Characterization of Varicella-Zoster virus glycoprotein K (open reading frame 5) and its role in virus growth.
Varicella-zoster virus (VZV) is an alphaherpesvirus that is the causative agent of chickenpox and herpes zoster. VZV open reading frame 5 (ORF5) encodes glycoprotein K (gK), which is conserved among alphaherpesviruses. While VZV gK has not been characterized, and its role in viral replication is unknown, homologs of VZV gK in herpes simplex virus type 1 (HSV-1) and pseudorabies virus (PRV) have been well studied. To identify the VZV ORF5 gene product, we raised a polyclonal antibody against a fusion protein of ORF5 codons 25 to 122 with glutathione S-transferase and used it to study the protein in infected cells. A 40,000-molecular-weight protein was detected in cell-free virus by Western blotting. In immunogold electron microscopic studies, VZV gK was in enveloped virions and was evenly distributed in the cytoplasm in infected cells. To determine the function of VZV gK in virus growth, a series of gK deletion mutants were constructed with VZV cosmid DNA derived from the Oka strain. Full and partial deletions in gK prevented viral replication when the gK mutant cosmids were transfected into melanoma cells. Insertion of the HSV-1 (KOS) gK gene into the endogenous VZV gK site did not compensate for the deletion of VZV gK. The replacement of VZV gK at a nonnative AvrII site in the VZV genome restored the phenotypic characteristics of intact recombinant Oka (rOka) virus. Moreover, gK complementing cells transfected with a full gK deletion mutant exhibited viral plaques indistinguishable from those of rOka. Our results are consistent with the studies of gK proteins of HSV-1 and PRV showing that gK is indispensable for viral replication.
Mo
C
C
Department of Pediatrics, Stanford University School of Medicine, Stanford, California 94305, USA. cmo@cmgm.stanford.edu
Suen
J
J
Sommer
M
M
Arvin
A
A
D000595:Amino Acid Sequence; D002462:Cell Membrane / Q000378:metabolism; D003360:Cosmids; D003593:Cytoplasm / Q000378:metabolism; D015252:Deoxyribonucleases, Type II Site-Specific; D006023:Glycoproteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D014645:Herpesvirus 3, Human / Q000235:genetics / Q000254:growth & development* / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D014407:Tumor Cells, Cultured; D014759:Viral Envelope Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D010948:Viral Plaque Assay; D014771:Virion / Q000378:metabolism / Q000648:ultrastructure
D006023:Glycoproteins; C118859:ORF5 protein, Human herpesvirus 3; D014759:Viral Envelope Proteins; C064053:CCTAGG-specific type II deoxyribonucleases; D015252:Deoxyribonucleases, Type II Site-Specific