The detailed mechanism(s) by which porcine reproductive and respiratory syndrome virus (PRRSV) impairs alveolar Mφ homeostasis and function remains to be elucidated. We used differential display reverse-transcription PCR (DDRT-PCR) to identify molecular genetic changes within PRRSV-infected Mφ over a 24 h post infection period. From over 4000 DDRT-PCR amplicons examined, 19 porcine-derived DDRT-PCR products induced by PRRSV were identified and cloned. Northern blot analysis confirmed that four gene transcripts were induced during PRRSV infection. PRRSV attachment and penetration alone did not induce these gene transcripts. DNA sequence revealed that one PRRSV-induced expressed sequence tag (EST) encoded porcine Mx1, while the remaining 3 clones represented novel ESTs. A full-length cDNA clone for EST G3V16 was obtained from a porcine blood cDNA library. Sequence data suggests that it encodes an ubiquitin-specific protease (UBP) that regulates protein trafficking and degradation. In pigs infected in vivo, upregulated transcript levels were observed for Mx1 and Ubp in lung and tonsils, and for Mx1 in tracheobronchial lymph node (TBLN). These tissues correspond to sites for PRRSV persistence, suggesting that the Mxl and Ubp genes may play important roles in clinical disease during PRRSV infection.
Bibliographical noteFunding Information:
This research was supported by the National Pork and Producers Council (M.S.R), the Minnesota Pork Producers Association (T.W.M), the U.S.D.A. grant 95–3205-3846 (L.B.S.), and the University of Minnesota Agricultural Experiment Station (M.S.R.). The authors thank Drs. M. P. Murtaugh and K. S. Faaberg for supplying the PRRSV VR2332 strain, ORF7 PCR primers, and CL2621 cell culture, and Dr. C. W. Beattie for porcine cDNA library. The authors further acknowledge Dr. J. E. Collins for assistance with experimental design, and Drs. A. Bhattacharjee and A. Rink for technical advice on the DDRT-PCR assays and cDNA library screening.
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