title: Immunocompetent truncated E2 glycoprotein of bovine viral diarrhea virus (BVDV) expressed in Nicotiana tabacum plants: a candidate antigen for new generation of veterinary vaccines.
abstract: The bovine viral diarrhea virus (BVDV) is the etiological agent responsible for a wide spectrum of clinical diseases in cattle. The glycoprotein E2 is the major envelope protein of this virus and the strongest inductor of the immune response. There are several available commercial vaccines against bovine viral diarrhea (BVD), which show irregular performances. Here, we report the use of tobacco plants as an alternative productive platform for the expression of the truncated version of E2 glycoprotein (tE2) from the BVDV. The tE2 sequence, lacking the transmembrane domain, was cloned into the pK7WG2 Agrobacterium binary vector. The construct also carried the 2S2 Arabidopsis thaliana signal for directing the protein into the plant secretory pathway, the Kozak sequence, an hexa-histidine tag to facilitate protein purification and the KDEL endoplasmic reticulum retention signal. The resulting plasmid (pK-2S2-tE2-His-KDEL) was introduced into Agrobacterium tumefaciens strain EHA101 by electroporation. The transformed A. tumefaciens was then used to express tE2 in leaves of Nicotiana tabacum plants. Western blot and ELISA using specific monoclonal antibodies confirmed the presence of the recombinant tE2 protein in plant extracts. An estimated amount of 20 μg of tE2 per gram of fresh leaves was regularly obtained with this plant system. Injection of guinea pigs with plant extracts containing 20 μg of rtE2 induced the production of BVDV specific antibodies at equal or higher levels than those induced by whole virus vaccines. This is the first report of the production of an immunocompetent tE2 in N. tabacum plants, having the advantage to be free of any eventual animal contaminant.
authors.affiliation: Instituto de Ciencia y Tecnología Dr. César Milstein, CONICET - Fundación Pablo Cassará, Saladillo 2468, Ciudad de Buenos Aires, C11440FFX, Argentina.
authors.forename: Guillermo
authors.initials: G
authors.lastname: Nelson
authors.forename: Patricia
authors.initials: P
authors.lastname: Marconi
authors.forename: Osvaldo
authors.initials: O
authors.lastname: Periolo
authors.forename: José
authors.initials: J
authors.lastname: La Torre
authors.forename: María Alejandra
authors.initials: MA
authors.lastname: Alvarez
mesh_terms: D060054:Agrobacterium; D000818:Animals; D000914:Antibodies, Viral / Q000097:blood; D000956:Antigens, Viral / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology*; D001912:Bovine Virus Diarrhea-Mucosal Disease / Q000276:immunology / Q000517:prevention & control*; D002417:Cattle; D001908:Diarrhea Viruses, Bovine Viral / Q000235:genetics / Q000276:immunology; D005260:Female; D005818:Genetic Engineering / Q000379:methods; D006168:Guinea Pigs; D009500:Neutralization Tests; D030821:Plants, Genetically Modified / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D014026:Nicotiana / Q000378:metabolism*; D014759:Viral Envelope Proteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology*; D014765:Viral Vaccines / Q000276:immunology*
chemical_list: D000914:Antibodies, Viral; D000956:Antigens, Viral; D011994:Recombinant Proteins; D014759:Viral Envelope Proteins; D014765:Viral Vaccines