Plasma membrane topology of syncytial domains of herpes simplex virus type 1 glycoprotein K (gK): the UL20 protein enables cell surface localization of gK but not gK-mediated cell-to-cell fusion.
Most spontaneously occurring mutations that cause extensive herpes simplex virus type 1 (HSV-1)-induced cell fusion are single amino acid changes within glycoprotein K (gK). Despite the strong genetic association of gK with virus-induced cell fusion, its direct involvement in cellular membrane fusion has been controversial, largely due to previously unsuccessful efforts to detect gK expression on virion and cellular surfaces. Recently, we showed that gK is expressed on HSV-1 virions and functioned in virus entry (T. P. Foster, G. V. Rybachuk, and K. G. Kousoulas, J. Virol. 75:12431-12438, 2001). To determine whether gK is expressed on cellular surfaces, as well as its membrane topology, we generated the recombinant viruses gKV5DI, gKV5DII, gKV5DIII, and gKV5DIVcontaining insertions of the V5 antigenic epitope within each of four domains of gK predicted to localize either in the cytoplasmic side or in the extracytoplasmic side of cellular membranes. Immunohistochemical and confocal microscopy analyses of infected cells showed that both wild-type and syncytial forms of gK were expressed on cell surfaces. Analysis of the topology of the V5-tagged gK revealed that gK domains I and IV were located extracellularly, whereas domains II and III were localized intracellularly. Transiently expressed gK failed to localize in cellular plasma membranes. In contrast, infection of gK-transfected cells with the gK-null virus DeltagK enabled expression of gK on cell surfaces, as well as gK-mediated membrane fusion. Transient-coexpression experiments revealed that the UL20 protein enabled cell surface expression of gK, but not gK-mediated cell-to-cell fusion, indicating that additional viral proteins are required for expression of the gK syncytial phenotype.
Foster
Timothy P
TP
Division of Biotechnology and Molecular Medicine, School of Veterinary Medicine, Louisiana State University, Baton Rouge 70803, USA.
Alvarez
Xavier
X
Kousoulas
Konstantin G
KG
D000595:Amino Acid Sequence; D000818:Animals; D002462:Cell Membrane / Q000737:chemistry*; D002522:Chlorocebus aethiops; D006031:Glycosylation; D006056:Golgi Apparatus / Q000737:chemistry / Q000502:physiology; D018259:Herpesvirus 1, Human / Q000502:physiology; D008561:Membrane Fusion; D008969:Molecular Sequence Data; D014709:Vero Cells; D014759:Viral Envelope Proteins / Q000032:analysis; D014764:Viral Proteins / Q000032:analysis / Q000737:chemistry / Q000502:physiology*
C121742:UL20 protein, Herpes simplex virus type 1; C072448:UL53 protein, Human herpesvirus 1; D014759:Viral Envelope Proteins; D014764:Viral Proteins; C040495:glycoprotein B, Simplexvirus