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ARS Home » Midwest Area » Ames, Iowa » National Animal Disease Center » Virus and Prion Research » Research » Publications at this Location » Publication #251478
Title: Porcine Reproductive and Respiratory Syndrome Virus Replicase - Isoforms of Nonstructural Protein 2 and Interaction with Heat Shock 70kDa Protein 5

Author
item Faaberg, Kay
item HAN, JUN - University Of Minnesota
item RUTHERFORD, MARK - University Of Minnesota

Submitted to: International Positive Strand RNA Virus Symposium
Publication Type: Abstract Only
Publication Acceptance Date: 3/26/2010
Publication Date: 5/17/2010
Citation: Faaberg, K.S., Han, J., Rutherford, M.S. 2010. Porcine reproductive and respiratory syndrome virus replicase - isoforms of nonstructural protein 2 and interaction with heat shock 70kDa protein 5 [abstract]. Ninth International Positive Strand RNA Virus Symposium, Atlanta, GA. Paper No. P1-F16.

Interpretive Summary:

Technical Abstract: The nsp2 replicase protein of porcine reproductive and respiratory syndrome virus (PRRSV), when expressed independently, was recently demonstrated to be processed from its precursor by the PL2 protease at or near the G**1196|G**1197 dipeptide in transfected CHO cells. The proteolytic cleavage of nsp2 was further investigated in virally infected MARC-145 cells by the use of two recombinant viruses expressing epitope-tagged nsp2. The data revealed that nsp2 exists as different isoforms during PRRSV infection, termed nsp2a, b, c, d, e and f. Moreover, these nsp2 species appeared to share the same N-terminus but differed in their respective C-termini based on deletion mutagenesis and antibody probing. The largest protein, nsp2a, corresponded to the nsp2 product identified in transfected CHO cells. Nsp2b and c were processed within or near the TM region, presumably at or near the conserved sites G**981|G**982 and G**828|G**829|G**830, respectively. The C-termini for nsp2d, e and f were mapped within the nsp2 middle hypervariable region, but no conserved cleavage sites could be predicted, suggesting involvement of other viral or host cell proteases. The larger nsp2 species emerged almost simultaneously in the early stage of PRRSV infection. Pulse-chase analysis revealed that all six nsp2 species were relatively stable and had low turnover rates. Deletion mutagenesis revealed that the smaller nsp2 species (e.g. nsp2d, e and f) were not essential for viral replication in cell culture. Lastly, we identified a cellular chaperone, heat shock 70kDa protein 5 (HSPA5), that was strongly associated with nsp2 and which may have important implications for PRRSV replication. Overall, these findings indicate that the PRRSV nsp2 is increasingly emerging as a multifunctional protein and may have a profound impact on PRRSV replication and viral pathogenesis.