title: Scanning Mutagenesis of Human Cytomegalovirus Glycoprotein gH/gL.
abstract: The core, conserved function of the herpesvirus gH/gL is to promote gB-mediated membrane fusion during entry, although the mechanism is poorly understood. The human cytomegalovirus (HCMV) gH/gL can exist as either the gH/gL/gO trimer or the gH/gL/UL128/UL130/UL131 (gH/gL/UL128-131) pentamer. One model suggests that gH/gL/gO provides the core fusion role during entry into all cells within the broad tropism of HCMV, whereas gH/gL/UL128-131 acts at an earlier stage, by a distinct receptor-binding mechanism to enhance infection of select cell types, such as epithelial cells, endothelial cells, and monocytes/macrophages. To further study the distinct functions of these complexes, mutants with individual charged cluster-to-alanine (CCTA) mutations of gH and gL were combined to generate a library of 80 mutant gH/gL heterodimers. The majority of the mutant gH/gL complexes were unable to facilitate gB-mediated membrane fusion in transient-expression cell-cell fusion experiments. In contrast, these mutants supported the formation of gH/gL/UL128-131 complexes that could block HCMV infection in receptor interference experiments. These results suggest that receptor interactions with gH/gL/UL128-131 involve surfaces contained on the UL128-131 proteins but not on gH/gL. gH/gL/UL128-131 receptor interference could be blocked with anti-gH antibodies, suggesting that interference is a cell surface phenomenon and that anti-gH antibodies can block gH/gL/UL128-131 in a manner that is distinct from that for gH/gL/gO.
  
  
  OBJECTIVE
  Interest in the gH/gL complexes of HCMV (especially gH/gL/UL128-131) as vaccine targets has far outpaced our understanding of the mechanism by which they facilitate entry and contribute to broad cellular tropism. For Epstein-Barr virus (EBV), gH/gL and gH/gL/gp42 are both capable of promoting gB fusion for entry into epithelial cells and B cells, respectively. In contrast, HCMV gH/gL/gO appears to be the sole fusion cofactor that promotes gB fusion activity, whereas gH/gL/UL128-131 expands cell tropism through a distinct yet unknown mechanism. This study suggests that the surfaces of HCMV gH/gL are critical for promoting gB fusion but are dispensable for gH/gL/UL128-131 receptor interaction. This underscores the importance of gH/gL/gO in HCMV entry into all cell types and reaffirms the complex as a candidate target for vaccine development. The two functionally distinct forms of gH/gL present in HCMV make for a useful model with which to study the fundamental mechanisms by which herpesvirus gH/gL regulates gB fusion.
authors.affiliation: Division of Biological Sciences, University of Montana, Missoula, Montana, USA Cellular, Molecular and Microbial Biology Program, University of Montana, Missoula, Montana, USA Center for Biomolecular Structure and Dynamics, University of Montana, Missoula, Montana, USA.
authors.forename: Eric P
authors.initials: EP
authors.lastname: Schultz
authors.affiliation: Division of Biological Sciences, University of Montana, Missoula, Montana, USA Cellular, Molecular and Microbial Biology Program, University of Montana, Missoula, Montana, USA Center for Biomolecular Structure and Dynamics, University of Montana, Missoula, Montana, USA.
authors.forename: Jean-Marc
authors.initials: JM
authors.lastname: Lanchy
authors.affiliation: Division of Biological Sciences, University of Montana, Missoula, Montana, USA Cellular, Molecular and Microbial Biology Program, University of Montana, Missoula, Montana, USA Center for Biomolecular Structure and Dynamics, University of Montana, Missoula, Montana, USA.
authors.forename: Erin E
authors.initials: EE
authors.lastname: Ellerbeck
authors.affiliation: Division of Biological Sciences, University of Montana, Missoula, Montana, USA Cellular, Molecular and Microbial Biology Program, University of Montana, Missoula, Montana, USA Biochemistry and Biophysics Program, University of Montana, Missoula, Montana, USA Center for Biomolecular Structure and Dynamics, University of Montana, Missoula, Montana, USA brent.ryckman@mso.umt.edu.
authors.forename: Brent J
authors.initials: BJ
authors.lastname: Ryckman
mesh_terms: D019943:Amino Acid Substitution; D002460:Cell Line; D003587:Cytomegalovirus / Q000235:genetics / Q000502:physiology*; D004252:DNA Mutational Analysis; D006801:Humans; D016297:Mutagenesis, Site-Directed; D055503:Protein Multimerization; D014759:Viral Envelope Proteins / Q000235:genetics* / Q000378:metabolism; D053586:Virus Internalization
chemical_list: C077406:UL115 protein, Human herpesvirus 5; D014759:Viral Envelope Proteins; C499358:glycoprotein H, Human cytomegalovirus