Characterization of the interactome of the porcine reproductive and respiratory syndrome virus glycoprotein-5
Porcine reproductive and respiratory syndrome virus (PRRSV) is one of the most important pathogens in the swine industry, causing reproductive failure in sows and respiratory disorders in piglets. Glycosylated protein 5 (GP5) is a major envelope protein of the virus. It is essential for virus particle assembly and involved in viral pathogenesis. In the present study, we identified the host cellular proteins that interact with GP5 by performing immunoprecipitation in MARC-145 cells infected by a recombinant PRRSV containing a FLAG-tag insertion in GP5. In total, 122 cellular proteins were identified by LC-MS/MS. Gene Ontology and KEGG databases were used to map these proteins to different cellular processes, locations and functions. Interestingly, 10.24% of identified cellular proteins were involved in the process of translation. Follow up experiments demonstrated that expression of GP5 in transfected cells led to inhibition of translation of reporter genes. Interaction between GP5 and ATP synthase subunit alpha (ATP5A) was further confirmed by co-immunoprecipitation suggesting a possible role of GP5 in regulation of ATP production in cells. These data contribute to a better understanding of GP5’s role in viral pathogenesis and virus-host interactions.
We thank Dr. X-J Meng (Virginia Tech) for supplying us with plasmid pIR-VR2385-CA. Also, we are thankful to Dr. Aviad Levin (University of Alberta) for designing a peptide for rabbit immunization, and Dr. Samuel Attah-Poku for making peptide conjugates. We acknowledge the Proteomics Center at the University of Missouri-Columbia for mass spectrometry analysis, and Dr. Aaron White for critical reading of the manuscript. This paper was published with the permission of the Director of VIDO-InterVac, journal series no. 817.
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflict of interest. The study was funded in part by funding from the University of Saskatchewan President’s NSERC fund grant (A.Z. is a principal investigator), and M.Z. is supported by a China Scholarship Council.
Research involving human participants and/or animals
This article does not contain any studies with human participants. Work with animals was approved by the University of Saskatchewan’s Animal Research Ethics Board and adhered to the Canadian Council on Animal Care guidelines for humane animal use.
Informed consent was not obtained since human participants were not involved in the study.
- 1.Adams MJ, Lefkowitz EJ, King AM, Harrach B, Harrison RL, Knowles NJ, Kropinski AM, Krupovic M, Kuhn JH, Mushegian AR, Nibert M, Sabanadzovic S, Sanfacon H, Siddell SG, Simmonds P, Varsani A, Zerbini FM, Gorbalenya AE, Davison AJ (2016) Ratification vote on taxonomic proposals to the International Committee on Taxonomy of Viruses (2016). Arch Virol 161:2921–2949CrossRefPubMedGoogle Scholar
- 2.Cervantes-Salazar M, Angel-Ambrocio AH, Soto-Acosta R, Bautista-Carbajal P, Hurtado-Monzon AM, Alcaraz-Estrada SL, Ludert JE, Del Angel RM (2015) Dengue virus NS1 protein interacts with the ribosomal protein RPL18: this interaction is required for viral translation and replication in Huh-7 cells. Virology 484:113–126CrossRefPubMedGoogle Scholar
- 6.Dong S, Liu L, Wu W, Armstrong SD, Xia D, Nan H, Hiscox JA, Chen H (2016) Determination of the interactome of non-structural protein12 from highly pathogenic porcine reproductive and respiratory syndrome virus with host cellular proteins using high throughput proteomics and identification of HSP70 as a cellular factor for virus replication. J Proteom 146:58–69CrossRefGoogle Scholar
- 8.Gao J, Ji P, Zhang M, Wang X, Li N, Wang C, Xiao S, Mu Y, Zhao Q, Du T, Sun Y, Hiscox JA, Zhang G, Zhou EM (2014) GP5 expression in Marc-145 cells inhibits porcine reproductive and respiratory syndrome virus infection by inducing beta interferon activity. Vet Microbiol 174:409–418CrossRefPubMedGoogle Scholar
- 9.Gao J, Xiao S, Xiao Y, Wang X, Zhang C, Zhao Q, Nan Y, Huang B, Liu H, Liu N, Lv J, Du T, Sun Y, Mu Y, Wang G, Syed SF, Zhang G, Hiscox JA, Goodfellow I, Zhou EM (2016) MYH9 is an essential factor for porcine reproductive and respiratory syndrome virus infection. Sci Rep 6:25120CrossRefPubMedPubMedCentralGoogle Scholar
- 14.Liu L, Lear Z, Hughes DJ, Wu W, Zhou EM, Whitehouse A, Chen H, Hiscox JA (2015) Resolution of the cellular proteome of the nucleocapsid protein from a highly pathogenic isolate of porcine reproductive and respiratory syndrome virus identifies PARP-1 as a cellular target whose interaction is critical for virus biology. Vet Microbiol 176:109–119CrossRefPubMedPubMedCentralGoogle Scholar
- 17.Mu Y, Li L, Zhang B, Huang B, Gao J, Wang X, Wang C, Xiao S, Zhao Q, Sun Y, Zhang G, Hiscox JA, Zhou EM (2015) Glycoprotein 5 of porcine reproductive and respiratory syndrome virus strain SD16 inhibits viral replication and causes G2/M cell cycle arrest, but does not induce cellular apoptosis in Marc-145 cells. Virology 484:136–145CrossRefPubMedGoogle Scholar
- 20.Ni YY, Huang YW, Cao D, Opriessnig T, Meng XJ (2011) Establishment of a DNA-launched infectious clone for a highly pneumovirulent strain of type 2 porcine reproductive and respiratory syndrome virus: identification and in vitro and in vivo characterization of a large spontaneous deletion in the nsp2 region. Virus Res 160:264–273CrossRefPubMedGoogle Scholar
- 35.Xiao Y, Wu W, Gao J, Smith N, Burkard C, Xia D, Zhang M, Wang C, Archibald A, Digard P, Zhou EM, Hiscox JA (2016) Characterization of the interactome of the porcine reproductive and respiratory syndrome virus nonstructural protein 2 reveals the hyper variable region as a binding platform for association with 14-3-3 proteins. J Proteome Res 15:1388–1401CrossRefPubMedGoogle Scholar