Bovine herpesvirus 1 U(s) open reading frame 4 encodes a glycoproteoglycan.
Sequence analysis of the short unique (Us) segment of the bovine herpesvirus 1 (BHV-1) genome predicted that the Us open reading frame (ORF) 4 encodes a protein with homology to glycoprotein G (gG) of other alpha-herpesviruses (P. Leung-Tack, J.-C. Audonnet, and M. Riviere, Virology 199:409-421, 1994). RNA analysis showed that the Us ORF4 is contained within two transcripts of 3.5 and 1.8 kb. The 3.5 kb RNA represents a structurally bicistronic RNA which encompasses the Us ORF3 and Us ORF4, whereas the 1.8-kb RNA constitutes the monocistronic Us ORF4 mRNA. To identify the predicted BHV-I gG, recombinant vaccinia virus expressing the Us ORF4 was used to raise specific antibodies in rabbits. The antiserum recognized a 65-kDa polypeptide and a very diffusely migrating species of proteins with an apparent molecular mass of between 90 and greater than 240 kDa in supernatants of BHV-1-infected cells which was also precipitated together with 61- and 70-kDa polypeptides from cell-associated proteins. The specificity of the reaction was demonstrated by the absence of these proteins from the supernatant of cells infected with the Us ORF4 deletion mutant BHV-l/gp1-8. Treatment of the immunoprecipitated proteins with glycosidases and chondroitinase AC showed that the 65-kDa protein constitutes gG, which contains both N- and O-linked carbohydrates, and that the high-molecular-mass proteins contain glycosaminoglycans linked to a 65-kDa glycoprotein that is antigenically related to gG. These molecules were therefore named glycoproteoglycan C (gpgG). Pulse chase experiments indicated that gG and gpgG were processed from a common precursor molecule with an apparent molecular mass of 61 kDa via a 70-kDa intermediate. Both gG and gpgG could not be found associated with purified virions. In summary, our results identify the BHV-I gG protein and demonstrate the presence of a form of posttranslational modification, glycosamino-glycosylation, that has not yet been described for a herpesvirus-encoded protein.
Keil
G M
GM
Institute for Molecular and Cellular Virology, Insel Riems, Germany.
Engelhardt
T
T
Karger
A
A
Enz
M
M
D000818:Animals; D000906:Antibodies; D015153:Blotting, Western; D002241:Carbohydrates / Q000032:analysis; D002417:Cattle; D002460:Cell Line; D005814:Genes, Viral; D006031:Glycosylation; D007242:Herpesvirus 1, Bovine / Q000235:genetics* / Q000378:metabolism*; D007668:Kidney; D016366:Open Reading Frames; D011509:Proteoglycans / Q000096:biosynthesis*; D012333:RNA, Messenger / Q000378:metabolism; D012367:RNA, Viral / Q000378:metabolism; D011817:Rabbits; D014158:Transcription, Genetic; D014764:Viral Proteins / Q000096:biosynthesis*; D015678:Viral Structural Proteins / Q000235:genetics
D000906:Antibodies; D002241:Carbohydrates; D011509:Proteoglycans; D012333:RNA, Messenger; D012367:RNA, Viral; D014764:Viral Proteins; D015678:Viral Structural Proteins; C046998:bovine herpesvirus type-1 glycoproteins