Title Packaging of Porcine Reproductive and Respiratory Syndrome Virus Replicon RNA by a Stable Cell Line Expressing Its Nucleocapsid Protein
Author Byung-Hak Song1, Jeong-Min Kim1, Jin-Kyoung Kim1, Han-Saem Jang1, Gil-Nam Yun1, Eun-Jin Choi2, Jae-Young Song2, Sang-Im Yun1, and Young-Min Lee1*
Address 1Department of Microbiology, College of Medicine and Medical Research Institute, Chungbuk National University, Cheongju 361-763, Republic of Korea, 2National Veterinary Research and Quarantine Service, Ministry for Food, Agriculture, Forestry, and Fisheries, Anyang 430-757, Republic of Korea
Bibliography Journal of Microbiology, 49(3),516-523, 2011,
Key Words porcine reproductive and respiratory syndrome virus, viral replicon, nucleocapsid, stable cell line, trans-complementation
Abstract Porcine reproductive and respiratory syndrome virus (PRRSV), a member of the Arteriviridae family, is one of the most common and economically important swine pathogens. Although both live-attenuated and killed-inactivated vaccines against the virus have been available for a decade, PRRSV is still a major problem in the swine industry worldwide. To explore the possibility of producing single-round infectious PRRSV replicon particles as a potential vaccine strategy, we have now generated two necessary components: 1) a stable cell line (BHK/Sinrep19/PRRSV-N) that constitutively expresses the viral nucleocapsid (N) protein localized to the cytoplasm and the nucleolus and 2) a PRRSV replicon vector (pBAC/PRRSV/Replicon-ΔN) with a 177-nucleotide deletion, removing the 3′-half portion of ORF7 in the viral genome, from which the self-replicating propagation-defective replicon RNAs were synthesized in vitro by SP6 polymerase run-off transcription. Transfection of this replicon RNA into N protein-expressing BHK-21 cells led to the secretion of infectious particles that packaged the replicon RNA, albeit with a low production efficiency of 0.4×102 to 1.1×102 infectious units/ml; the produced particles had only single-round infectivity with no cell-to-cell spread. This trans-complementation system for PRRSV provides a useful platform for studies to define the packaging signals and motifs present within the viral genome and N protein, respectively, and to develop viral replicon-based antiviral vaccines that will stop the infection and spread of this pathogen.