Herpes Virus Fusion and Entry: A Story with Many Characters

Herpesviridae comprise a large family of enveloped DNA viruses all of whom employ orthologs of the same three glycoproteins, gB, gH and gL. Additionally, herpesviruses often employ accessory proteins to bind receptors and/or bind the heterodimer gH/gL or even to determine cell tropism. Sorting out how these proteins function has been resolved to a large extent by structural biology coupled with supporting biochemical and biologic evidence. Together with the G protein of vesicular stomatitis virus, gB is a charter member of the Class III fusion proteins. Unlike VSV G, gB only functions when partnered with gH/gL. However, gH/gL does not resemble any known viral fusion protein and there is evidence that its function is to upregulate the fusogenic activity of gB. In the case of herpes simplex virus, gH/gL itself is upregulated into an active state by the conformational change that occurs when gD, the receptor binding protein, binds one of its receptors. In this review we focus primarily on prototypes of the three subfamilies of herpesviruses. We will present our model for how herpes simplex virus (HSV) regulates fusion in series of highly regulated steps. Our model highlights what is known and also provides a framework to address mechanistic questions about fusion by HSV and herpesviruses in general

Random mutagenesis of the gene encoding a viral ligand for multiple cell entry receptors to obtain viral mutants altered for receptor usage

Herpes simplex virus type 1 (HSV-1) can enter cells expressing any one of multiple entry receptors, including the herpesvirus entry mediator (HVEM), nectin-1, and sites in heparan sulfate generated by specific 3-O-sulfotransferases. The viral ligand for these receptors is glycoprotein D (gD). To define structural requirements for functional interactions of gD with its receptors and to obtain viral mutants altered for receptor usage, we generated a library of HSV-1 mutants with random mutations in the gD gene. Viral isolates selected on a monkey cell line (Vero) were screened for the loss of ability to infect cells expressing each of the HSV-1 receptors. The 10 HSV-1 mutants obtained had 12 mutations in gD, affecting 11 amino acids. All mutations reduced or abrogated viral entry through HVEM and 3-O-sulfated heparan sulfate, indicating that similar features of gD are critical for functional interactions with both these receptors. None of the mutations reduced viral entry through nectin-1, whereas a subset of the mutations conferred ability to use nectin-2 as an entry receptor. These and other results show that features of gD, including conformation of the N terminus, critical for functional interactions with HVEM/3-O-sulfated heparan sulfate, differ from those critical for interactions with nectin-1

The multipartite system that mediates entry of herpes simplex virus into the cell

The multipartite entry-fusion system of herpes simplex virus is made of a quartet of glycoproteins-gD, gB, gH.gL-and three alternative gD receptors, herpesvirus entry mediator (HVEM), nectin1 and modified sites on heparan sulphate. This multipartite system recapitulates the basic steps of virus-cell fusion, i.e. receptor recognition, triggering of fusion and fusion execution. Specifically, in addition to serving as the receptor-binding glycoprotein, gD triggers fusion through a specialised domain, named pro-fusion domain (PFD), located C-terminally in the ectodomain. In the unliganded gD the C-terminal region folds around the N-terminal region, such that gD adopts a closed autoinhibited conformation. In HVEM- and nectin1-bound gD the C-terminal region is displaced (opened conformation). gD is the tool for modification of HSV tropism, through insertion of ligands to heterologous tumour-specific receptors. It is discussed whether gD responds to the interaction with the natural and the heterologous receptors by adopting similar conformations, and whether the closed-to-open switch in conformation is a generalised mechanism of activation. A peculiar recombinant highlighted that the central Ig-folded core of gD may not encode executable functions for entry and that the 219-314 aa segment may be sufficient to trigger fusion. With respect to fusion execution, gB appears to be a prospective fusogen based on its coiled-coil trimeric structure, similar to that of another fusion glycoprotein. On the other hand, gH exhibits molecular elements typical of class 1 fusion glycoproteins, in particular heptad repeats and strong tendency to interact with lipids. Whether fusion execution is carried out by gB or gH.gL, or both glycoproteins in complex or sequentially remains to be determined