scholarly journals The Herpes Simplex Virus Type 1 UL17 Gene Encodes Virion Tegument Proteins That Are Required for Cleavage and Packaging of Viral DNA

1998 ◽  
Vol 72 (5) ◽  
pp. 3779-3788 ◽  
Author(s):  
Brandy Salmon ◽  
Charles Cunningham ◽  
Andrew J. Davison ◽  
Wendy J. Harris ◽  
Joel D. Baines
Keyword(s):  
Vero Cells ◽  
Open Reading Frame ◽  
Wild Type Virus ◽  
Type Virus ◽  
Wild Type ◽  
Viral Dna ◽  
Reading Frame ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACT Previous studies have suggested that the UL17 gene of herpes simplex virus type 1 (HSV-1) is essential for virus replication. In this study, viral mutants incorporating either a lacZexpression cassette in place of 1,490 bp of the 2,109-bp UL17 open reading frame [HSV-1(ΔUL17)] or a DNA oligomer containing an in-frame stop codon inserted 778 bp from the 5′ end of the UL17 open reading frame [HSV-1(UL17-stop)] were plaque purified on engineered cell lines containing the UL17 gene. A virus derived from HSV-1(UL17-stop) but containing a restored UL17 gene was also constructed and was designated HSV-1(UL17-restored). The latter virus formed plaques and cleaved genomic viral DNA in a manner indistinguishable from wild-type virus. Neither HSV-1(ΔUL17) nor HSV-1(UL17-stop) formed plaques or produced infectious progeny when propagated on noncomplementing Vero cells. Furthermore, genomic end-specific restriction fragments were not detected in DNA purified from noncomplementing cells infected with HSV-1(ΔUL17) or HSV-1(UL17-stop), whereas end-specific fragments were readily detected when the viruses were propagated on complementing cells. Electron micrographs of thin sections of cells infected with HSV-1(ΔUL17) or HSV-1(UL17-stop) illustrated that empty capsids accumulated in the nuclei of Vero cells, whereas DNA-containing capsids accumulated in the nuclei of complementing cells and enveloped virions were found in the cytoplasm and extracellular space. Additionally, protein profiles of capsids purified from cells infected with HSV-1(ΔUL17) compared to wild-type virus show no detectable differences. These data indicate that the UL17 gene is essential for virus replication and is required for cleavage and packaging of viral DNA. To characterize the UL17 gene product, an anti-UL17 rabbit polyclonal antiserum was produced. The antiserum reacted strongly with a major protein of apparent M r 77,000 and weakly with a protein of apparent M r 72,000 in wild-type infected cell lysates and in virions. Bands of similar sizes were also detected in electrophoretically separated tegument fractions of virions and light particles and yielded tryptic peptides of masses characteristic of the predicted UL17 protein. We therefore conclude that the UL17 gene products are associated with the virion tegument and note that they are the first tegument-associated proteins shown to be required for cleavage and packaging of viral DNA.

scholarly journals Assembly of Infectious Herpes Simplex Virus Type 1 Virions in the Absence of Full-Length VP22

2000 ◽  
Vol 74 (21) ◽  
pp. 10041-10054 ◽  
Author(s):  
Lisa E. Pomeranz ◽  
John A. Blaho
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Vero Cells ◽  
Wild Type Virus ◽  
Type Virus ◽  
Wild Type ◽  
Tegument Proteins ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACT VP22, the 301-amino-acid phosphoprotein product of the herpes simplex virus type 1 (HSV-1) UL49 gene, is incorporated into the tegument during virus assembly. We previously showed that highly modified forms of VP22 are restricted to infected cell nuclei (L. E. Pomeranz and J. A. Blaho, J. Virol. 73:6769–6781, 1999). VP22 packaged into infectious virions appears undermodified, and nuclear- and virion-associated forms are easily differentiated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (J. A. Blaho, C. Mitchell, and B. Roizman, J. Biol. Chem. 269:17401–17410, 1994). As VP22 packaging-associated undermodification is unique among HSV-1 tegument proteins, we sought to determine the role of VP22 during viral replication. We now show the following. (i) VP22 modification occurs in the absence of other viral factors in cell lines which stably express its gene. (ii) RF177, a recombinant HSV-1 strain generated for this study, synthesizes only the amino-terminal 212 amino acids of VP22 (Δ212). (iii) Δ212 localizes to the nucleus and incorporates into virions during RF177 infection of Vero cells. Thus, the carboxy-terminal region is not required for nuclear localization of VP22. (iv) RF177 synthesizes the tegument proteins VP13/14, VP16, and VHS (virus host shutoff) and incorporates them into infectious virions as efficiently as wild-type virus. However, (v) the loss of VP22 in RF177 virus particles is compensated for by a redistribution of minor virion components. (vi) Mature RF177 virions are identical to wild-type particles based on electron microscopic analyses. (vii) Single-step growth kinetics of RF177 in Vero cells are essentially identical to those of wild-type virus. (viii) RF177 plaque size is reduced by nearly 40% compared to wild-type virus. Based on these results, we conclude that VP22 is not required for tegument formation, virion assembly/maturation, or productive HSV-1 replication, while the presence of full-length VP22 in the tegument is needed for efficient virus spread in Vero cell monolayers.

scholarly journals Point Mutations in Herpes Simplex Virus Type 1 oriL, but Not in oriS, Reduce Pathogenesis during Acute Infection of Mice and Impair Reactivation from Latency

2006 ◽  
Vol 80 (1) ◽  
pp. 440-450 ◽  
Author(s):  
John W. Balliet ◽  
Priscilla A. Schaffer
Keyword(s):  
Herpes Simplex ◽  
Point Mutations ◽  
Tear Film ◽  
Wild Type Virus ◽  
Type Virus ◽  
Wild Type ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACT In vitro studies of herpes simplex virus type 1 (HSV-1) viruses containing mutations in core sequences of the viral origins of DNA replication, oriL and oriS, that eliminate the ability of these origins to initiate viral-DNA synthesis have demonstrated little or no effect on viral replication in cultured cells, leading to the conclusion that the two types of origins are functionally redundant. It remains unclear, therefore, why origins that appear to be redundant are maintained evolutionarily in HSV-1 and other neurotropic alphaherpesviruses. To test the hypothesis that oriL and oriS have distinct functions in the HSV-1 life cycle in vivo, we determined the in vivo phenotypes of two mutant viruses, DoriL-ILR and DoriS-I, containing point mutations in oriL and oriS site I, respectively, that eliminate origin DNA initiation function. Following corneal inoculation of mice, tear film titers of DoriS-I were reduced relative to wild-type virus. In all other tests, however, DoriS-I behaved like wild-type virus. In contrast, titers of DoriL-ILR in tear film, trigeminal ganglia (TG), and hindbrain were reduced and mice infected with DoriL-ILR exhibited greatly reduced mortality relative to wild-type virus. In the TG explant and TG cell culture models of reactivation, DoriL-ILR reactivated with delayed kinetics and, in the latter model, with reduced efficiency relative to wild-type virus. Rescuant viruses DoriL-ILR-R and DoriS-I-R behaved like wild-type virus in all tests. These findings demonstrate that functional differences exist between oriL and oriS and reveal a prominent role for oriL in HSV-1 pathogenesis.

scholarly journals A Novel Herpes Simplex Virus Type 1 Transcript (AL-RNA) Antisense to the 5′ End of the Latency-Associated Transcript Produces a Protein in Infected Rabbits

2002 ◽  
Vol 76 (16) ◽  
pp. 8003-8010 ◽  
Author(s):  
Guey-Chuen Perng ◽  
Barak Maguen ◽  
Ling Jin ◽  
Kevin R. Mott ◽  
John Kurylo ◽  
...  
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Herpes Simplex Virus Type ◽  
Open Reading Frame ◽  
Wild Type ◽  
Reading Frame ◽  
Viral Virulence ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACT Following primary ocular infection, herpes simplex virus type 1 (HSV-1) establishes a lifelong latent infection in sensory neurons of the trigeminal ganglia. Latency-associated transcript (LAT), the only known viral gene abundantly transcribed during HSV-1 neuronal latency, is required for high levels of reactivation. Recently we showed that three different mutants that do not alter the LAT promoter but contain deletions within the 5′ end of the primary LAT transcript affect viral virulence (G. C. Perng et al., J. Virol. 75:9018-9028, 2001). In contrast, in LAT-null mutants viral virulence appears unaltered (T. M. Block et al., Virology 192:618-630, 1993; D. C. Bloom et al., J. Virol. 68:1283-1292, 1994; J. M. Hill et al., Virology 174:117-125, 1990; G. C. Perng et al., J. Virol. 68:8045-8055, 1994; F. Sedarati, K. M. Izumi, E. K. Wagner, and J. G. Stevens, J. Virol. 63:4455-4458, 1989). We therefore hypothesized that the 5′ end of LAT and/or an as yet unidentified gene that overlaps part of this region is involved in viral virulence. We report here on the discovery and initial characterization of a novel HSV-1 RNA consistent with such a putative gene. The novel RNA was antisense to the 5′ end of LAT and was designated AL-RNA (anti-LAT sense RNA). The AL-RNA overlapped the core LAT promoter and the first 158 nucleotides of the 5′ end of the primary LAT transcript. AL-RNA was detected in extracts from neuron-like cells (PC-12) infected with wild-type HSV-1 but not in cells infected with a mutant with the AL region deleted. The deletions in each of the above three mutants with altered virulence encompass the 5′ end of the AL-RNA, and these mutants cannot transcribe AL. This supports the hypothesis that the AL gene may play a role in viral virulence. Based on comparison to the corresponding genomic sequence, the AL-RNA did not appear to be spliced. The AL-RNA was polyadenylated and contained an open reading frame capable of encoding a protein 56 amino acids in length with a predicted molecular mass of 6.8 kDa. Sera from three of three rabbits infected with wild-type HSV-1 but not sera from any of three rabbits infected with a mutant with the AL-RNA region deleted recognized the Escherichia coli recombinantly expressed AL open reading frame on Western blots. In addition, four of six rabbits infected with wild-type virus developed enzyme-linked immunosorbent assay titers against one or more AL synthetic peptides. These results suggest that an AL protein is produced in vivo.

scholarly journals Recombinant Herpes Simplex Virus Type 1 Expressing Murine Interleukin-4 Is Less Virulent than Wild-Type Virus in Mice

2001 ◽  
Vol 75 (19) ◽  
pp. 9029-9036 ◽  
Author(s):  
Homayon Ghiasi ◽  
Yanira Osorio ◽  
Guey-Chuen Perng ◽  
Anthony B. Nesburn ◽  
Steven L. Wechsler
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Herpes Simplex Virus Type ◽  
Interleukin 4 ◽  
Wild Type Virus ◽  
Type Virus ◽  
Wild Type ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACT The effect of interleukin-4 (IL-4) on herpes simplex virus type 1 (HSV-1) infection in mice was evaluated by construction of a recombinant HSV-1 expressing the gene for murine IL-4 in place of the latency-associated transcript (LAT). The mutant virus (HSV-IL-4) expressed high levels of IL-4 in cultured cells. The replication of HSV-IL-4 in tissue culture and in trigeminal ganglia was similar to that of wild-type virus. In contrast, HSV-IL-4 appeared to replicate less well in mouse eyes and brains. Although BALB/c mice are highly susceptible to HSV-1 infection, ocular infection with HSV-IL-4 resulted in 100% survival. Furthermore, 57% of the mice survived coinfection with a mixture of HSV-IL-4 and a lethal dose of wild-type McKrae, compared with only 10% survival following infection with McKrae alone. Similar to wild-type BALB/c mice, 100% of IL-4−/− mice also survived HSV-IL-4 infection. T-cell depletion studies suggested that protection against HSV-IL-4 infection was mediated by a CD4+-T-cell response.

scholarly journals Functional Genomic Analysis of Herpes Simplex Virus Type 1 Counteraction of the Host Innate Response

2006 ◽  
Vol 80 (15) ◽  
pp. 7600-7612 ◽  
Author(s):  
Tracy Jo Pasieka ◽  
Tracey Baas ◽  
Victoria S. Carter ◽  
Sean C. Proll ◽  
Michael G. Katze ◽  
...  
Keyword(s):  
Herpes Simplex ◽  
Antiviral Response ◽  
Wild Type Virus ◽  
Type Virus ◽  
Wild Type ◽  
Mouse Embryo Fibroblasts ◽  
Infected Cells ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACTHerpes simplex virus type 1 (HSV-1) mutants lacking the ICP34.5 gene are severely attenuated in mouse models and have a significant growth defect in confluent mouse embryo fibroblasts. Previously, ICP34.5 was demonstrated to have a crucial role in evading the innate immune response to infection by mediating the dephosphorylation of eIF2α, a translation initiation factor phosphorylated by PKR during the antiviral response. To further understand the role of ICP34.5 in evasion of the antiviral response, we used transcriptional profiling to examine host cell gene expression in both wild-type and ICP34.5-null virus-infected mouse embryo fibroblasts over a time course of infection. Our study revealed that cells responded to infection within 3 h through PKR-dependent eIF2α phosphorylation and that the majority of up-regulated genes at 3 h postinfection were involved in the antiviral response. HSV-1 counters this response through early expression of ICP34.5 and dephosphorylation of eIF2α. By 12 h postinfection, the differences between the number and functional classification of genes differentially up- and down-regulated between wild-type and ICP34.5-null virus-infected cells were maximal. Specifically, in wild-type virus-infected cells, the majority of changed genes were involved in metabolic and biosynthetic processes, while in ICP34.5-null virus-infected cells, mostly antiviral genes were up-regulated. Further, ICP34.5-null virus-infected cells produced greater amounts of beta interferon than wild-type virus-infected cells. These results indicate that ICP34.5 expression and function at early times postinfection have a pivotal role in the ability of HSV-1 to gain control of the host cell and maintain an environment for successful viral replication.

scholarly journals Diverse Herpes Simplex Virus Type 1 Thymidine Kinase Mutants in Individual Human Neurons and Ganglia

2007 ◽  
Vol 81 (13) ◽  
pp. 6817-6826 ◽  
Author(s):  
Kening Wang ◽  
Gowtham Mahalingam ◽  
Susan E. Hoover ◽  
Erik K. Mont ◽  
Steven M. Holland ◽  
...  
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Herpes Simplex Virus Type ◽  
Wild Type Virus ◽  
Type Virus ◽  
Wild Type ◽  
Mixed Populations ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACT Mutations in the thymidine kinase gene (tk) of herpes simplex virus type 1 (HSV-1) explain most cases of virus resistance to acyclovir (ACV) treatment. Mucocutaneous lesions of patients with ACV resistance contain mixed populations of tk mutant and wild-type virus. However, it is unknown whether human ganglia also contain mixed populations since the replication of HSV tk mutants in animal neurons is impaired. Here we report the detection of mutated HSV tk sequences in human ganglia. Trigeminal and dorsal root ganglia were obtained at autopsy from an immunocompromised woman with chronic mucocutaneous infection with ACV-resistant HSV-1. The HSV-1 tk open reading frames from ganglia were amplified by PCR, cloned, and sequenced. tk mutations were detected in a seven-G homopolymer region in 11 of 12 ganglia tested, with clonal frequencies ranging from 4.2 to 76% HSV-1 tk mutants per ganglion. In 8 of 11 ganglia, the mutations were heterogeneous, varying from a deletion of one G to an insertion of one to three G residues, with the two-G insertion being the most common. Each ganglion had its own pattern of mutant populations. When individual neurons from one ganglion were analyzed by laser capture microdissection and PCR, 6 of 14 HSV-1-positive neurons were coinfected with HSV tk mutants and wild-type virus, 4 of 14 were infected with wild-type virus alone, and 4 of 14 were infected with tk mutant virus alone. These data suggest that diverse tk mutants arise independently under drug selection and establish latency in human sensory ganglia alone or together with wild-type virus.

scholarly journals The Product of the Herpes Simplex Virus Type 1 UL25 Gene Is Required for Encapsidation but Not for Cleavage of Replicated Viral DNA

1998 ◽  
Vol 72 (2) ◽  
pp. 1060-1070 ◽  
Author(s):  
Alistair R. McNab ◽  
Prashant Desai ◽  
Stan Person ◽  
Lori L. Roof ◽  
Darrell R. Thomsen ◽  
...  
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Virus Type ◽  
Herpes Simplex Virus Type ◽  
Vero Cells ◽  
Open Reading Frame ◽  
Viral Dna ◽  
Reading Frame ◽  
Simplex Virus

ABSTRACT The herpes simplex virus type 1 (HSV-1) UL25 gene contains a 580-amino-acid open reading frame that codes for an essential protein. Previous studies have shown that the UL25 gene product is a virion component (M. A. Ali et al., Virology 216:278–283, 1996) involved in virus penetration and capsid assembly (C. Addison et al., Virology 138:246–259, 1984). In this study, we describe the isolation of a UL25 mutant (KUL25NS) that was constructed by insertion of an in-frame stop codon in the UL25 open reading frame and propagated on a complementing cell line. Although the mutant was capable of synthesis of viral DNA, it did not form plaques or produce infectious virus in noncomplementing cells. Antibodies specific for the UL25 protein were used to demonstrate that KUL25NS-infected Vero cells did not express the UL25 protein. Western immunoblotting showed that the UL25 protein was associated with purified, wild-type HSV A, B, and C capsids. Transmission electron microscopy indicated that the nucleus of Vero cells infected with KUL25NS contained large numbers of both A and B capsids but no C capsids. Analysis of infected cells by sucrose gradient sedimentation analysis confirmed that the ratio of A to B capsids was elevated in KUL25NS-infected Vero cells. Following restriction enzyme digestion, specific terminal fragments were observed in DNA isolated from KUL25NS-infected Vero cells, indicating that the UL25 gene was not required for cleavage of replicated viral DNA. The latter result was confirmed by pulsed-field gel electrophoresis (PFGE), which showed the presence of genome-size viral DNA in KUL25NS-infected Vero cells. DNase I treatment prior to PFGE demonstrated that monomeric HSV DNA was not packaged in the absence of the UL25 protein. Our results indicate that the product of the UL25 gene is required for packaging but not cleavage of replicated viral DNA.

scholarly journals The Herpes Simplex Virus Type 1 US11 Protein Binds the Coterminal UL12, UL13, and UL14 RNAs and Regulates UL13 Expression In Vivo

2002 ◽  
Vol 76 (16) ◽  
pp. 8090-8100 ◽  
Author(s):  
Helen L. Attrill ◽  
Sarah A. Cumming ◽  
J. Barklie Clements ◽  
Sheila V. Graham
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Wild Type Virus ◽  
Type Virus ◽  
Wild Type ◽  
Infected Cells ◽  
Simplex Virus ◽  
Us11 Protein ◽  
Hsv 1

ABSTRACT The US11 protein of herpes simplex virus type 1 (HSV-1) is a small, highly basic phosphoprotein expressed at late times during infection. US11 localizes to the nucleolus in infected cells, can associate with ribosomes, and has been shown to bind RNA. The RNA substrates of US11 identified thus far have no apparent role in the virus lytic cycle, so we set out to identify a novel, biologically relevant RNA substrate(s) for this protein in HSV-1-infected cells. We designed a reverse transcriptase PCR-based protocol that allowed specific selection of a 600-bp RNA binding partner for US11. This RNA sequence, designated 12/14, is present in the coterminal HSV-1 mRNAs UL12, UL13, and UL14. We show that the binding of US11 to 12/14 is sequence-specific and mediated by the C-terminal domain of the protein. To elucidate the role of US11 in the virus life cycle, we infected cells with wild-type virus, a cosmid-reconstructed US11 HSV-1 null mutant, and a cosmid-reconstructed wild-type virus and analyzed expression of UL12, -13, and -14 during a time course of infection. These experiments revealed that this interaction has biological activity; at early times of infection, US11 down-regulates UL13 protein kinase mRNA and protein.

scholarly journals Cell-to-Cell Spread of Wild-Type Herpes Simplex Virus Type 1, but Not of Syncytial Strains, Is Mediated by the Immunoglobulin-Like Receptors That Mediate Virion Entry, Nectin1 (PRR1/HveC/HIgR) and Nectin2 (PRR2/HveB)

2000 ◽  
Vol 74 (8) ◽  
pp. 3909-3917 ◽  
Author(s):  
Francesca Cocchi ◽  
Laura Menotti ◽  
Patrice Dubreuil ◽  
Marc Lopez ◽  
Gabriella Campadelli-Fiume
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Wild Type Virus ◽  
Human Cell Lines ◽  
Type Virus ◽  
Wild Type ◽  
Simplex Virus ◽  
Cell Spread ◽  
Hsv 1

ABSTRACT The immunoglobulin-like receptors that mediate entry of herpes simplex virus type 1 (HSV-1) into human cells were found to mediate the direct cell-to-cell spread of wild-type virus. The receptors here designated Nectin1α and -δ and Nectin2α were originally designated HIgR, PRR1/HveC, and PRR2α/HveB, respectively. We report the following. (i) Wild-type HSV-1 spreads from cell to cell in J cells expressing nectin1α or nectin1δ but not in parental J cells that are devoid of entry receptors. A monoclonal antibody to nectin1, which blocks entry, also blocked cell-to-cell spread in nectin1-expressing J cells. Moreover, wild-type virus did not spread from a receptor-positive to a receptor-negative cell. (ii) The antibody to nectin1 blocked transmission of wild-type virus in a number of human cell lines, with varying efficiencies, suggesting that nectin1 is the principal mediator of wild-type virus spread in a variety of human cell lines. (iii) Nectin1 did not mediate cell fusion induced by the syncytial strains HSV-1(MP) and HFEM-syn. (iv) Nectin2α could serve as a receptor for spread of a mutant virus carrying the L25P substitution in glycoprotein D, but not of wild-type virus, in agreement with its ability to mediate entry of the mutant but not of wild-type virus.

scholarly journals The Conserved Carboxyl-Terminal Half of Herpes Simplex Virus Type 1 Regulatory Protein ICP27 Is Dispensable for Viral Growth in the Presence of Compensatory Mutations

2000 ◽  
Vol 74 (16) ◽  
pp. 7362-7374 ◽  
Author(s):  
Scott M. Bunnell ◽  
Stephen A. Rice
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Herpes Simplex Virus Type ◽  
Vero Cells ◽  
Terminal Portion ◽  
Viral Dna ◽  
Infected Cells ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACT ICP27 is an essential herpes simplex virus type 1 (HSV-1) immediate-early protein that regulates viral gene expression by poorly characterized mechanisms. Previous data suggest that its carboxyl (C)-terminal portion is absolutely required for productive viral infection. In this study, we isolated M16R, a second-site revertant of a viral ICP27 C-terminal mutant. M16R harbors an intragenic reversion, as demonstrated by the fact that its cloned ICP27 allele can complement the growth of an HSV-1 ICP27 deletion mutant. DNA sequencing demonstrated that the intragenic reversion is a frameshift alteration in a homopolymeric run of C residues at codons 215 to 217. This results in the predicted expression of a truncated, 289-residue molecule bearing 72 novel C-terminal residues derived from the +1 reading frame. Consistent with this, M16R expresses an ICP27-related molecule of the predicted size in the nuclei of infected cells. Transfection-based viral complementation assays confirmed that the truncated, frameshifted protein can partially substitute for ICP27 in the context of viral infection. Surprisingly, its novel C-terminal residues are required for this activity. To see if the frameshift mutation is all that is required for M16R's viability, we re-engineered the M16R ICP27 allele and inserted it into a new viral background, creating the HSV-1 mutant M16exC. An additional mutant, exCd305, was constructed which possesses the frameshift in the context of an ICP27 gene with the C terminus deleted. We found that both M16exC and exCd305 are nonviable in Vero cells, suggesting that one or more extragenic mutations are also required for the viability of M16R. Consistent with this interpretation, we isolated two viable derivatives ofexCd305 which grow productively in Vero cells despite being incapable of encoding the C-terminal portion of ICP27. Studies of viral DNA synthesis in mutant-infected cells indicated that the truncated, frameshifted ICP27 protein can enhance viral DNA replication. In summary, our results demonstrate that the C-terminal portion of ICP27, conserved widely in herpesviruses and previously believed to be absolutely essential, is dispensable for HSV-1 lytic replication in the presence of compensatory genomic mutations.

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