title: Substitution of herpes simplex virus 1 entry glycoproteins with those of saimiriine herpesvirus 1 reveals a gD-gH/gL functional interaction and a region within the gD profusion domain that is critical for fusion.
abstract: To gain insight into the mechanism of herpesvirus entry into cells, the four glycoproteins that are necessary for herpes simplex virus (HSV) fusion were cloned from the saimiriine herpesvirus 1 (SaHV-1) genome, a primate member of the alphaherpesvirus family. Cell-cell fusion assays indicate that SaHV-1 entry glycoproteins function with the previously identified alphaherpesvirus entry receptors nectin-1 and CD155 but not with herpesvirus entry mediator (HVEM) or paired immunoglobulin-like type 2 receptor alpha (PILRα). Replacement of HSV-1 gD with the SaHV-1 gD homolog resulted in a complete loss of fusion function when coexpressed with HSV-1 gB and gH/gL. HSV-1 gD was also unable to substitute for SaHV-1 gD when coexpressed with SaHV-1 gB and gH/gL. Similarly, the gH/gL heterodimers from HSV-1 and SaHV-1 were not interchangeable. In contrast, both the HSV-1 and SaHV-1 gB homologs retained function in a heterotypic context. These results suggest that an essential interaction between homotypic gD and gH/gL occurs during both HSV-1 and SaHV-1 entry. To map the site of this homotypic interaction, we created a series of gD chimeras, focusing on the "profusion domain" (PFD) that consists of HSV-1 gD residues 261 to 305 or SaHV-1 gD residues 264 to 307. We identified a seven-amino-acid stretch (264 RTLPPPK 270) at the N terminus of the SaHV-1 gD PFD that contributes to homotypic fusion. Finally, we found that the gD receptor-binding region and PFD cannot function independently but that both can inhibit the function of wild-type gD.
  
  
  OBJECTIVE
  The herpesvirus entry machinery requires the concerted action of at least four glycoproteins; however, details of the interactions among these glycoproteins are not well understood. Like HSV-1, SaHV-1 belongs to the alphaherpesvirus subfamily. Using cell-cell fusion experiments, we found that SaHV-1 uses the entry receptors nectin-1 and CD155 but not HVEM or PILRα. By swapping the entry glycoproteins between HSV-1 and SaHV-1, we revealed a functional interaction between gD and gH/gL. To examine the homotypic interaction site on gD, we evaluated the function of a panel of HSV-1/SaHV-1 gD chimeras and identified a small region in the SaHV-1 gD profusion domain that is critical for SaHV-1 fusion. This study contributes to our understanding of the molecular mechanisms of herpesvirus entry and membrane fusion.
authors.affiliation: Department of Microbiology-Immunology, Feinberg School of Medicine of Northwestern University, Chicago, Illinois, USA.
authors.forename: Qing
authors.initials: Q
authors.lastname: Fan
authors.forename: Richard
authors.initials: R
authors.lastname: Longnecker
authors.forename: Sarah A
authors.initials: SA
authors.lastname: Connolly
mesh_terms: D000818:Animals; D015153:Blotting, Western; D016466:CHO Cells; D015815:Cell Adhesion Molecules / Q000378:metabolism; D002459:Cell Fusion; D003001:Cloning, Molecular; D006224:Cricetinae; D003412:Cricetulus; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D018259:Herpesvirus 1, Human / Q000235:genetics*; D000075983:Nectins; D010957:Plasmids / Q000235:genetics; D011991:Receptors, Virus / Q000378:metabolism; D018139:Simplexvirus / Q000235:genetics*; D014760:Viral Fusion Proteins / Q000235:genetics* / Q000378:metabolism*; D053586:Virus Internalization
chemical_list: D015815:Cell Adhesion Molecules; C000619010:NECTIN1 protein, human; D000075983:Nectins; D011991:Receptors, Virus; D014760:Viral Fusion Proteins; C066100:poliovirus receptor