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.
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