Expression and stability of foreign tags inserted into nsp2 of porcine reproductive and respiratory syndrome virus (PRRSV)

Kim, Dal–Young; Calvert, Jay G.; Chang, Kyung-Ro; Horlen, Kyle; Kerrigan, Maureen; Rowland, Raymond R. R.

doi:10.1016/j.virusres.2007.04.019

Cited by 55 publications

(57 citation statements)

References 29 publications

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“…Meanwhile, Nsp2 has become one of the vital regions for monitoring the evolution of PRRSV and for molecular epidemiology research on PRRSV (2,10,12,13,16,35,47). Recently, mutations in the Nsp2-coding region resulting in a 403-aa manual deletion and expression of a foreign protein were reported (11,15,20). Therefore, further investigations are required, particularly on the role of Nsp2 in the cell and tissue tropism, replication and growth, and variation and pathogenicity of PRRSV, as well as in the difference in virulence among various strains.…”

Section: Discussionmentioning

confidence: 99%

The 30-Amino-Acid Deletion in the Nsp2 of Highly Pathogenic Porcine Reproductive and Respiratory Syndrome Virus Emerging in China Is Not Related to Its Virulence

Zhou

Zhang

Zeng

et al. 2009

J Virol

244

201

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During the past 2 years, an atypical clinical outbreak, caused by a highly pathogenic porcine reproductive and respiratory syndrome virus (PRRSV) with a unique 30-amino-acid deletion in its Nsp2-coding region, was pandemic in China. In this study, we generated four full-length infectious cDNA clones: a clone of the highly virulent PRRSV strain JXwn06 (pWSK-JXwn), a clone of the low-virulence PRRSV strain HB-1/3.9 (pWSK-HB-1/3.9), a chimeric clone in which the Nsp2 region containing the 30-amino-acid deletion was replaced by the corresponding region of the low-virulence PRRSV strain HB-1/3.9 (pWSK-JXwn-HB1nsp2), and a mutated HB-1/3.9 clone with the same deletion in Nsp2 as JXwn06 (pWSK-HB1-ND30). We also investigated the pathogenicities of the rescued viruses (designated RvJXwn, RvJXwn-HB1nsp2, RvHB-1/3.9, and RvHB1-ND30, respectively) in specific-pathogen-free piglets in order to determine the role of the 30-amino-acid deletion in the virulence of the highly pathogenic PRRSV. All the rescued viruses could replicate stably in MARC-145 cells. Our findings indicated that RvJXwn-HB1nsp2 retained high virulence for piglets, like RvJXwn and the parental virus JXwn06, although the survival time of piglets infected with RvJXwn-HB1nsp2 was obviously prolonged. RvHB1-ND30 exhibited low virulence for piglets, like RvHB-1/3.9 and the parental virus HB-1/3.9. Therefore, we conclude that the 30-amino-acid deletion is not related to the virulence of the highly pathogenic PRRSV emerging in China.

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Section: Discussionmentioning

confidence: 99%

The 30-Amino-Acid Deletion in the Nsp2 of Highly Pathogenic Porcine Reproductive and Respiratory Syndrome Virus Emerging in China Is Not Related to Its Virulence

Zhou

Zhang

Zeng

et al. 2009

J Virol

244

201

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“…Previous studies proved that PRRSV could tolerate the addition of foreign genes, which were fused to ORF1b, ORF7, or Nsp2 [27][28][29][30][31]. However, insertion of extra nonviral genes has reduced levels of replication or resulted in genetic instability such that the recombinant viruses no longer encode a functional foreign gene after serial passage.…”

Section: Discussionmentioning

confidence: 99%

Construction and in vitro evaluation of a recombinant live attenuated PRRSV expressing GM-CSF

Zhou

Jiang

et al. 2014

Virology Journal

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Background: Porcine reproductive and respiratory syndrome virus (PRRSV) continues to be an important problem for the swine industry. Inactivated vaccines and modified-live virus vaccines are widely used in the field; however, the efficacy of these PRRSV vaccines is suboptimal due to poor immunogenicity. Granulocyte-macrophage colony stimulating factor (GM-CSF) has been extensively used as an effective genetic and protein adjuvant to enhance the efficiencies vaccines expressing tumor or pathogen antigens. The purpose of this study was to determine if GM-CSF could increase the efficiency of PRRSV vaccine. Methods: The GM-CSF gene was inserted in the HuN4-F112 vaccine strain by overlap PCR. The expression of GM-CSF by the recombinant virus was confirmed with methods of indirect immunofluorescent assay (IFA) and Western blotting. The stability of recombinant virus was assessed by cDNA sequence and IFA after 20 passages. To detect the biological activity of GM-CSF expressed by the recombinant virus, bone marrow-derived dendritic cells (BMDCs) were isolated and co-cultured with the recombinant virus or parental virus and the surface phenotypes of BMDCs were examined by flow cytometric analysis. The cytokines secreted by BMDCs infected with PRRSV, or treated with LPS, GM-CSF or medium alone were evaluated by ProcartaPlexTM Multiplex Immunoassays and qRT-PCR. Results: A novel modified-live PRRSV vaccine strain expressing GM-CSF (rHuN4-GM-CSF) was successfully constructed and rescued. The GM-CSF protein was stable expressed in recombinant virus-infected cells after 20 passages. Analysis of virus replication kinetics showed that the novel vaccine strain expressing GM-CSF had a similar replication rate as the parental virus. In vitro studies showed that infection of porcine BMDCs with rHuN4-GM-CSF resulted in increased surface expression of MHCI+, MHCII + and CD80/86+ that was dependent on virus expressed GM-CSF. The expression of representative cytokines was significantly up-regulated when BMDCs were incubated with the recombinant GM-CSF expressing virus.

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“…The causative agent, porcine reproductive and respiratory syndrome virus (PRRSV), is an enveloped, single-strand positive RNA virus which is a member of the genus Arterivirus, family Arteriviridae, order Nidovirales (2). In 2006, a highly pathogenic strain of PRRSV (HP-PRRSV) with a discontinuous 30-amino-acid depletion in the Nsp2 protein was identified in China (3,4). The atypical PRRS caused by HP-PRRSV was characterized by high fever (Ͼ41°C for at least 4 days), high morbidity, and high mortality in pigs of all ages.…”

Section: Importancementioning

confidence: 99%

Highly Pathogenic Porcine Reproductive and Respiratory Syndrome Virus Induces Prostaglandin E ₂ Production through Cyclooxygenase 1, Which Is Dependent on the ERK1/2-p-C/EBP-β Pathway

Guo

Zhao

et al. 2014

J Virol

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Atypical porcine reproductive and respiratory syndrome (PRRS) caused by highly pathogenic porcine reproductive and respiratory syndrome virus (HP-PRRSV) is characterized by high fever and high mortality. However, the mechanism underlying the fever induction is still unknown. Prostaglandin E 2 (PGE 2 ), synthesized by cyclooxygenase type 1/2 (COX-1/2) enzymes, is essential for inducing fever. In this study, we found that PGE 2 , together with COX-1, was significantly elevated by HP-PRRSV. We subsequently demonstrated that extracellular signal-regulated kinase 1/2 (ERK1/2) and phosphorylated ERK (p-ERK) were the key nodes to trigger COX-1 expression after HP-PRRSV infection. Furthermore, we proved the direct binding of p-C/EBP-␤ to the COX-1 promoter by luciferase reporter and chromatin immunoprecipitation assays. In addition, silencing of C/EBP-␤ remarkably impaired the enhancement of COX-1 production induced by HP-PRRSV infection. Taken together, our results indicate that HP-PPRSV elicits the expression of COX-1 through the ERK1/2-p-C/EBP-␤ signaling pathway, resulting in the increase of PGE 2 , which might be the cause of high fever in infected pigs. Our findings might provide new insights into the molecular mechanisms underlying the pathogenesis of HP-PRRSV infection. IMPORTANCEThe atypical PRRS caused by HP-PRRSV was characterized by high fever, high morbidity, and high mortality in pigs of all ages, yet how HP-PRRSV induces high fever in pigs remains unknown. In the present study, we found out that HP-PRRSV infection could increase PGE 2 production by upregulation of COX-1, and we subsequently characterized the underlying mechanisms about how HP-PRRSV enhances COX-1 production. PGE 2 plays a critical role in inducing high temperature in hosts during pathogen infections. Thus, our findings here could help us have a better understanding of HP-PRRSV pathogenesis.

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Expression and stability of foreign tags inserted into nsp2 of porcine reproductive and respiratory syndrome virus (PRRSV)

Cited by 55 publications

References 29 publications

The 30-Amino-Acid Deletion in the Nsp2 of Highly Pathogenic Porcine Reproductive and Respiratory Syndrome Virus Emerging in China Is Not Related to Its Virulence

The 30-Amino-Acid Deletion in the Nsp2 of Highly Pathogenic Porcine Reproductive and Respiratory Syndrome Virus Emerging in China Is Not Related to Its Virulence

Construction and in vitro evaluation of a recombinant live attenuated PRRSV expressing GM-CSF

Highly Pathogenic Porcine Reproductive and Respiratory Syndrome Virus Induces Prostaglandin E ₂ Production through Cyclooxygenase 1, Which Is Dependent on the ERK1/2-p-C/EBP-β Pathway

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Expression and stability of foreign tags inserted into nsp2 of porcine reproductive and respiratory syndrome virus (PRRSV)

Cited by 55 publications

References 29 publications

The 30-Amino-Acid Deletion in the Nsp2 of Highly Pathogenic Porcine Reproductive and Respiratory Syndrome Virus Emerging in China Is Not Related to Its Virulence

The 30-Amino-Acid Deletion in the Nsp2 of Highly Pathogenic Porcine Reproductive and Respiratory Syndrome Virus Emerging in China Is Not Related to Its Virulence

Construction and in vitro evaluation of a recombinant live attenuated PRRSV expressing GM-CSF

Highly Pathogenic Porcine Reproductive and Respiratory Syndrome Virus Induces Prostaglandin E 2 Production through Cyclooxygenase 1, Which Is Dependent on the ERK1/2-p-C/EBP-β Pathway

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Highly Pathogenic Porcine Reproductive and Respiratory Syndrome Virus Induces Prostaglandin E ₂ Production through Cyclooxygenase 1, Which Is Dependent on the ERK1/2-p-C/EBP-β Pathway