The intracellular assembly, maturation, and transport of varicella-zoster virus (VZV) will be studied. VZV, the highly contagious etiologic agent of chickenpox (varicella) and shingles (zoster) is spread by infectious virions that are present in fluid that fills vesicular skin lesions. Virions secreted by cells in vitro, however, are not infectious and yield infectious particles only when the cells are disrupted. EM observations suggest that nucleocapsids of VZV are enveloped while exiting from the nucleus through the inner nuclear membrane. Virions then travel within the cisternal compartment of the Golgi apparatus where they are diverted from the secretory pathway to prelysosomal structures. Diverted virions accumulate in acid phosphatase-containing vacuoles in which the particles appear to be degraded. These observations are consistent with the hypothesis that the intracellular route of transport of newly synthesized VZV particles is similar to that of newly synthesized acid hydrolases. We have found that VZV envelope glycoproteins (gps), like the acid hydrolases, contain mannose 6-phosphate (Man 6-P) residues. We postulate that Man 6-P on the viral envelope leads VZV to bind to Man 6-P receptors (MPRs) in the Golgi, which, in turn, direct newly synthesized VZV to endosomes. MPRs also cycle to the cell surface; therefore, if VZV were to escape degradation in the epidermis of infected patients, then Man 6-P on the viral envelope might bind to MPRs on the plasma membrane of target cells, facilitation infection. We will test the hypotheses that VZV binds to MPRs, that intracellular binding of VZV to MPRs in cultured cells diverts virions from the secretory to the lysosomal pathway, that binding of intact VZV to surface MPRs facilities viral infection of target cells, and that cells in suprabasal layers of the epidermis of infected patients lack intracellular MPRs and thus secrete infectious virions into vesicular fluid. Preliminary data indicate that: (i) Man 6-P is present on complex oligosaccharides of viral envelope gps. (ii) phosphorylated monosaccharides protect cells from infection by cell-free VZV in vitro; (iii) the rank order of efficacy (Man 6-P>>Glu 6-P>Glu 1-P) is the same as that for affinity for MPRs; (iv) chloroquine, which prevents recycling of free MPRs to the cell surface, reversibly protects cells from infection by VZV; (v) VZV accumulates in endosomes and the trans Golgi network of infected cells in culture and VZV gp immunoreactivity is co-distributed in cells with that of MPRs. We propose to definitively identify the compartments of the intracellular pathway of viral maturation. We will ascertain where VZV gps become incorporated into the viral envelope. We will determine the structures of VZV-associated oligosaccharides and how they are synthesized. We will establish if VZV-associated Man 6-P bearing oligosaccharides are sufficient to mediate binding of VZV to MPRs, we will assay the presence of Man 6-P on intact VZV virions obtained from vesicular fluid of cutaneous lesions, we will test the role of MPRs in mediating VZV infection of cultured cells, and we will investigate how VZV escapes intracellular degradation and reaches vesicular fluid in cutaneous lesions of patients with varicella.
| Gershon, Anne A; Gershon, Michael D (2010) Perspectives on vaccines against varicella-zoster virus infections. Curr Top Microbiol Immunol 342:359-72 |
| Gershon, Michael D; Gershon, Anne A (2010) VZV infection of keratinocytes: production of cell-free infectious virions in vivo. Curr Top Microbiol Immunol 342:173-88 |
