Herpes simplex viruses (HSVs) cause cold sores, eye and genital infections, neonatal infections and encephalitis. The virus establishes lifelong latent infections in sensory ganglia. The envelope contains eleven virus-encoded glycoproteins of which four are essential for virus entry. These are gD, the receptor binding protein, gB and a complex of gH and gL. Several cell molecules serve as entry receptors and in each case the receptor binds gD. Most strains can use two of these, HveA (HVEM) and nectin- 1 (HveC). Whereas HveA (a TNF receptor) is found most abundantly on T lymphocytes, nectin- 1 (a cell adhesion molecule) is abundant on epithelial and neuronal cells.
The specific aims of this grant are: (1) to characterize the interaction between gD and nectin- 1; (2) to determine the role HSV entry receptors in a mouse model of virus infection; and (3) to carry out structure-function studies of HSV gB.
In Aim 1, our goal is to increase our understanding of the interaction between gD and nectin-1 in vitro, and on cells. When gD binds HveA, it undergoes two conformational changes as revealed by crystallographic data. We hypothesize that at least one of these also occurs when gD binds nectin- 1 and this change may play a role in later steps of entry. Solution of the structure of the gD/nectin-1 complex will test this hypothesis and is a major goal of this project. These studies combined with targeted mutagenesis will enhance our understanding of this protein-protein interaction. Nectin-1 can interact in trans with itself as well as with other nectins at cellular adherens junctions. We speculate that gD acts as a ligand for nectin-1 at these junctions so that receptor is available for virus spread to the next cell. Although nectin-1 appears to be more abundant than HveA on epithelial and neuronal cells, the role of these two receptors in HSV pathogenesis is not known. We developed a panel of gD mutants with altered ability to use one or the other receptor.
In Aim 2, we will test the ability of viruses carrying these mutations to infect and cause zosteriform disease in mice. Although gB, gH and gL are required for fusion of the viral envelope with the plasma membrane of the host cell, their role in this process is not understood.
In Aim 3, we will carry out structure function studies of gB in order to clarify its role in virus entry. We will follow up on the observation that gB partitions into lipid rafts on cells at the time of entry. Ultimately, we want to define the cellular targets of this interaction. Recently, we cloned, expressed and purified large quantities of the gB ectodomain. Crystals of this protein diffract and we therefore propose to solve the structure of gB by X-ray crystallography. The studies in this grant may lead to development of new approaches for antiviral therapeutics based on targets of the HSV entry process. ? ? ?

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI056045-03
Application #
7010099
Study Section
Virology Study Section (VR)
Program Officer
Beisel, Christopher E
Project Start
2004-02-01
Project End
2009-01-31
Budget Start
2006-02-01
Budget End
2007-01-31
Support Year
3
Fiscal Year
2006
Total Cost
$386,938
Indirect Cost
Name
University of Pennsylvania
Department
Pathology
Type
Schools of Veterinary Medicine
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
Atanasiu, Doina; Saw, Wan Ting; Eisenberg, Roselyn J et al. (2016) Regulation of HSV glycoprotein induced cascade of events governing cell-cell fusion. J Virol :
Cairns, Tina M; Huang, Zhen-Yu; Gallagher, John R et al. (2015) Patient-Specific Neutralizing Antibody Responses to Herpes Simplex Virus Are Attributed to Epitopes on gD, gB, or Both and Can Be Type Specific. J Virol 89:9213-31
Saw, Wan Ting; Matsuda, Zene; Eisenberg, Roselyn J et al. (2015) Using a split luciferase assay (SLA) to measure the kinetics of cell-cell fusion mediated by herpes simplex virus glycoproteins. Methods 90:68-75
Cairns, Tina M; Huang, Zhen-Yu; Whitbeck, J Charles et al. (2014) Dissection of the antibody response against herpes simplex virus glycoproteins in naturally infected humans. J Virol 88:12612-22
Gallagher, John R; Atanasiu, Doina; Saw, Wan Ting et al. (2014) Functional fluorescent protein insertions in herpes simplex virus gB report on gB conformation before and after execution of membrane fusion. PLoS Pathog 10:e1004373
Whitbeck, J Charles; Huang, Zhen-Yu; Cairns, Tina M et al. (2014) Repertoire of epitopes recognized by serum IgG from humans vaccinated with herpes simplex virus 2 glycoprotein D. J Virol 88:7786-95
Cairns, Tina M; Fontana, Juan; Huang, Zhen-Yu et al. (2014) Mechanism of neutralization of herpes simplex virus by antibodies directed at the fusion domain of glycoprotein B. J Virol 88:2677-89
Lazear, Eric; Whitbeck, J Charles; Zuo, Yi et al. (2014) Induction of conformational changes at the N-terminus of herpes simplex virus glycoprotein D upon binding to HVEM and nectin-1. Virology 448:185-95
Atanasiu, Doina; Cairns, Tina M; Whitbeck, J Charles et al. (2013) Regulation of herpes simplex virus gB-induced cell-cell fusion by mutant forms of gH/gL in the absence of gD and cellular receptors. MBio 4:
Maurer, Ulrike E; Zeev-Ben-Mordehai, Tzviya; Pandurangan, Arun Prasad et al. (2013) The structure of herpesvirus fusion glycoprotein B-bilayer complex reveals the protein-membrane and lateral protein-protein interaction. Structure 21:1396-405

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