title: The class II membrane glycoprotein G of bovine respiratory syncytial virus, expressed from a synthetic open reading frame, is incorporated into virions of recombinant bovine herpesvirus 1.
abstract: The bovine herpesvirus 1 (BHV-1) recombinants BHV-1/eG(ori) and BHV-1/eG(syn) were isolated after insertion of expression cassettes which contained either a genomic RNA-derived cDNA fragment (BHV-1/eG(ori)) or a modified, chemically synthesized open reading frame (ORF) (BHV-1/eG(syn)), which both encode the attachment glycoprotein G of bovine respiratory syncytial virus (BRSV), a class II membrane glycoprotein. Northern blot analyses and nuclear runoff transcription experiments indicated that transcripts encompassing the authentic BRSV G ORF were unstable in the nucleus of BHV-1/eG(ori)-infected cells. In contrast, high levels of BRSV G RNA were detected in BHV-1/eG(syn)-infected cells. Immunoblots showed that the BHV-1/eG(syn)-expressed BRSV G glycoprotein contains N- and O-linked carbohydrates and that it is incorporated into the membrane of infected cells and into the envelope of BHV-1/eG(syn) virions. The latter was also demonstrated by neutralization of BHV-1/eG(syn) infectivity by monoclonal antibodies or polyclonal anti-BRSV G antisera and complement. Our results show that expression of the BRSV G glycoprotein by BHV-1 was dependent on the modification of the BRSV G ORF and indicate that incorporation of class II membrane glycoproteins into BHV-1 virions does not necessarily require BHV-1-specific signals. This raises the possibility of targeting heterologous polypeptides to the viral envelope, which might enable the construction of BHV-1 recombinants with new biological properties and the development of improved BHV-1-based live and inactivated vector vaccines.
authors.affiliation: Institute of Molecular and Cellular Virology, Friedrich-Loeffler-Institutes, Federal Research Centre for Virus Diseases of Animals, Riems, Germany.
authors.forename: G
authors.initials: G
authors.lastname: Kühnle
authors.forename: A
authors.initials: A
authors.lastname: Heinze
authors.forename: J
authors.initials: J
authors.lastname: Schmitt
authors.forename: K
authors.initials: K
authors.lastname: Giesow
authors.forename: G
authors.initials: G
authors.lastname: Taylor
authors.forename: I
authors.initials: I
authors.lastname: Morrison
authors.forename: F A
authors.initials: FA
authors.lastname: Rijsewijk
authors.forename: J T
authors.initials: JT
authors.lastname: van Oirschot
authors.forename: G M
authors.initials: GM
authors.lastname: Keil
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002417:Cattle; D002460:Cell Line; D004279:DNA, Viral; D015870:Gene Expression; D005822:Genetic Vectors / Q000235:genetics*; D016679:Genome, Viral; D007242:Herpesvirus 1, Bovine / Q000235:genetics*; D008969:Molecular Sequence Data; D009500:Neutralization Tests; D016366:Open Reading Frames; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D011995:Recombination, Genetic; D018112:Respiratory Syncytial Virus, Bovine / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic; D014759:Viral Envelope Proteins / Q000096:biosynthesis* / Q000235:genetics; D014771:Virion / Q000378:metabolism
chemical_list: D004279:DNA, Viral; C066028:G protein, Bovine respiratory syncytial virus; D011993:Recombinant Fusion Proteins; D014759:Viral Envelope Proteins