title: Mutagenesis of varicella-zoster virus glycoprotein I (gI) identifies a cysteine residue critical for gE/gI heterodimer formation, gI structure, and virulence in skin cells.
abstract: Varicella-zoster virus (VZV) is the alphaherpesvirus that causes chicken pox (varicella) and shingles (zoster). The two VZV glycoproteins gE and gI form a heterodimer that mediates efficient cell-to-cell spread. Deletion of gI yields a small-plaque-phenotype virus, ΔgI virus, which is avirulent in human skin using the xenograft model of VZV pathogenesis. In the present study, 10 mutant viruses were generated to determine which residues were required for the typical function of gI. Three phosphorylation sites in the cytoplasmic domain of gI were not required for VZV virulence in vivo. Two deletion mutants mapped a gE binding region in gI to residues 105 to 125. A glycosylation site, N116, in this region did not affect virulence. Substitution of four cysteine residues highly conserved in the Alphaherpesvirinae established that C95 is required for gE/gI heterodimer formation. The C95A and Δ105-125 (with residues 105 to 125 deleted) viruses had small-plaque phenotypes with reduced replication kinetics in vitro similar to those of the ΔgI virus. The Δ105-125 virus was avirulent for human skin in vivo. In contrast, the C95A mutant replicated in vivo but with significantly reduced kinetics compared to those of the wild-type virus. In addition to abolished gE/gI heterodimer formation, gI from the C95A or the Δ105-125 mutant was not recognized by monoclonal antibodies that detect the canonical conformation of gI, demonstrating structural disruption of gI in these viruses. This alteration prevented gI incorporation into virus particles. Thus, residues C95 and 105 to 125 are critical for gI structure required for gE/gI heterodimer formation, virion incorporation, and ultimately, effective viral spread in human skin.
authors.affiliation: Stanford University School of Medicine, Stanford, CA 94305, USA. sloliver@stanford.edu
authors.forename: Stefan L
authors.initials: SL
authors.lastname: Oliver
authors.forename: Marvin H
authors.initials: MH
authors.lastname: Sommer
authors.forename: Mike
authors.initials: M
authors.lastname: Reichelt
authors.forename: Jaya
authors.initials: J
authors.lastname: Rajamani
authors.forename: Leonssia
authors.initials: L
authors.lastname: Vlaycheva-Beisheim
authors.forename: Shaye
authors.initials: S
authors.lastname: Stamatis
authors.forename: Jason
authors.initials: J
authors.lastname: Cheng
authors.forename: Carol
authors.initials: C
authors.lastname: Jones
authors.forename: James
authors.initials: J
authors.lastname: Zehnder
authors.forename: Ann M
authors.initials: AM
authors.lastname: Arvin
mesh_terms: D019943:Amino Acid Substitution; D002460:Cell Line; D003545:Cysteine / Q000235:genetics; D004252:DNA Mutational Analysis; D014645:Herpesvirus 3, Human / Q000235:genetics / Q000472:pathogenicity*; D006801:Humans; D025941:Protein Interaction Mapping; D055503:Protein Multimerization; D017384:Sequence Deletion; D012867:Skin / Q000473:pathology / Q000821:virology*; D014759:Viral Envelope Proteins / Q000235:genetics / Q000378:metabolism*; D010948:Viral Plaque Assay; D014774:Virulence; D037521:Virulence Factors / Q000235:genetics / Q000378:metabolism*; D014779:Virus Replication
chemical_list: D014759:Viral Envelope Proteins; D037521:Virulence Factors; C056913:glycoprotein E, varicella-zoster virus; D003545:Cysteine