Functional and physical interactions of the herpes simplex virus type 1 UL20 membrane protein with glycoprotein K.
Herpes simplex virus type 1 glycoprotein K (gK) and the UL20 protein (UL20p) are coordinately transported to the trans-Golgi network (TGN) and cell surfaces and are required for cytoplasmic virion envelopment at the TGN. In addition, cell surface expression of gK and UL20p is required for virus-induced cell fusion. Previously, confocal microscopy colocalization and intracellular transport experiments strongly suggested direct protein-protein interactions between gK and UL20p. Direct protein-protein interactions between gK and UL20p were demonstrated through reciprocal coimmunoprecipitation experiments, as well as with glutathione S-transferase (GST) pull-down experiments. A fusion protein consisting of the amino-terminal 66 amino acids of UL20p fused in-frame with GST was expressed in Escherichia coli and purified via glutathione column chromatography. Precipitation of GST-UL20p from mixtures of GST-UL20p fusion protein with cellular extracts containing gK specifically coprecipitated gK but not other viral glycoproteins. The purified UL20p-GST fusion protein reacted with all gK-associated protein species. It was concluded that the amino terminus of UL20p, most likely, interacted with gK domain III, which is predicted to lie intracellularly. UL20p-gK domain-specific interactions must serve important functions in the coordinate transport of UL20p and gK to the TGN, because retention of UL20p in the endoplasmic reticulum (ER) via the addition of an ER retention signal at the carboxyl terminus of UL20p forced the ER retention of gK and drastically inhibited intracellular virion envelopment and virus-induced cell fusion.
Foster
Timothy P
TP
Division of Biotechnology and Molecular Medicine, School of Veterinary Medicine, Louisiana State University, Baton Rouge, LA 70803, USA.
Chouljenko
Vladimir N
VN
Kousoulas
K G
KG
D000818:Animals; D015153:Blotting, Western; D002522:Chlorocebus aethiops; D004591:Electrophoresis, Polyacrylamide Gel; D047468:Immunoprecipitation; D008565:Membrane Proteins / Q000378:metabolism*; D046529:Microscopy, Electron, Transmission; D021381:Protein Transport / Q000502:physiology; D014709:Vero Cells; D014759:Viral Envelope Proteins / Q000378:metabolism; D014760:Viral Fusion Proteins / Q000378:metabolism*; D014764:Viral Proteins / Q000378:metabolism*; D014771:Virion / Q000378:metabolism*; D021601:trans-Golgi Network / Q000378:metabolism* / Q000648:ultrastructure
D008565:Membrane Proteins; C121742:UL20 protein, Herpes simplex virus type 1; C072448:UL53 protein, Human herpesvirus 1; D014759:Viral Envelope Proteins; D014760:Viral Fusion Proteins; D014764:Viral Proteins