title: Expression and immunogenicity of the spike glycoprotein of porcine respiratory coronavirus encoded in the E3 region of adenovirus.
abstract: The full length spike (S) gene of porcine respiratory coronavirus (PRCV) was inserted into the genome of human adenovirus type 5 downstream of the early transcription region 3 promoter. The recombinant virus replicated in cultures of the swine testicle ST cell line and directed the synthesis of S antigen to an amount of approximately 33 micrograms per 10(6) cells, as determined by ELISA. The antigen was cell-associated except in the late phase of the infection, when a low amount (4 micrograms per 10(6) cells) was released in the culture supernatant. The cell-associated antigen consisted of 2 polypeptides of 160 K and 175 K, respectively. The 160 K polypeptide comigrated with the authentic S' precursor from PRCV-infected cells. The 175 K polypeptide had the same mobility as the authentic mature S protein from PRCV-infected cells and from PRCV released in the supernatant. The extracellular recombinant antigen corresponded with the 175 K mature protein. Immunofluorescent staining gave evidence that some recombinant S protein was exposed on the cell surface; it also showed that the protein was recognized by conformation-specific anti-S monoclonal antibodies. Piglets, immunized oronasally with the recombinant adenovirus vector developed PRCV-neutralizing serum antibodies and were partially protected against PRCV-challenge.
authors.affiliation: Laboratory of Virology, Faculty of Veterinary Medicine, University of Gent, Belgium.
authors.forename: P
authors.initials: P
authors.lastname: Callebaut
authors.forename: M
authors.initials: M
authors.lastname: Pensaert
mesh_terms: D000260:Adenoviruses, Human / Q000235:genetics / Q000502:physiology; D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D017934:Coronavirus / Q000276:immunology* / Q000378:metabolism / Q000502:physiology; D018352:Coronavirus Infections / Q000276:immunology / Q000662:veterinary* / Q000821:virology; D017931:DNA Primers; D015870:Gene Expression; D005822:Genetic Vectors; D006801:Humans; D008297:Male; D008562:Membrane Glycoproteins / Q000096:biosynthesis* / Q000276:immunology*; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D011995:Recombination, Genetic; D012141:Respiratory Tract Infections / Q000276:immunology / Q000662:veterinary* / Q000821:virology; D064370:Spike Glycoprotein, Coronavirus; D013552:Swine; D013553:Swine Diseases; D013737:Testis / Q000821:virology; D013997:Time Factors; D014158:Transcription, Genetic; D014614:Vaccines, Synthetic; D014759:Viral Envelope Proteins / Q000096:biosynthesis* / Q000276:immunology*; D014765:Viral Vaccines; D014779:Virus Replication
chemical_list: D017931:DNA Primers; D008562:Membrane Glycoproteins; D064370:Spike Glycoprotein, Coronavirus; D014614:Vaccines, Synthetic; D014759:Viral Envelope Proteins; D014765:Viral Vaccines; C578557:spike glycoprotein, SARS-CoV; C578553:spike protein, mouse hepatitis virus