We observed that HSV 1 induces a receptor for the third component of complement (C3b) following infection of endothelial cells, and a variety of other cell types. The receptor is a viral glycoprotein, gC. Infection with other herpes viruses, including HSV 2, CMV and VZV do not induce a C3b receptor (C3bR). The ability to bind C3b assigns a new function to an abundant HSV 1 glycoprotein. Our objectives are to examine the importance of the C3bR in the pathogenesis of HSV 1 disease and to determine the structure of gC that promotes binding to C3b. We will use monoclonal and polyclonal antibodies that react with gC and prepare complement-coated erthyrocytes which bind to the C3bR to characterize some of the properties of the receptor on infected cells. These include: the kinetics of appearance of the receptor; antigenic similarities between the C3bR on the virus and circulating blood cell; whether additional types of complement receptors (CR2, CR3) are produced; whether the receptor can be detected on human tissues during HSV 1 infection in vivo; whether ligand binding to the receptor effects viral synthesis; whether the receptor protects the virus from complement-mediated lysis; and whether ligand binding to the receptor triggers receptor mobility and capping as it does on various types of blood cells. To date we have only demonstrated C3bR on the surface of infected cells. We will purify the glycoprotein by affinity chromatography and develop an immunoblot assay to determine if the purified glycoprotein binds C3b. If we detect receptor activity on purified gC, it will be the first demonstration of a C3 receptor on an infectious agent. We will prepare tryptic peptide or formic acid digests of purified gC and determine which fragments retain C3b-binding activity. We will examine mutants of HSV 1 gC which lack C3bR and identify the missing fragments by N-terminal amino acid sequencing, tryptic-peptide digests and V8 proteolysis. By expanding our knowledge on the structure and functions of gC, we hope to unravel some of the mysteries as to how this virus causes disease, wit a particular emphasis on injury to the vascular endothelium.

National Institute of Health (NIH)
National Heart, Lung, and Blood Institute (NHLBI)
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Virology Study Section (VR)
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University of Pennsylvania
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Awasthi, Sita; Balliet, John W; Flynn, Jessica A et al. (2014) Protection provided by a herpes simplex virus 2 (HSV-2) glycoprotein C and D subunit antigen vaccine against genital HSV-2 infection in HSV-1-seropositive guinea pigs. J Virol 88:2000-10
Awasthi, Sita; Zumbrun, Elizabeth E; Si, Huaxin et al. (2012) Live attenuated herpes simplex virus 2 glycoprotein E deletion mutant as a vaccine candidate defective in neuronal spread. J Virol 86:4586-98
Lubinski, John M; Lazear, Helen M; Awasthi, Sita et al. (2011) The herpes simplex virus 1 IgG fc receptor blocks antibody-mediated complement activation and antibody-dependent cellular cytotoxicity in vivo. J Virol 85:3239-49
Awasthi, Sita; Lubinski, John M; Shaw, Carolyn E et al. (2011) Immunization with a vaccine combining herpes simplex virus 2 (HSV-2) glycoprotein C (gC) and gD subunits improves the protection of dorsal root ganglia in mice and reduces the frequency of recurrent vaginal shedding of HSV-2 DNA in guinea pigs compared to J Virol 85:10472-86
Huang, Jialing; Lazear, Helen M; Friedman, Harvey M (2011) Completely assembled virus particles detected by transmission electron microscopy in proximal and mid-axons of neurons infected with herpes simplex virus type 1, herpes simplex virus type 2 and pseudorabies virus. Virology 409:12-6
King, Ryan D; Lubinski, John M; Friedman, Harvey M (2009) Herpes simplex virus type 1 infection increases the carbohydrate binding activity and the secretion of cellular galectin-3. Arch Virol 154:609-18
Awasthi, Sita; Lubinski, John M; Friedman, Harvey M (2009) Immunization with HSV-1 glycoprotein C prevents immune evasion from complement and enhances the efficacy of an HSV-1 glycoprotein D subunit vaccine. Vaccine 27:6845-53
Awasthi, Sita; Lubinski, John M; Eisenberg, Roselyn J et al. (2008) An HSV-1 gD mutant virus as an entry-impaired live virus vaccine. Vaccine 26:1195-203
Hook, Lauren M; Huang, Jialing; Jiang, Ming et al. (2008) Blocking antibody access to neutralizing domains on glycoproteins involved in entry as a novel mechanism of immune evasion by herpes simplex virus type 1 glycoproteins C and E. J Virol 82:6935-41
Sutherland, M R; Friedman, H M; Pryzdial, E L G (2007) Thrombin enhances herpes simplex virus infection of cells involving protease-activated receptor 1. J Thromb Haemost 5:1055-61

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