Herpes simplex virus (HSV) remains a major pathogen world-wide causing human diseases including cold sores, eye and genital infections, neonatal infections, and encephalitis. HSV establishes a life-long latent infection which reactivates periodically to produce recurrent disease. The virus envelope contains ten glycoproteins that are important for infection and pathogenesis. This grant focuses on the structure-function relationship of two of the glycoproteins, gD and gB, both of which are crucial for viral entry and are in Phase III trials as human vaccines. Few details are known about the entry process or about structural changes that occur to the glycoproteins during entry. Our goal is to better define the structures of gD and gB, and to understand how structure is related to function.
In Aim 1, three approaches will be used to probe gD structure at increasing levels of resolution; (i) antigenic and biophysical studies; (ii) visualization by EM methods including scanning transmission electron microscopy and 2-D electron crystallography; and (iii) crystallization of gD and of gD complexed with MAb DL11 leading to X-ray diffraction analysis. The DL11 epitope overlaps a functional site as well as a virus neutralizing site on gD. All of the studies in Aim 1 will employ biologically active and antigenically correct, truncated forms of gD-1 and gD-2 obtained from baculovirus recombinants.
In Aim 2 the recombinant proteins and non-functional but antigenically active mutants will be used in in vitro assays to dissect the effects of the mutations on entry. We will look at the effect of mutations on the ability of gD to bind to cells, block HSV infection, interact with the mannose-6-phosphate receptor, and function in fusion. The goal is to understand what regions of gD are needed for receptor binding as well as to define the regions of gD involved in any additional role played by the protein in virus infection.
Aim 3 focuses on a structure-function analysis of gB. Dimeric gB found in virus-infected cells and in virions is closely associated with fusion leading to virus entry and cell to cell spread. Our hypothesis it that a """"""""triggering"""""""" event causes conformational changes in dimeric gB resulting in fusion between the virion envelope and the cellular membrane. We will examine: (i) gB purified from HSV-infected cells, or truncated and purified from a baculovirus expression system; (ii) gB as it exits in virions; (iii) gB as it exists in the cell surface; and (iv) the role of gB in fusion using a new assay. These studies will involve the use of a panel of MAbs, some of which recognize only the dimeric form of gB. Thus the specific aims are: (l) to study the immunological and biophysical properties of HSV gD; (2) to relate structural properties of gD to its function in virus infection; and (3) to relate structural properties of HSV gB to its function in virus infection.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI018289-17
Application #
2671730
Study Section
Virology Study Section (VR)
Project Start
1981-09-30
Project End
2000-07-31
Budget Start
1998-08-01
Budget End
1999-07-31
Support Year
17
Fiscal Year
1998
Total Cost
Indirect Cost
Name
University of Pennsylvania
Department
Microbiology/Immun/Virology
Type
Schools of Dentistry
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
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Hook, Lauren M; Cairns, Tina M; Awasthi, Sita et al. (2018) Vaccine-induced antibodies to herpes simplex virus glycoprotein D epitopes involved in virus entry and cell-to-cell spread correlate with protection against genital disease in guinea pigs. PLoS Pathog 14:e1007095
Hensel, Michael T; Marshall, Jason D; Dorwart, Michael R et al. (2017) Prophylactic Herpes Simplex Virus 2 (HSV-2) Vaccines Adjuvanted with Stable Emulsion and Toll-Like Receptor 9 Agonist Induce a Robust HSV-2-Specific Cell-Mediated Immune Response, Protect against Symptomatic Disease, and Reduce the Latent Viral Reservoir. J Virol 91:
Cairns, Tina M; Ditto, Noah T; Lou, Huan et al. (2017) Global sensing of the antigenic structure of herpes simplex virus gD using high-throughput array-based SPR imaging. PLoS Pathog 13:e1006430
Fontana, Juan; Atanasiu, Doina; Saw, Wan Ting et al. (2017) The Fusion Loops of the Initial Prefusion Conformation of Herpes Simplex Virus 1 Fusion Protein Point Toward the Membrane. MBio 8:
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 :
Persson, Josefine; Zhang, Yuan; Olafsdottir, Thorunn A et al. (2016) Nasal Immunization Confers High Avidity Neutralizing Antibody Response and Immunity to Primary and Recurrent Genital Herpes in Guinea Pigs. Front Immunol 7:640
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

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