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Archives of Virology

, Volume 159, Issue 8, pp 1927–1940 | Cite as

Genetic manipulation of a transcription-regulating sequence of porcine reproductive and respiratory syndrome virus reveals key nucleotides determining its activity

  • Haihong Zheng
  • Keyu Zhang
  • Xing-Quan Zhu
  • Changlong Liu
  • Jiaqi Lu
  • Fei Gao
  • Yan Zhou
  • Hao Zheng
  • Tao Lin
  • Liwei Li
  • Guangzhi Tong
  • Zuzhang Wei
  • Shishan Yuan
Original Article

Abstract

The factors that determine the transcription-regulating sequence (TRS) activity of porcine reproductive and respiratory syndrome virus (PRRSV) remain largely unclear. In this study, the effect of mutagenesis of conserved C nucleotides at positions 5 and 6 in the leader TRS (TRS-L) and/or canonical body TRS7 (TRS-B7) on the synthesis of subgenomic (sg) mRNA and virus infectivity was investigated in the context of a type 2 PRRSV infectious cDNA clone. The results showed that a double C mutation in the leader TRS completely abolished sg mRNAs synthesis and virus infectivity, but a single C mutation did not. A single C or double C mutation in TRS-B7.1 or/and TRS-B7.2 impaired or abolished the corresponding sg mRNA synthesis. Introduction of identical mutations in the leader and body TRSs partially restored sg mRNA7.1 and/or sg mRNA7.2 transcription, indicating that the base-pairing interaction between sense TRS-L and cTRS-B is a crucial factor influencing sg mRNA synthesis. Analysis of the mRNA leader-body junctions of mutants provided evidence for a mechanism of discontinuous minus-strand transcription. This study also showed that mutational inactivation of TRS-B7.1 or TRS-B7.2 did not affect the production of infectious progeny virus, and the sg mRNA formed from each of them could express N protein. However, TRS-B7.1 plays more important roles than TRS-B7.2 in maintaining the growth characteristic of type 2 PRRSV. These results provide more insight into the molecular mechanism of genome expression and subgenomic mRNA transcription of PRRSV.

Keywords

Mutant Virus Shuttle Plasmid Equine Arteritis Virus Infectious Progeny Virus Discontinuous Transcription 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Notes

Acknowledgements

This study was supported from the National Natural Science Foundation of China (30972204, 31372444) and the EU Seventh Framework Program (No. 245141).

Supplementary material

705_2014_2018_MOESM1_ESM.docx (24 kb)
Supplementary material 1 (DOC 72 kb)

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Copyright information

© Springer-Verlag Wien 2014

Authors and Affiliations

  • Haihong Zheng
    • 1
  • Keyu Zhang
    • 2
  • Xing-Quan Zhu
    • 3
  • Changlong Liu
    • 1
  • Jiaqi Lu
    • 1
  • Fei Gao
    • 1
  • Yan Zhou
    • 1
  • Hao Zheng
    • 1
  • Tao Lin
    • 1
  • Liwei Li
    • 1
  • Guangzhi Tong
    • 1
  • Zuzhang Wei
    • 1
    • 4
  • Shishan Yuan
    • 1
  1. 1.Department of Swine Infectious Diseases, Shanghai Veterinary Research InstituteChinese Academy of Agricultural SciencesShanghaiChina
  2. 2.Key Laboratory of Veterinary Drug Safety Evaluation and Residues Research, Shanghai Veterinary Research InstituteChinese Academy of Agricultural SciencesShanghaiChina
  3. 3.State Key Laboratory of Veterinary Etiological Biology, Lanzhou Veterinary Research InstituteChinese Academy of Agricultural SciencesLanzhouChina
  4. 4.College of Animal Science and TechnologyGuangxi UniversityNanningChina

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