Herpes simplex virus 1 (HSV-1) is a pathogen which primarily infects the oral mucosa. Glycoprotein B (gB) plays an essential role in HSV infection and is highly conserved among different herpesviruses. It is component of the virion envelope, elicits high titers of neutralizing antibody, functions in penetration of virions into host cells (ts mutation), and evidence suggests that gB forms complexes with submembrane proteins (syn mutation). Analyses of the nucleotide sequence and hydropathicity of gB predict that it contains an extracellular hydrophilic domain, a hydrophobic transmembrane region, and a charged carboxyl terminus projecting into the cytoplasm. Studies are designed to elucidate the requirements for the various functions of gB and to verify aspects of the structure of the molecule focusing on the extracellular region and the carboxyl terminus. 1) We are continuing to map antigenic and functional domains on the hydrophilic region of gB by marker rescue of mutants resistant to potent neutralizing monoclonal antibodies. Cognitive sites of antibodies will be mapped by reconstructing the gB with small, in-frame deletions using a panel of DNaseI deletions clones. 2) We will map the boundies of the domain of the syn locus at the charged carboxyl terminus of mutants in gB. By inducing compensatory mutations across the genome, genes encoding proteins which form complexes with gB will be identified by marker rescue with wild type DNA fragments. 3) Contributions of specific N- linked glycosylation sites to the structure, function, and transport of gB will be studied using site-directed mutagenesis to alter the consenus sequence for carbohydrate addition. gB constructs will be expressed in cells containing a resident HSV-1 alpha 4 gene by transfecting with DNA of plasmids carrying the gB gene linked to the dihydrofolate reductase gene resistant to methotrexate. Analysis of gB functions in virions will be carried out by superinfecting cells producing gB constitutively with conditional lethal mutants in gB or by recombining gB constructs into the viral genome. Ultimately, these data will contribute to our understanding of the functional topology of the gB molecule.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI030873-07
Application #
3145854
Study Section
Experimental Virology Study Section (EVR)
Project Start
1987-05-01
Project End
1995-07-31
Budget Start
1993-08-01
Budget End
1994-07-31
Support Year
7
Fiscal Year
1993
Total Cost
Indirect Cost
Name
University of California San Francisco
Department
Type
Schools of Dentistry
DUNS #
073133571
City
San Francisco
State
CA
Country
United States
Zip Code
94143
Navarro, D; Qadri, I; Pereira, L (1993) Transport and secretion of truncated derivatives of herpes simplex virus 1 glycoprotein B. Virology 192:234-45
Sanchez-Pescador, L; Pereira, L; Charlebois, E D et al. (1993) Antibodies to epitopes of herpes simplex virus type 1 glycoprotein B (gB) in human sera: analysis of functional gB epitopes defined by inhibition of murine monoclonal antibodies. J Infect Dis 168:844-53
Mirda, D P; Navarro, D; Paz, P et al. (1992) The fibroblast growth factor receptor is not required for herpes simplex virus type 1 infection. J Virol 66:448-57
Carter, S R; Pereira, L; Paz, P et al. (1992) A quantitative assay of retrograde transported HSV in the trigeminal ganglion. Invest Ophthalmol Vis Sci 33:1934-9
Navarro, D; Paz, P; Pereira, L (1992) Domains of herpes simplex virus I glycoprotein B that function in virus penetration, cell-to-cell spread, and cell fusion. Virology 186:99-112
Torrisi, M R; Di Lazzaro, C; Pavan, A et al. (1992) Herpes simplex virus envelopment and maturation studied by fracture label. J Virol 66:554-61
Basgoz, N; Qadri, I; Navarro, D et al. (1992) The amino terminus of human cytomegalovirus glycoprotein B contains epitopes that vary among strains. J Gen Virol 73 ( Pt 4):983-8
Sanchez-Pescador, L; Paz, P; Navarro, D et al. (1992) Epitopes of herpes simplex virus type 1 glycoprotein B that bind type-common neutralizing antibodies elicit type-specific antibody-dependent cellular cytotoxicity. J Infect Dis 166:623-7
Qadri, I; Navarro, D; Paz, P et al. (1992) Assembly of conformation-dependent neutralizing domains on glycoprotein B of human cytomegalovirus. J Gen Virol 73 ( Pt 11):2913-21
Wooden, S K; Li, L J; Navarro, D et al. (1991) Transactivation of the grp78 promoter by malfolded proteins, glycosylation block, and calcium ionophore is mediated through a proximal region containing a CCAAT motif which interacts with CTF/NF-I. Mol Cell Biol 11:5612-23

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