1. A herpes simplex virus 2 glycoprotein D mutant generated by bacterial artificial chromosome mutagenesis is severely impaired for infecting neuronal cells and infects only Vero cells expressing exogenous HVEM.
- Author
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Wang K, Kappel JD, Canders C, Davila WF, Sayre D, Chavez M, Pesnicak L, and Cohen JI
- Subjects
- Animals, Cell Adhesion Molecules metabolism, Cell Line, Chlorocebus aethiops, Chromosomes, Artificial, Bacterial genetics, Herpesvirus 2, Human metabolism, Humans, Mice, Mice, Inbred BALB C, Mutagenesis, Nectins, Oligonucleotides genetics, Point Mutation genetics, Polymerase Chain Reaction, Receptors, Tumor Necrosis Factor, Member 14 metabolism, Vero Cells metabolism, Herpesvirus 2, Human genetics, Neurons virology, Vero Cells virology, Viral Envelope Proteins genetics, Virus Internalization
- Abstract
We constructed a herpes simplex virus 2 (HSV-2) bacterial artificial chromosome (BAC) clone, bHSV2-BAC38, which contains full-length HSV-2 inserted into a BAC vector. Unlike previously reported HSV-2 BAC clones, the virus genome inserted into this BAC clone has no known gene disruptions. Virus derived from the BAC clone had a wild-type phenotype for growth in vitro and for acute infection, latency, and reactivation in mice. HVEM, expressed on epithelial cells and lymphocytes, and nectin-1, expressed on neurons and epithelial cells, are the two principal receptors used by HSV to enter cells. We used the HSV-2 BAC clone to construct an HSV-2 glycoprotein D mutant (HSV2-gD27) with point mutations in amino acids 215, 222, and 223, which are critical for the interaction of gD with nectin-1. HSV2-gD27 infected cells expressing HVEM, including a human epithelial cell line. However, the virus lost the ability to infect cells expressing only nectin-1, including neuronal cell lines, and did not infect ganglia in mice. Surprisingly, we found that HSV2-gD27 could not infect Vero cells unless we transduced the cells with a retrovirus expressing HVEM. High-level expression of HVEM in Vero cells also resulted in increased syncytia and enhanced cell-to-cell spread in cells infected with wild-type HSV-2. The inability of the HSV2-gD27 mutant to infect neuronal cells in vitro or sensory ganglia in mice after intramuscular inoculation suggests that this HSV-2 mutant might be an attractive candidate for a live attenuated HSV-2 vaccine.
- Published
- 2012
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