Viral and cellular mechanisms of HSV fusion and entry

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Herpesviruses are ubiquitous pathogens that cause significant morbidity and mortality worldwide. Herpes simplex virus (HSV) causes cold sores and sexually transmitted infections. Grave outcomes of HSV infection include neonatal disease, blindness, and disseminated infections of the immunocompromised. Our long-term goal is a comprehensive understanding of the entry mechanisms of this important group of pathogens. Although much progress has been made, the complex mechanisms of herpesvirus fusion and entry remain enigmatic. This has posed a roadblock to developing clinically effective entry inhibitors. HSV enters most cultured mammalian cells and commandeers diverse entry pathways. An emerging concept in herpesvirology is that endosomal pH of the host cell is required for viral entry, often in a cell type specific manner. However, the mechanistic role that low pH plays in herpesviral entry is not clear. The HSV envelope proteins gB, gD, and the gH/gL heterodimer are required but are likely not sufficient for membrane fusion and entry in the context of HSV infection. Envelope protein gC is the principal mediator of viral attachment to cell surface heparan sulfate proteoglycans during entry. However, this interaction is dispensable for viral entry in cell culture and does not explain the infection defect of gC-deleted viruses. We recently revealed a new, post-attachment function for gC in HSV entry. gC influences the fusion protein gB and promotes efficient penetration of HSV from endosomes during entry. gC elevates the pH-threshold of fusion- associated conformational changes in the fusion protein gB. We formulated complementary yet independent aims to delineate previously unrecognized functions of HSV-1 gC. In Specific Aim 1, we will elucidate gC's role in fusion and entry by employing a battery of assays in the context of the required HSV envelope proteins and cellular receptors. Specific Aim 2 encompasses structure-function studies of gC pertinent to entry and infection of epithelial cells, the main target of primary and recurrent HSV infection. Our experimental design employs techniques of cell biology, biochemistry, and molecular virology. Completion of the aims will reveal detailed insight into the multifunctional nature of HSV-1 gC and will fill critical knowledge gaps about how the complex fusion mechanism of herpesviruses is triggered in pathophysiologically relevant cell types. These studies will aid in the development of new preventions and therapeutics for HSV.