the bovine foamy virus micro RNAs have been analyzed. Here, we report that the minimal BFV micro RNA cassette is significantly weaker than a U6 promoter-based construct and strongly suppressed by flanking sequences. The primary micro RNA sequence can be manipulated and chimerized as long as the dumbbell-like folding of the primary micro RNA is maintained. Since more subtle changes are associated with reduced functionality, the overall structure and shape, but possibly individual elements and residues also, are important for the expression and processing of the bovine foamy virus micro RNAs.


Bovine foamy virus MiRNA BFV Spumaretrovirus Gene expression and regulation 



We thank the Baden-Württemberg Stiftung for financing these studies by the research Grant SID 49. We thank Bryan Cullen, Duke University, Durham, USA for thoughful discussions, Dirk Grimm, BioQuant, Heidelberg, Germany for providing the U6 promoter-containing plasmid pBS–U6-GFP plasmid and the German Cancer Research Center Core Facility Genomics and Proteomics for sequencing of miRNAs.

Author contributions

WC performed and analyzed the experiments, AH and AH-W evaluated sequencing data and conducted pri-miRNA structural analyses, and ML designed and supervised the study and wrote the manuscript. All authors read and approved the final manuscript.


The funding bodies played no role in the study design, data collection or analysis, or writing of the manuscript.

Compliance with ethical standards

Conflict of interest

The authors declare that there are no conflicts of interest.

Ethical approval

This article does not include any studies with human participants or animals performed by any of the authors.

Informed consent

Not applicable as no human participants are involved.


  1. 1.
    A.S. Khan, J. Bodem, F. Buseyne, A. Gessain, W. Johnson, J.H. Kuhn, J. Kuzmak, D. Lindemann, M.L. Linial, M. Löchelt, M. Materniak-Kornas, M.A. Soares, W.M. Switzer, Virology 516, 158–164 (2018)CrossRefPubMedGoogle Scholar
  2. 2.
    D. Lindemann, A. Rethwilm, Viruses 3, 561–585 (2011)CrossRefPubMedPubMedCentralGoogle Scholar
  3. 3.
    A. Rethwilm, Med. Microbiol. Immunol. 199, 197–207 (2010)CrossRefPubMedGoogle Scholar
  4. 4.
    T. Kehl, J. Tan, M. Materniak, Viruses 5, 2169–2209 (2013)CrossRefPubMedPubMedCentralGoogle Scholar
  5. 5.
    M. Linial, Trends Microbiol. 8, 284–289 (2000)CrossRefPubMedGoogle Scholar
  6. 6.
    A. Rethwilm, J. Bodem, Viruses 5, 2349–2374 (2013)CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    M. Löchelt, Curr. Top. Microbiol. Immunol. 277, 27–61 (2003)PubMedGoogle Scholar
  8. 8.
    M. Löchelt, F. Romen, P. Bastone, H. Muckenfuss, N. Kirchner, Y.B. Kim, U. Truyen, U. Rosler, M. Battenberg, A. Saib, E. Flory, K. Cichutek, C. Munk, Proc. Natl. Acad. Sci. USA 102, 7982–7987 (2005)CrossRefPubMedGoogle Scholar
  9. 9.
    M. Löchelt, S.F. Yu, M.L. Linial, R.M. Flugel, Virology 206, 601–610 (1995)CrossRefPubMedGoogle Scholar
  10. 10.
    B.R. Cullen, Proc. Natl. Acad. Sci. USA 109, 2695–2696 (2012)CrossRefPubMedGoogle Scholar
  11. 11.
    Z.A. Klase, G.C. Sampey, F. Kashanchi, Retrovirology 10, 15 (2013)CrossRefPubMedPubMedCentralGoogle Scholar
  12. 12.
    Y. Yao, L.P. Smith, V. Nair, M. Watson, J. Virol. 88, 2–9 (2014)CrossRefPubMedPubMedCentralGoogle Scholar
  13. 13.
    R.P. Kincaid, Y. Chen, J.E. Cox, A. Rethwilm, C.S. Sullivan, mBio 5, e00074 (2014)CrossRefPubMedPubMedCentralGoogle Scholar
  14. 14.
    A.W. Whisnant, T. Kehl, Q. Bao, M. Materniak, J. Kuzmak, M. Löochelt, B.R. Cullen, J. Virol. 88, 4679–4686 (2014)CrossRefPubMedPubMedCentralGoogle Scholar
  15. 15.
    J. Krol, I. Loedige, W. Filipowicz, Nat. Rev. Genet. 11, 597–610 (2010)CrossRefPubMedGoogle Scholar
  16. 16.
    M. Sand, Methods Mol. Biol. 1095, 3–10 (2014)CrossRefPubMedGoogle Scholar
  17. 17.
    N. Rosewick, M. Momont, K. Durkin, H. Takeda, F. Caiment, Y. Cleuter, C. Vernin, F. Mortreux, E. Wattel, A. Burny, M. Georges, A. Van den Broeke, Proc. Natl. Acad. Sci. USA 110, 2306–2311 (2013)CrossRefPubMedGoogle Scholar
  18. 18.
    B. Van Driessche, A. Rodari, N. Delacourt, S. Fauquenoy, C. Vanhulle, A. Burny, O. Rohr, C. Van Lint, Sci. Rep. 6, 31125 (2016)CrossRefPubMedPubMedCentralGoogle Scholar
  19. 19.
    Q. Bao, M. Hipp, A. Hugo, J. Lei, Y. Liu, T. Kehl, T. Hechler, M. Löchelt, Viruses 7, 5855–5874 (2015)CrossRefPubMedPubMedCentralGoogle Scholar
  20. 20.
    L. Dong, Q. Cheng, Z. Wang, P. Yuan, Z. Li, Y. Sun, S. Han, J. Yin, B. Peng, X. He, W. Liu, Viruses 7, 1668–1684 (2015)CrossRefPubMedPubMedCentralGoogle Scholar
  21. 21.
    T. Hechler, M. Materniak, T. Kehl, J. Kuzmak, M. Löchelt, J. Virol. 86, 10905–10906 (2012)CrossRefPubMedPubMedCentralGoogle Scholar
  22. 22.
    J. Lei, W. Osen, A. Gardyan, A. Hotz-Wagenblatt, G. Wei, L. Gissmann, S. Eichmuller, M. Löchelt, PLoS ONE 10, e0138458 (2015)CrossRefPubMedPubMedCentralGoogle Scholar
  23. 23.
    R. Lorenz, S.H. Bernhart, C. Honer Zu Siederdissen, H. Tafer, C. Flamm, P.F. Stadler, I.L. Hofacker, Algorithms Mol. Biol. 6, 26 (2011)CrossRefPubMedPubMedCentralGoogle Scholar
  24. 24.
    J. Wang, J. Tan, H. Guo, Q. Zhang, R. Jia, X. Xu, Y. Geng, W. Qiao, J. Virol. 84, 11888–11897 (2010)CrossRefPubMedPubMedCentralGoogle Scholar
  25. 25.
    G.R. Kunkel, R.L. Maser, J.P. Calvet, T. Pederson, Proc. Natl. Acad. Sci. USA 83, 8575–8579 (1986)CrossRefPubMedGoogle Scholar
  26. 26.
    J.M. Burke, R.P. Kincaid, F. Aloisio, N. Welch, C.S. Sullivan, Nucleic Acids Res. 45, e154 (2017)CrossRefPubMedPubMedCentralGoogle Scholar
  27. 27.
    J. Tan, P. Hao, R. Jia, W. Yang, R. Liu, J. Wang, Z. Xi, Y. Geng, W. Qiao, Virology 405, 408–413 (2010)CrossRefPubMedGoogle Scholar
  28. 28.
    J. Kimpton, M. Emerman, J. Virol. 66, 2232–2239 (1992)PubMedPubMedCentralGoogle Scholar
  29. 29.
    R.P. Kincaid, J.M. Burke, C.S. Sullivan, Proc. Natl. Acad. Sci. USA 109, 3077–3082 (2012)CrossRefPubMedGoogle Scholar
  30. 30.
    M.C. Frie, C.J. Droscha, A.E. Greenlick, P.M. Coussens, Front. Vet. Sci. 4, 245 (2017)CrossRefPubMedGoogle Scholar
  31. 31.
    K. Durkin, N. Rosewick, M. Artesi, V. Hahaut, P. Griebel, N. Arsic, A. Burny, M. Georges, A. Van den Broeke, Retrovirology 13, 33 (2016)CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  • Wenhu Cao
    • 1
  • Anke Heit
    • 2
  • Agnes Hotz-Wagenblatt
    • 2
  • Martin Löchelt
    • 1
  1. 1.Division Molecular Diagnostics of Oncogenic InfectionsResearch Focus Infection, Inflammation and CancerHeidelbergGermany
  2. 2.Core Facility Omics IT and Data ManagementGerman Cancer Research Center (Deutsches Krebsforschungszentrum, DKFZ)HeidelbergGermany

Personalised recommendations