VP22 of Herpes Simplex Virus 1 Promotes Protein Synthesis at Late Times in Infection and Accumulation of a Subset of Viral mRNAs at Early Times in Infection

Herpes simplex virus 1 (HSV-1) virions are composed of a nucleocapsid enclosing the double-stranded linear DNA genome, a proteinaceous layer termed the tegument that surrounds the nucleocapsid, and a host-derived lipid membrane envelope that contains viral glycoproteins. The tegument is unique to herpesviruses and is composed of at least 20 different viral proteins of varied stoichiometries. Tegument proteins have been shown to play a variety of roles in infection, including the regulation of viral and host gene expression and the promotion of virus assembly and egress (1, 16, 34, 38). Tegument proteins enter the cell upon fusion of the viral envelope with the host cell membrane during infection initiation. Thus, these proteins provide a potential means to modulate the host cell and advance viral infection at both very early times in infection upon tegument delivery and at late times in infection, when the tegument proteins are produced in high amounts. VP22, encoded by the UL49 gene, is one of the most abundant HSV-1 tegument proteins, with an average stoichiometry of 2,000 copies per virion. A number of functions have been attributed to VP22, including association with and reorganization of microtubules in infected and uninfected cells (10, 22) and incorporation of RNA into the virion (42). In previous studies, UL49 truncation or deletion mutants propagated on VP22-expressing cell lines were shown to be significantly debilitated in production of infectious virus on noncomplementing cells at low multiplicities of infection and produced plaques of significantly smaller size (6, 35). Further characterization revealed significant defects in release of infectious virus into the medium in the absence of VP22, thereby decreasing viral spread (6). Virions produced by this UL49− virus upon infection of noncomplementing cells contained decreased levels of ICP0, gD, and gE. These observations are potentially consistent with reports that VP22 interacts with the cytoplasmic tails of gD and gE (4, 14, 33) and is necessary for localization of ICP0 to putative sites of viral assembly within the cytoplasm (8). In contrast to the small plaque phenotype displayed by the UL49− virus on noncomplementing cells after propagation on complementing cells, passage of the UL49− virus on noncomplementing cells rapidly and consistently caused the normally defective virus to produce plaques of wild-type size (5). These data suggest a strong selection for a secondary compensatory mutation(s) that rescues the UL49− small plaque phenotype. Also pertinent to this report is the observation that VP22 interacts with VP16 (9). VP16 was one of the first transcription factors described and binds host octamer transcription factor 1 and other host factors to mediate transcription from viral immediate-early promoters expressed in the absence of viral protein production (23-26, 40). Thus, VP16 within the tegument is delivered to the nucleus upon initiation of infection to mediate preferential transcription of viral mRNAs over those of the host. Previous studies indicated that VP16 and the VP16/VP22 complex can interact with the virion host shutoff protein (vhs) (46). vhs, encoded by the UL41 gene, is a riboendonuclease that specifically cleaves host and viral mRNAs (7, 13, 27, 28, 38, 45, 48). The RNase activity of vhs is believed to promote expression of viral proteins early in infection by degrading host mRNAs. Likewise, vhs is believed to promote the transition from the early phase of viral transcription to the late phase by degrading early viral mRNAs at the appropriate time in infection. In contrast, the production of VP16 late in infection suppresses the RNase activity of vhs (29, 41, 44), suggesting a mechanism to ensure appropriate accumulation of late transcripts. Interestingly, a previously described UL49-null mutant derived from an HSV-1 bacterial artificial chromosome accumulated deletion and frameshift mutations in UL41 when propagated on noncomplementing cells (43), suggesting the presence of vhs was detrimental in the absence of VP22. The present studies were initiated to further characterize the phenotype of the UL49− virus we described previously (6). The data presented herein show VP22 is required for optimal protein synthesis at late times in infection and the accumulation of at least gE, gD, and vhs mRNAs at early times in infection. Interestingly, the protein and mRNA phenotypes are separable with regard to both timing during infection and the genes affected, suggesting separate roles for VP22 in both protein and mRNA accumulation. Moreover, these effects were mediated in the absence of mutations in VP16 or vhs.