Delineation of the essential function of bovine herpesvirus 1 gD: an indication for the modulatory role of gD in virus entry.
The entry process of alphaherpesviruses consists of two steps, initial virus attachment and subsequent virus penetration involving membrane fusion. Glycoprotein D (gD) of the alphaherpesvirus bovine herpesvirus 1 (BHV 1) is an essential envelope protein, and it has been previously documented that gD plays a significant part in both of the virus entry steps. In order to gain further insight into the virus entry process, we attempted to define the essential function of BHV 1 gD. We replaced the gD transmembrane and cytoplasmic domains with a lipid-addition signal sequence from human decay accelerating factor and produced a stably transfected Madin Darby bovine kidney (MDBK) cell line that expresses a nonfusogenic, glycosylphosphatidylinositol (GPI)-anchored gD. We found that this cell line was able to support the growth of a gD gene-deletion mutant; the resultant gD mutant progeny contained the GPI-anchored gD on its virions and was able to enter into and produce a production infection in MDBK cells. This result suggests that fusion activity does not constitute the essential function of gD. In addition, we found that a gD-null virus (a virus containing no gD on its virion) could infect gD-expressing cells, but not normal MDBK cells. The ability of the gD-null virus to infect gD-expressing cells was dependent on the gD present on the cell surface, since either treating cells with phosphatidylinositol-specific phospholipase C to remove the GPI-anchored gD or incubating cells with gD monoclonal antibodies could block gD-null virus infection. This demonstrates that gD present on the cell surface can act in trans to facilitate the entry of virion lacking gD. This indicates that essential gD function can take place in the absence of gD-mediated virus attachment and membrane fusion. We also found that the gD monoclonal antibodies that block gD-null virus entry into gD-expressing cells are strictly restricted to the monoclonal antibodies that show postadsorption neutralization activity, indicating that the trans-acting function exhibited by the gD present on the cell surface represents the same function as defined by postadsorption antibody neurtralization. The results from this study suggest that the essential function of gD in virus entry is to modulate other virus-cell interaction(s) involved in productive virus penetration.
Liang
X
X
Veterinary Infectious Disease Organization, University of Saskatchewan, Saskatoon, Canada.
Pyne
C
C
Li
Y
Y
Babiuk
L A
LA
Kowalski
J
J
D000595:Amino Acid Sequence; D000818:Animals; D001287:Attachment Sites, Microbiological; D001483:Base Sequence; D002417:Cattle; D002478:Cells, Cultured; D004279:DNA, Viral / Q000235:genetics; D017353:Gene Deletion; D015870:Gene Expression; D005814:Genes, Viral; D006017:Glycolipids / Q000737:chemistry; D007242:Herpesvirus 1, Bovine / Q000235:genetics / Q000472:pathogenicity* / Q000502:physiology*; D008561:Membrane Fusion; D008969:Molecular Sequence Data; D014162:Transfection; D014759:Viral Envelope Proteins / Q000737:chemistry / Q000235:genetics / Q000502:physiology*
D004279:DNA, Viral; D006017:Glycolipids; D014759:Viral Envelope Proteins