Role of the VP16-Binding Domain of vhs in Viral Growth, Host Shutoff Activity, and Pathogenesis

Abstract

ABSTRACT The virion host shutoff (vhs) protein of herpes simplex virus type 1 causes the degradation of host and viral mRNA immediately upon infection of permissive cells. vhs can interact with VP16 through a 20-amino-acid binding domain, and viruses containing a deletion of this VP16-binding domain of vhs (Δ20) and a corresponding marker rescue (Δ20R) were constructed and characterized. Transient-transfection assays showed that this domain was dispensable for vhs activity. The Δ20 recombinant virus, however, was unable to induce mRNA degradation in the presence of actinomycin D, while degradation induced by Δ20R was equivalent to that for wild-type virus. Δ20, Δ20R, and KOS caused comparable RNA degradation in the absence of actinomycin D. Western blot analysis of infected cells indicated that comparable levels of vhs were expressed by Δ20, Δ20R, and KOS, and there was only a modest reduction of vhs packaging in Δ20. Immunoprecipitation of protein from cells infected with Δ20 and Δ20R showed equivalent coprecipitation of vhs and VP16. Pathogenesis studies with Δ20 showed a significant decrease in replication in the corneas, trigeminal ganglia, and brains, as well as a significant reduction in clinical disease and lethality, but no significant difference in the establishment of, or reactivation from, latency compared to results with KOS and Δ20R. These results suggest that the previously described VP16-binding domain is not required for vhs packaging or for binding to VP16. It is required, however, for RNA degradation activity of tegument-derived vhs and wild-type replication and virulence in mice.