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DA Leib, SS Strand; Domains of the HSV-1 Virion Host Shutoff Protein Involved in Packaging and Pathogenesis . Invest. Ophthalmol. Vis. Sci. 2002;43(13):3857.
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Purpose: It has been demonstrated that the HSV-1 virion host shutoff (vhs) protein interacts with VP16 during infection and in the viral particle. The purpose of this study was to assess the role of this interaction in the lifecycle of HSV-1 in cell culture and in pathogenesis in a mouse ocular model. Methods: A recombinant virus was generated (termed F1) which lacked the VP16-interaction domain of vhs along with a marker-rescued virus (termed F1R). These viruses were tested in vitro for their abilities to replicate in cell culture, to induce RNA degradation as measured by northern blotting and to package vhs into the tegument as measured by western blotting. In mice, these viruses were tested for their abilities to replicate in the cornea and trigeminal ganglia and for their abilities to reactivate from dissociated latently infected trigeminal ganglia. Results: Both F1 and F1R replicated equivalently in cell culture. By northern blotting in the presence of actinomycin D (to inhibit de novo synthesis of vhs), F1 was completely unable to induce RNA degradation, while F1R was equivalent to wild-type virus, with high levels of degradation induced. In contrast, F1 and F1R in the absence of actinomycin D caused significant and equivalent RNA degradation. By western blotting F1 was unable to package significant levels of vhs compared to wild-type viruses. In mice, F1 was deficient for growth in the cornea and trigeminal ganglia, growth was intermediate compared to wild-type and vhs null viruses. Conclusion: Interaction with VP16 is necessary for the packaging of vhs into the tegument of HSV-1 and the lack of this interaction alters vhs activity during lytic replication. Packaged vhs is required for maximal lytic replication in vivo but is not required for growth in culture or for efficient reactivation. These data also support the idea that packaged vhs may play a role in the modulation of the immune response.
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