Functional inaccessibility of quiescent herpes simplex virus genomes
© Minaker et al; licensee BioMed Central Ltd. 2005
Received: 21 September 2005
Accepted: 21 November 2005
Published: 21 November 2005
Newly delivered herpes simplex virus genomes are subject to repression during the early stages of infection of human fibroblasts. This host defence strategy can limit virus replication and lead to long-term persistence of quiescent viral genomes. The viral immediate-early protein ICP0 acts to negate this negative regulation, thereby facilitating the onset of the viral replication cycle. Although few mechanistic details are available, the host repression machinery has been proposed to assemble the viral genome into a globally inaccessible configuration analogous to heterochromatin, blocking access to most or all trans-acting factors. The strongest evidence for this hypothesis is that ICP0-deficient virus is unable to reactivate quiescent viral genomes, despite its ability to undergo productive infection given a sufficiently high multiplicity of infection. However, recent studies have shown that quiescent infection induces a potent antiviral state, and that ICP0 plays a key role in disarming such host antiviral responses. These findings raise the possibility that cells containing quiescent viral genomes may be refractory to superinfection by ICP0-deficient virus, potentially providing an alternative explanation for the inability of such viruses to trigger reactivation. We therefore asked if ICP0-deficient virus is capable of replicating in cells that contain quiescent viral genomes.
We found that ICP0-deficient herpes simplex virus is able to infect quiescently infected cells, leading to expression and replication of the superinfecting viral genome. Despite this productive infection, the resident quiescent viral genome was neither expressed nor replicated, unless ICP0 was provided in trans.
These data document that quiescent HSV genomes fail to respond to the virally modified host transcriptional apparatus or viral DNA replication machinery provided in trans by productive HSV infection in the absence of ICP0. These results point to global repression as the basis for HSV genome quiescence, and indicate that ICP0 induces reactivation by overcoming this global barrier to the access of trans-acting factors.
Herpes simplex virus (HSV) is a significant human pathogen and the prototypical member of the herpesviridae, a large family of enveloped nuclear DNA viruses. HSV displays two modes of interaction with its human host: lytic and latent (reviewed in ). Primary infection of epithelial cells produces the lytic response – productive virus replication followed by cell death. Progeny virions then infect adjacent sensory neurons, establishing a life-long latent interaction. Productive infection is characterized by the sequential expression of three classes of viral genes, immediate-early (IE), early (E) and late (L). This regulatory cascade is initiated by VP16, an abundant tegument protein that activates transcription of the IE genes. Four of the IE proteins (ICP0, ICP4, ICP22 and ICP27) then serve to drive further progression into the lytic program. Three of these, ICP4, ICP22 and ICP27, contribute in various ways to the activation of the E and/or L genes . The role of ICP0 appears to be distinct, in that it is also required for efficient IE gene expression [2–4]. Thus, ICP0 mutant viruses display reduced levels of IE gene expression during infection [3–5], and ICP0 activates expression of IE, E and L genes in trans ient transfection assays [6–10]. Moreover, expression of ICP0 in trans substantially complements the defect of VP16 mutants [11, 12], which are otherwise arrested prior to the IE phase following low multiplicity infection. The function of ICP0 therefore seems to lie upstream of those of the other IE gene products in the HSV regulatory cascade.
ICP0 has been described as a promiscuous activator capable of stimulating the expression of a wide range of viral and cellular promoters in transient co-transfection assays (reviewed in ). It acts to enhance mRNA accumulation, at least in part by stimulating transcription [14, 15]. However it does not bind DNA and there is no evidence that it acts directly on the transcriptional apparatus. Rather, ICP0 appears to stimulate HSV gene expression at least in part by counteracting one or more cellular repression mechanisms that otherwise silence newly delivered viral genomes (reviewed in ). This hypothesis emerged from the finding that viral genomes unable to express ICP0 often fail to engage the viral lytic program of gene expression and instead persist for extended periods in the nucleus in an extrachromosomal non-linear configuration without giving rise to appreciable levels of viral gene products [17–21]. Such quiescent genomes however remain potentially functional, as they can be efficiently reactivated by superinfecting the cultures with HSV or human cytomegalovirus (HCMV, another herpesvirus) or by providing ICP0 or HCMV pp71 in trans [17–19, 22–24]. The IE promoters residing in quiescent HSV genomes appear to be repressed rather than simply inactive, as they fail to respond to VP16 and several other stimuli that otherwise augment their activity ; however, they remain susceptible to activation by ICP0 or pp71 [18, 23]. Repression of genomes entering quiescence occurs gradually: newly delivered IE promoters are initially responsive to VP16 and other stimuli and are only later rendered refractory to stimuli other than ICP0 . Perhaps unexpectedly, the otherwise constitutively active HCMV IE promoter is also repressed as recombinant HSV genomes enter quiescence [18–20]. Taken in combination, these data suggest that newly delivered HSV and HCMV IE promoters are targeted by a cellular repression mechanism that is inactivated by ICP0. HSV E and L promoters are also inactive during quiescence; however it is not yet clear if they are actively repressed like the IE promoters or simply inactive due to the absence of the IE proteins.
The mechanisms underlying repression and reactivation of quiescent HSV genomes remain unclear. ICP0 interacts with numerous cellular proteins (reviewed in ) including some that could plausibly contribute to gene silencing (for example, type II histone deacetylases  and the coREST/REST repressor complex ). In addition, ICP0 bears a RING-finger E3 ubiquitin ligase domain [28–30] that is essential for reactivation , suggesting that it may act at least in part by targeting key components of the cellular repression machinery for ubiquitination and degradation. Consistent with this view, reactivation is blocked by proteasome inhibitors . However, the crucial target(s) of ICP0 relevant to reactivation have yet to be defined. It may be significant that infecting HSV genomes initially localize to the periphery of nuclear ND10 domains [31–33], and that ICP0 disrupts ND10 [34–36] by targeting several components, including PML, for destruction [37–39]. However, the intranuclear location of quiescent genomes has yet to be determined, and current evidence suggests that transcriptional activity is required for the association of viral genomes with ND10 . Thus, it is not clear what, if any, role ND10 play in quiescence.
A remarkable feature of quiescent HSV genomes is that they fail to detectably respond to superinfection with ICP0-deficient HSV [22, 40–42]. The result is striking because ICP0-deficient HSV is itself capable of productively infecting many cell types including those used to establish quiescence, giving rise to infectious progeny. One interpretation of these data is that quiescent HSV genomes are inaccessible to the virally modified transcriptional apparatus and HSV DNA replication machinery provided in trans by the superinfecting virus in the absence of ICP0 [16, 41]. If this interpretation is correct, then it follows that: (1) quiescence involves a global restriction in the accessibility of the viral genome to trans-acting factors perhaps akin to that associated with the heterochromatinization of silent host chromosomal loci, and (2) ICP0 induces reactivation by overcoming this generalized barrier to genome activity. However, another hypothesis to explain the inability of ICP0-deficient viruses to induce reactivation is that ICP0 may be required for productive infection of cells harboring quiescent HSV. Under this alternative scenario, ICP0-deficient HSV is effectively excluded from the cells harboring the resident virus, thereby precluding genome reactivation. This "superinfection-immunity" model has not been examined in previous studies; however several considerations suggest that it should be carefully evaluated. First, the severity of the phenotype of ICP0-deficient mutants varies markedly between cell types  and during cell cycle progression , raising the possibility that such mutants may be unusually sensitive to any perturbations of cellular physiology induced by quiescent HSV infection. Second, the data of Hobbs et al  indicate that the replication of ICP0-deficient HSV is severely compromised under the conditions used by those authors in their reactivation assays. Third, HSV virions trigger the induction of a potent antiviral state associated with activation of a subset of IFN-inducible genes in human fibroblasts under conditions where viral gene expression is prevented [45–48], as in quiescence. Moreover, ICP0 serves to block this cellular antiviral response , by preventing the activation of IRF3 through unknown mechanisms . Consistent with this particular mechanism of superinfection immunity, ICP0 mutants are hypersensitive to the antiviral effects of type I IFN [50–52] and thus might also be expected to be unusually sensitive to the IFN-independent antiviral state provoked by HSV virions. Fourth, it is possible that quiescent HSV itself gives rise to one or more gene products that interfere with replication of superinfecting ICP0-deficient HSV in a fashion analogous to the repressors produced by temperate bacteriophages.
Considering the foregoing, we examined the susceptibility of human embryonic lung (HEL) fibroblasts harboring quiescent HSV-1 genomes to productive superinfection by ICP0-deficient HSV. We found that such cells are capable of supporting expression and replication of superinfecting ICP0-deficient genomes, given a sufficiently high input multiplicity of infection (MOI). However, the resident quiescent viral genomes were not detectably expressed or replicated in these superinfected cultures. Our results therefore rule out the superinfection-immunity model for the inability of ICP0-deficient HSV to reactivate quiescent HSV, and document that quiescent HSV genomes fail to respond to the virally modified host transcriptional apparatus or viral DNA replication machinery during productive HSV infection in the absence of ICP0. These results point to global repression as the basis for HSV genome quiescence, and indicate that ICP0 induces reactivation by overcoming this global barrier to trans-acting factors.
Results and Discussion
ICP0 is specifically required for reactivation of gene expression from quiescent HSV-1 KM110-R genomes
ICP0 is required for reactivation of viral DNA replication
Previous work has implied that ICP0 is required for replication of the resident viral genome following superinfection of cells harboring quiescent HSV [40, 41]. To determine if this is the case in our system, we used Southern blot hybridization to monitor replication of the resident KM110-R genome following superinfection with wild-type and mutant virus (figure 2B). The genome of KM110-R can be readily distinguished from that of wild-type HSV-1 because it bears an NheI linker at the VP16 locus that marks the V422 mutation . As a result, the 8.1 kb BamHI fragment that spans the VP16 locus is cleaved by NheI in KM110-R, yielding fragments of 4.9 and 3.2 kb (figure 2B). Quiescent KM110-R genomes were not detectable prior to reactivation under the conditions used in our Southern blot assay; however the expected KM110-R signal was readily detected following genome amplification induced by super-infection with wild-type KOS (figure 2B). Mutants lacking ICP4, ICP22, and ICP27 (d120, d22-lacZ and d27-1 respectively) triggered replication of the KM110-R genome as effectively as wild-type KOS; in contrast, no amplified KM110-R signal was observed following infection with the ICP0-deficient mutant n212. As expected , the ICP4 and ICP27 null mutants each displayed a severe DNA replication defect in cells lacking KM1110-R. These defects were however complemented in cells harboring KM110-R, presumably due to provision of the missing gene products in trans from the reactivated KM110-R genome.
Taken in combination, the data presented above clearly document that ICP0 is essential for the reactivation of expression from the HCMV IE and HSV VP16 promoters and replication of quiescent HSV-1 genomes in superinfected cultures, confirming and extending the results of previous studies.
ICP0-deficient HSV-1 is able to infect cells that harbor quiescent KM110-R
Viral progeny recovered from superinfected cells. HEL cells containing or lacking quiescent KM110-R (MOI 6) were superinfected on day 4 with either KOS or n212 (MOIs of 10 and 30 respectively). Progeny virus harvested 18 hours later was then titrated on U2OS cells in the presence of HMBA (Methods).
RFP- titre (PFU/mL)
RFP+ titre (PFU/mL)
1.25 × 103 ± 1.5 × 103
3.66 × 107 ± 1.3 × 107
1.79 × 107 ± 8.6 × 106
1.00 × 104 ± 6.8 × 103
5.52 × 107 ± 2.2 × 107
2.63 × 107 ± 3.9 × 106
8.30 × 106 ± 7.4 × 105
Summary and implications
Our results document that ICP0-deficient HSV is capable of productively infecting cells that harbor quiescent HSV genomes: given a sufficiently high multiplicity of infection the superinfecting virus initiates gene expression and progresses to at least the stage of viral DNA replication in the majority of such cells. Remarkably, this productive infection does not provoke reactivation of the resident viral genomes. These data exclude the superinfection-immunity model for the failure of ICP0-deficient HSV to trigger reactivation and provide strong support for the suggestion that quiescent HSV genomes are functionally inaccessible to the modified transcription apparatus and viral DNA replication factors provided by the superinfecting virus [16, 41]. As pointed out by Preston , the refractory state of quiescent HSV genomes appears to be distinct from that adopted by the viral genome during latent infection of sensory neurons, as latent HSV genomes can be reactivated in response to external signals or by expression of any of HSV VP16, ICP4 or ICP0 ; in contrast, the only known means of reactivating quiescent genomes is via expression of ICP0 or its HCMV functional counterpart pp71. The implication is that quiescent genomes are more effectively shielded from trans-acting factors than latent genomes.
The mechanisms that prevent quiescent HSV genomes from responding to trans-acting factors are of great interest, as is the mode of action of ICP0 in overcoming this barrier to gene expression and DNA replication. Sequence-specific repression seems unlikely, for two reasons. First, the results of this and previous [18, 41] reports indicate that genes driven from at least three distinct categories of viral promoters (HCMV IE, HSV IE, and HSV VP16) remain silent in cells superinfected with ICP0-deficient HSV, despite the activity of the corresponding genes located in the superinfecting viral genome. Similarly, the quiescent genome fails to respond to the viral DNA replication and recombination machinery provided by the superinfecting virus. These data suggest that the inhibitory mechanism renders many (if not all) of the cis-acting elements (eg. promoters and origins of DNA replication) located in the quiescent genome non-operative. Second, the quiescent genome is not activated by replication of the superinfecting viral genome within the same nucleus, a condition that would likely titrate classical sequence-specific DNA-binding repressors. These data suggest that quiescent genomes may be stably associated with repressive material that does not readily equilibrate between viral genomes, or located at one or more inaccessible intranuclear sites.
The functional inaccessibility of quiescent HSV genomes documented here is reminiscent of that displayed by genes located in cellular heterochromatin ; however it is worth emphasizing that previous work has shown that quiescent HSV genomes lack regularly spaced nucleosomes at the tk locus , a feature that distinguishes them both from classical heterochromatin and the latent HSV genomes present in sensory neurons . Moreover, HSV infection (and ICP0) does not activate the heterochromatinized endogenous cellular β-globin gene in present fibroblasts, although transfected (and presumably euchromatic) copies of this gene are susceptible to activation by HSV infection [63, 64]. These considerations raise the possibility that HSV genome quiescence involves novel mechanisms, perhaps related to those that inhibit HSV transcription in response to type I IFN [51, 65]. Indeed, ICP0 is able to overcome the IFN-induced barrier to HSV transcription , in addition to triggering reactivation of quiescent HSV genomes. It therefore seems likely that further studies of the mode of action of ICP0 may illuminate one or more intranuclear mechanisms of antiviral defense.
Our results provide strong support for the hypothesis that quiescent HSV genomes are silenced by a cellular mechanism that renders them globally inaccessible to most trans-acting factors. The implication is that ICP0 triggers reactivation from quiescence by overcoming this generalized barrier to gene expression and DNA replication. Further studies designed to identify the components of this repression mechanism will clarify how the balance between host intranuclear repression mechanisms and viral countermeasures regulates the onset of the HSV lytic program of gene expression.
Cells and Virus
Human U2OS osteosarcoma cells, Human Embryonic Lung (HEL) fibroblasts and African green monkey kidney (Vero) cells were obtained from the American Type Culture Collection. E5  and V27  cells were gifts from N. A. DeLuca and S. Rice respectively. Cells were maintained in Dulbecco's Modified Eagle Medium (DMEM) (Gibco) supplemented with 10% (U2OS and HEL) or 5% (Vero) fetal bovine serum (FBS), 50 units/ml penicillin (P) and 5 μg/ml streptomycin (S). E5 and V27 cells were additionally supplemented with 100 μg/ml G418 (Geneticin®, GIBCO).
KOS 1.1 (a wild-type strain of HSV-1), KOS-G (see below) and d22lacZ ( ICP22-) were grown and titered on Vero cells. n212 ( ICP0-), n212-G, KM110 ( VP16/ICP0- double mutant) and KM110-R were grown and titered on U20S cells (in the presence of 3 mM HMBA for KM110). d120 ( ICP4-) and d27-1 ( ICP27-) were grown and titered on complementing E5 and V27 cells respectively.
In experiments where the progeny of superinfected cultures were examined for recovery of the dsRED gene (table 1), the superinfected cells were treated with an acid glycine wash to remove any input superinfecting virus that had not penetrated the host cells, as follows. 2 hrs post-superinfection, the growth medium from monolayers grown in 12 well plates was aspirated. The cells were then incubated with 1 ml Acid Glycine wash (8 g/L NaCl, 1.8 g/L KCl, 0.1 g/L MgCl2·6H2O, 0.1 g/L CaCl2·6H2O, 7.5 g/L glycine, pH 3) for 30 seconds. After two washes with 1 ml Phosphate Buffered Saline (PBS: 10 mg/ml NaCl, 0.25 mg/ml KCl, 1.8 mg/ml Na2HPO4, 0.3 mg/ml KH2PO4, pH 7.5), regular growth medium was added.
Construction of recombinant viruses
We modified KOS1.1, n212, and KM110 by inserting transgenes encoding eGFP (KOS-G, n212-G) or dsRed2 (KM110-R) driven from the human cytomegalovirus immediate-early promoter into the viral thymidine kinase (tk) locus in the tk sense orientation. To this end, 1.6 kbp Ase I-Mlu I fragments bearing the HCMV promoter, DsRed2 or eGFP coding sequence, and SV40 early polyadenylation signal were excised from pDsRed2-C1(Clontech) or pEGFP-C1 (Clontech) and inserted into SstI site in the tk coding sequences carried by pTK173 after making all ends blunt, generating pTK-Red and pTK-Green. The resulting tk-deficient insertion mutations were then transferred into the intact viral genomes of KOS1.1, n212, and KM110 via DNA-mediated marker rescue using standard methods. Briefly, 350 ng of pTK-Red or pTK-Green (cleaved with Afl III) was combined with 1–2 μg of total cellular DNA extracted from cells infected with the target virus, and the resulting mixture was transfected into U2OS cells using Fugene (Roche). Recombinants were then isolated from the progeny of the co-transfection by picking red or green fluorescent plaques. After several rounds of plaque purification the identity and purity of the recombinants was confirmed by Southern blot analysis of the viral tk, VP16, and ICP0 loci.
Samples were subject to electrophoresis through 12% SDS polyacrylamide gels along with 10 μl pre-stained molecular weight standards, Low Range (BIO-RAD), then transferred to a nitrocellulose membrane (Hybond ECL, Ambersham Pharmacia) using a wet protein transfer apparatus (Bio-Rad Trans-blot cell). Following the transfer, the membrane was incubated in 10% skim milk TBS-Tween (25 mM Tris, pH 8, 150 mM NaCl, 0.1% Tween-20) overnight at 4°C. Monoclonal antibodies to VP16 (LP1,  a generous gift from A. Minson) and β-actin (Sigma Aldrich) were used at dilutions of 1:16,000 and 1:5,000 respectively. The membrane was incubated with the primary antibody diluted in TBS-Tween/5% skim milk for 30 min at room temperature then washed three times for 10 min in TBS-Tween. The membrane was then incubated with secondary antibody, goat anti-mouse IgG-HRP (BioRad) diluted 1:3,000 in TBS-Tween/5% skim milk, for 30 min at room temperature. After washing three times as before, the membrane was developed using ECL+plus system (Amersham Biosciences) according to the manufacture's instructions. The signal was detected by exposure to Fuji Super RX X-Ray film.
Total cellular DNA extracted as previously described was cleaved with a mixture of Bam H I and Nhe I, then subjected to electrophoresis through a 1% agarose gel in Tris-acetate EDTA (TAE) for 2 hrs at 80 V in TAE buffer. The gel was then stained with SYBR Gold Nucleic Acid Gel Stain (Molecular Probes) according the manufacturer's instructions and quantified by phospho-imager analysis on a Storm 860 (Molecular Dynamics). The gel was washed sequentially in the following solutions for 15 min each: 0.25 M HCl, 0.5 M NaOH, 1 M Tris/1.5 M NaCl, and 10 × SSC. DNA was transferred to a GeneScreen Plus nylon membrane (NEN Life Sciences Products) in 10 × SSC. The membrane was UV-cross linked using Stratalinker 2400 (Stratagene) before being hybridized to a 32P-labelled 1537 nt probe VP16 probe generated by random priming. The probe fragment was obtained by polymerase chain reaction using pVP16 KOS  as the template and the primers 5' CGCCGTCGGGCGTCCCACAC 3' and 5' CGGGGGATGCGGATCCGGTCGCGC 3'. The 32P signal was detected by exposure to Kodak BioMax MS film at -80°C.
Cells were detached from the growth surface with trypsin, resuspended in DMEM and transferred to a 5 ml Falcon tube. Red and green fluorescence was quantified by passing the cells through a Becton Dickson FACScan and analyzed using CellQuest Software. HEL cells exhibit substantial levels of autofluorescence, potentially interfering with the analysis. However, we found that the intensities of the red and green autofluorerescent signals emitted by individual HEL cells are highly correlated (see for example figure 1) such that cells expressing neither dsRED2 nor eGFP fall on the diagonal of plots of green versus red signal intensity. This correlation allows cells expressing even low levels of dsRED2 to be readily detected as signals above the control diagonal (shown as the red dots figure 1). Note that this procedure uses the green autofluorescent signal emmited by each cell to estimate its autofluorence in the read channel. However, this procedure cannot be used if the cells also express eGFP (see figure 3), because the green autofluorescence is masked by the eGFP fluorescence. Hence, the only a minority of the RFP+ cells can be detected when the cells also express GFP (indicated by the purple dots in figure 3).
Detection of viral DNA replication compartments via indirect immunofluorescence of ICP4
Monolayers of HEL cells grown on 18 mm coverslips (Fisher Scientific) in a 12 well plate were fixed by washing twice with 1 ml PBS and incubating in 400 μl PBS containing 5% formaldehyde and 2% sucrose for 10 min. This and subsequent manipulations were at room temperature. The cells were then permeabilized by washing twice with 1 ml PBS and incubating in 400 μl PBS containing 0.6% NonidetP-40 and 10% sucrose for 10 min. After washing twice more with 1 ml PBS/1% FBS, the cells were incubated with 100 μl primary anti-ICP4 monoclonal antibody (#1114, Goodwin Institute) diluted 1:1000 in PBS/1%FBS for 1 hr, and washed six times with PBS/1% FBS over 15 min. The cells were then incubated in 100 μl Alexa Fluor® 488 labeled goat anti-mouse IgG (Molecular Probes) diluted 1:1000 in PBS/1%FBS for 1 hr and washed six times as before. The cell nuclei were stained by incubating in 100 μl of 500 ng/ml Hoescht 33342 (Molecular Probes) in PBS solution for 10 min, protected from the light. After washing three times in PBS/1% FBS, the coverslips were dipped in H2O, and allowed to dry for 15 min, protected from the light. The coverslips were mounted on slides using 20 μl Vectashield mounting medium, and secured with clear nail polish. Slides were examined using a Zeiss LSM 510, 2 photon Laser Scanning Microscope system with two lasers giving excitation lines at 488 nm (for Alexa Fluor 488) and 780 nm (for Hoescht stain), and using a 40× oil immersion objective lens.
List of abbreviations
infected cell protein
enhanced green fluorescent protein
human embryonic lung fibroblasts
herpes simplex virus
multiplicity of infection
nuclear domain 10
red fluorescent protein
viral protein 16
We thank Holly Saffran and Rob Maranchuk for technical support, and Jennifer Corcoran for comments on the manuscript. This work was supported by an operating grant from the Canadian Institutes for Health Research. JRS holds a Canada Research Chair in Molecular Virology.
- Roizman B, Knipe DM: Herpes simplex viruses and their replication. In Fields Virology. Volume 2. 4th edition. Edited by: Knipe DM, Howley PH. Philadelphia , Lippincott Williams and Wilkins; 2001:2399-2459.Google Scholar
- Cai W, Astor TL, Lipak LM, Cho C, Coen DM, Schaffer PA: The herpes simplex virus type 1 regulatory protein ICP0 enhances virus replication during acute infection and reactivation from latency. J Virol 1993, 67: 7501-7512.PubMed CentralPubMedGoogle Scholar
- Cai W, Schaffer PA: Herpes simplex virus type 1 ICP0 regulates expression of immediate-early, early, and late genes in productively infected cells. J Virol 1992, 66: 2904-2915.PubMed CentralPubMedGoogle Scholar
- Stow ND, Stow EC: Isolation and characterization of a herpes simplex virus type 1 mutant containing a deletion within the gene encoding the immediate early polypeptide Vmw110. J Gen Virol 1986, 67: 2571-2585.View ArticlePubMedGoogle Scholar
- Everett RD: Construction and characterization of herpes simplex virus type 1 mutants with defined lesions in immediate-early gene 1. J Gen Virol 1989, 70: 1185-1202.View ArticlePubMedGoogle Scholar
- Cai W, Schaffer PA: Herpes simplex virus type 1 ICPO plays a critical role in the de novo synthesis of infectious virus following transfection of viral DNA. J Virol 1989, 63: 4579-4589.PubMed CentralPubMedGoogle Scholar
- Everett RD: Transactivation of transcription by herpes virus products: requirement for two HSV-1 immediate-early polypeptides for maximum activity. EMBO J 1984, 3: 3135-3141.PubMed CentralPubMedGoogle Scholar
- Gelman IH, Silverstein S: Identification of immediate early genes from herpes simplex virus that transactivates the virus thymidine kinase gene. Proc Natl Sci USA 1985, 82: 5265-5269.View ArticleGoogle Scholar
- O'Hare P, Hayward GS: Three trans-acting regulatory proteins of herpes simplex virus modulate immediate-early gene expression in a pathway involving positive and negative feedback regulation. J Virol 1985, 56: 723-733.PubMed CentralPubMedGoogle Scholar
- Quinlan MP, Knipe DM: Stimulation of expression of a herpes simplex virus DNA-binding protein by two viral functions. Mol Cell Biol 1985, 5: 957-963.PubMed CentralView ArticlePubMedGoogle Scholar
- Ace CI, McKee TA, Ryan JM, Cameron JM, Preston CM: Construction and characterization of a herpes simplex virus type 1 mutant unable to transinduce immediate-early gene expression. Journal of Virology 1989, 63: 2260-2269.PubMed CentralPubMedGoogle Scholar
- Halford WP, Kemp CD, Isler JA, Davido DJ, Schaffer PA: ICP0, ICP4, or VP16 expressed from adenovirus vectors induces reactivation of latent herpes simplex virus type 1 in primary cultures of latently infected trigeminal ganglion cells. J Virol 2001, 75: 6143-6153. 10.1128/JVI.75.13.6143-6153.2001PubMed CentralView ArticlePubMedGoogle Scholar
- Everett RD: ICP0, a regulator of herpes simplex virus during lytic and latent infection. Bioessays 2000, 22: 761-770. 10.1002/1521-1878(200008)22:8<761::AID-BIES10>3.0.CO;2-AView ArticlePubMedGoogle Scholar
- Jordan R, Schaffer P: Activation of gene expression by herpes simplex virus type 1 ICP0 occurs at the level of mRNA synthesis. J Virol 1997, 71: 6850-6862.PubMed CentralPubMedGoogle Scholar
- Samaniego LA, Wu N, DeLuca NA: The herpes simplex virus immediate-early protein ICP0 affects transcription from the viral genome and infected-cell survival in the absence of ICP4 and ICP27. J Virol 1997, 71: 4614-4625.PubMed CentralPubMedGoogle Scholar
- Preston CM: Repression of viral transcription during herpes simplex virus latency. J Gen Virol 2000, 81: 1-19.View ArticlePubMedGoogle Scholar
- Stow EC, Stow ND: Complementation of a herpes simplex virus type 1 Vmw110 deletion mutant by human cytomegalovirus. J Gen Virol 1989, 70: 695-704.View ArticlePubMedGoogle Scholar
- Preston CM, Nicholl MJ: Repression of gene expression upon infection of cells with herpes simplex virus type 1 mutants impaired for immediate-early protein synthesis. J Virol 1997, 71: 7807-7813.PubMed CentralPubMedGoogle Scholar
- Samaniego LA, Neiderhiser L, DeLuca NA: Persistence and expression of the herpes simplex virus genome in the absence of immediate-early proteins. J Virol 1998, 72: 3307-3320.PubMed CentralPubMedGoogle Scholar
- Jamieson DR, Robinson LH, Daksis JI, M.J. N, Preston CM: Quiescent viral genomes in human fibroblasts after infection with herpes simplex virus type 1 Vmw65 mutants. J Gen Virol 1995, 76: 1417-1431.View ArticlePubMedGoogle Scholar
- Jackson SA, DeLuca NA: Relationship of herpes simplex virus genome configuration to productive and persistent infections. Proc Natl Acad Sci U S A 2003,100(13):7871-7876. 10.1073/pnas.1230643100PubMed CentralView ArticlePubMedGoogle Scholar
- Hobbs WE, Brough DE, Kovesdi I, DeLuca NA: Efficient activation of viral genomes by levels of herpes simplex virus ICP0 insufficient to affect cellular gene expression or cell survival. J Virol 2001, 75: 3391-3403. 10.1128/JVI.75.7.3391-3403.2001PubMed CentralView ArticlePubMedGoogle Scholar
- Preston CM, Nicholl MJ: Human cytomegalovirus tegument protein pp71 directs long-term gene expression from quiescent herpes simplex virus genomes. J Virol 2005, 79: 525-535. 10.1128/JVI.79.1.525-535.2005PubMed CentralView ArticlePubMedGoogle Scholar
- Everett RD, Orr A, Preston CM: A viral activator of gene expression functions via the ubiquitin- proteasome pathway. Embo J 1998,17(24):7161-7169. 10.1093/emboj/17.24.7161PubMed CentralView ArticlePubMedGoogle Scholar
- Hagglund R, Roizman B: Role of ICP0 in the strategy of conquest of the host cell by herpes simplex virus 1. J Virol 2004, 78: 2169-2178. 10.1128/JVI.78.5.2169-2178.2004PubMed CentralView ArticlePubMedGoogle Scholar
- Lomonte P, Thomas J, Texier P, Caron C, Khochbin S, Epstein AL: Functional interaction between class II histone deacetylases and ICP0 of herpes simplex virus type 1. J Virol 2004, 78: 6744-6757. 10.1128/JVI.78.13.6744-6757.2004PubMed CentralView ArticlePubMedGoogle Scholar
- Gu H, Liang Y, Mandel G, Roizman B: Components of the REST/CoREST/histone deacetylase repressor complex are disrupted, modified, and translocated in HSV-1-infected cells. Proc Natl Acad Sci U S A 2005,102(21):7571-7576. 10.1073/pnas.0502658102PubMed CentralView ArticlePubMedGoogle Scholar
- Boutell C, Sadis S, Everett RD: Herpes simplex virus type 1 immediate-early protein ICP0 and its isolated RING finger domain act as ubiquitin E3 ligases in vitro. J Virol 2002,76(2):841-850. 10.1128/JVI.76.2.841-850.2002PubMed CentralView ArticlePubMedGoogle Scholar
- Everett RD, O'Hare P, O'Rouke D, Barlow P, Orr A: Point mutations in the herpes simplex virus type 1 Vmw110 ring finger helix affect activation of gene expression, viral growth, and interaction with PML-containing structures. J Virol 1995, 69: 7339-7344.PubMed CentralPubMedGoogle Scholar
- Hagglund R, Van Sant C, Lopez P, Roizman B: Herpes simplex virus 1-infected cell protein 0 contains two E3 ubiquitin ligase sites specific for different E2 ubiquitin-conjugating enzymes. Proc Natl Acad Sci U S A 2002, 99: 631-636. 10.1073/pnas.022531599PubMed CentralView ArticlePubMedGoogle Scholar
- Ishov AM, Maul GG: The periphery of nuclear domain 10 (ND10) as site of DNA virus deposition. J Cell Biol 1996,134(4):815-826. 10.1083/jcb.134.4.815View ArticlePubMedGoogle Scholar
- Maul GG, Ishov AM, Everett RD: Nuclear domain 10 as preexisting potential replication start sites of herpes simplex virus type-1. Virology 1996, 217: 67-75. 10.1006/viro.1996.0094View ArticlePubMedGoogle Scholar
- Everett RD, Sourvinos G, Leiper C, Clements JB, Orr A: Formation of nuclear foci of the herpes simplex virus type 1 regulatory protein ICP4 at early times of infection: localization, dynamics, recruitment of ICP27, and evidence for the de novo induction of ND10-like complexes. J Virol 2004,78(4):1903-1917. 10.1128/JVI.78.4.1903-1917.2004PubMed CentralView ArticlePubMedGoogle Scholar
- Everett RD, Maul GG: HSV-1 IE protein Vmw110 causes redistribution of PML. EMBO J 1994, 13: 5062-5069.PubMed CentralPubMedGoogle Scholar
- Maul GG, Everett RD: The nuclear location of PML, a cellular member of the C3HC4 zinc-binding domain protein family, is rearranged during herpes simplex virus infection by the C3HC4 viral protein ICP0. J Gen Virol 1994, 75: 1223-1233.View ArticlePubMedGoogle Scholar
- Maul GG, Guldner HH, Spivack JG: Modification of discrete nuclear domains induced by herpes simplex virus type 1 immediate early gene 1 product (ICP0). J Gen Virol 1993, 74: 2679-2690.View ArticlePubMedGoogle Scholar
- Everett RD, Freemont P, Saitoh H, Dasso M, Orr A, Kathoria M, Parkinson J: The disruption of ND10 during herpes simplex virus infection correlates with the Vmw110- and proteasome-dependent loss of several PML isoforms. J Virol 1998,72(8):6581-6591.PubMed CentralPubMedGoogle Scholar
- Chelbi-Alix MK, de The H: Herpes virus induced proteasome-dependent degradation of the nuclear bodies-associated PML and Sp100 proteins. Oncogene 1999,18(4):935-941. 10.1038/sj.onc.1202366View ArticlePubMedGoogle Scholar
- Parkinson J, Everett RD: Alphaherpesvirus proteins related to herpes simplex virus type 1 ICP0 affect cellular structures and proteins. J Virol 2000,74(21):10006-10017. 10.1128/JVI.74.21.10006-10017.2000PubMed CentralView ArticlePubMedGoogle Scholar
- Russel J, Stow ND, Stow EC, Preston CM: Herpes simplex virus genes involved in latency in vitro. J Gen Virol 1987, 68: 3009-3018.View ArticleGoogle Scholar
- Harris RA, Everett RD, Zhu XX, Silverstein S, Preston CM: Herpes simplex virus type 1 immediate-early protein Vmw110 reactivates latent herpes simplex virus type 2 in an in vitro latency system. Journal of Virology 1989,63(8):3513-3515.PubMed CentralPubMedGoogle Scholar
- Harris RA, Preston CM: Establishment of latency in vitro by the herpes simplex virus type 1 mutant in1814. J Gen Virol 1991, 72: 907-913.View ArticlePubMedGoogle Scholar
- Everett RD, Boutell C, Orr A: Phenotype of a Herpes Simplex Virus Type 1 Mutant That Fails To Express Immediate-Early Regulatory Protein ICP0. J Virol 2004,78(4):1763-1774. 10.1128/JVI.78.4.1763-1774.2004PubMed CentralView ArticlePubMedGoogle Scholar
- Cai W, Schaffer PA: A cellular function can enhance gene expression and plating efficiency of a mutant defective in the gene for ICPO, a transactivating protein of herpes simplex virus type 1. J Virol 1991, 65: 4078-4090.PubMed CentralPubMedGoogle Scholar
- Nicholl MJ, Robinson LH, Preston CM: Activation of cellular interferon-responsive genes after infection of human cells with herpes simplex virus type 1. J Gen Virol 2000,81(Pt 9):2215-2218.View ArticlePubMedGoogle Scholar
- Preston CM, Harman AN, Nicholl MJ: Activation of interferon response factor-3 in human cells infected with herpes simplex virus type 1 or human cytomegalovirus. Journal of Virology 2001,75(19): 8909-8916. 10.1128/JVI.75.19.8909-8916.2001PubMed CentralView ArticlePubMedGoogle Scholar
- Mossman KL, Macgregor PF, Rozmus JJ, Goryachev AB, Edwards AM, Smiley JR: Herpes simplex virus triggers and then disarms a host antiviral response. J Virol 2001,75(2):750-758. 10.1128/JVI.75.2.750-758.2001PubMed CentralView ArticlePubMedGoogle Scholar
- Eidson KM, Hobbs WE, Manning BJ, Carlson P, DeLuca NA: Expression of herpes simplex virus ICP0 inhibits the induction of interferon-stimulated genes by viral infection. J Virol 2002,76(5):2180-2191. 10.1128/jvi.76.5.2180-2191.2002PubMed CentralView ArticlePubMedGoogle Scholar
- Lin R, Noyce RS, Collins SE, Everett RD, Mossman KL: The herpes simplex virus ICP0 RING finger domain inhibits IRF3- and IRF7-mediated activation of interferon-stimulated genes. Journal of Virology 2004,78(4):1675-1684. 10.1128/JVI.78.4.1675-1684.2004PubMed CentralView ArticlePubMedGoogle Scholar
- Mossman KL, Saffran HA, Smiley JR: Herpes simplex virus ICP0 mutants are hypersensitive to interferon. J Virol 2000,74(4):2052-2056. 10.1128/JVI.74.4.2052-2056.2000PubMed CentralView ArticlePubMedGoogle Scholar
- Mossman KL, Smiley JR: Herpes simplex virus ICP0 and ICP34.5 counteract distinct interferon-induced barriers to virus replication. J Virol 2002,76(4):1995-1998. 10.1128/JVI.76.4.1995-1998.2002PubMed CentralView ArticlePubMedGoogle Scholar
- Harle P, Sainz BJ, Carr DJ, Halford WP: The immediate-early protein, ICP0, is essential for the resistance of herpes simplex virus to interferon-alpha/beta. Virology 2002,293(2):295-304. 10.1006/viro.2001.1280View ArticlePubMedGoogle Scholar
- Mossman KL, Smiley JR: Truncation of the C-terminal acidic transcriptional activation domain of herpes simplex virus VP16 renders expression of the immediate-early genes almost entirely dependent on ICP0. J Virol 1999,73(12):9726-9733.PubMed CentralPubMedGoogle Scholar
- Meurens F, Schynts F, Keil GM, Muylkens B, Vanderplasschen A, Gallego P, Thiry E: Superinfection prevents recombination of the alphaherpesvirus bovine herpesvirus 1. J Virol 2004,78(8):3872-3879. 10.1128/JVI.78.8.3872-3879.2004PubMed CentralView ArticlePubMedGoogle Scholar
- Johnson RM, Spear PG: Herpes simplex virus glycoprotein D mediates interference with herpes simplex virus infection. J Virol 1989,63(2):819-827.PubMed CentralPubMedGoogle Scholar
- Banfield BW, Kaufman JD, Randall JA, Pickard GE: Development of pseudorabies virus strains expressing red fluorescent proteins: new tools for multisynaptic labeling applications. J Virol 2003,77(18):10106-10112. 10.1128/JVI.77.18.10106-10112.2003PubMed CentralView ArticlePubMedGoogle Scholar
- Preston CM: Control of herpes simplex virus type 1 mRNA synthesis in cells infected with wild-type virus or the temperature-sensitive mutant tsK. J Virol 1979, 29: 275-284.PubMed CentralPubMedGoogle Scholar
- DeLuca NA, McCarthy AM, Schaffer PA: Isolation and characterization of deletion mutants of herpes simplex virus type 1 in the gene encoding immediate-early regulatory protein ICP4. J Virol 1985,56(2):558-570.PubMed CentralPubMedGoogle Scholar
- Smiley JR, Duncan J: Truncation of the C-terminal acidic transcriptional activation domain of herpes simplex virus VP16 produces a phenotype similar to that of the in1814 linker insertion mutation. J Virol 1997,71(8):6191-6193.PubMed CentralPubMedGoogle Scholar
- Knipe DM, Senechek D, Rice SA, Smith JL: Stages in the nuclear association of the herpes simplex virus transcriptional activator protein ICP4. J Virol 1987,61(2):276-284.PubMed CentralPubMedGoogle Scholar
- Craig JM: Heterochromatin--many flavours, common themes. Bioessays 2005,27(1):17-28. 10.1002/bies.20145View ArticlePubMedGoogle Scholar
- Deshmane SL, Fraser NW: During latency, herpes simplex virus type 1 DNA is associated with nucleosomes in a chromatin structure. Journal of Virology 1989, 63: 943-947.PubMed CentralPubMedGoogle Scholar
- Everett RD: Activation of cellular promoters during herpes virus infection of biochemically transformed cells. Embo J 1985,4(8):1973-1980.PubMed CentralPubMedGoogle Scholar
- Cheung P, Panning B, Smiley JR: Herpes simplex virus immediate-early proteins ICP0 and ICP4 activate the endogenous human alpha-globin gene in nonerythroid cells. J Virol 1997,71(3):1784-1793.PubMed CentralPubMedGoogle Scholar
- Nicholl MJ, Preston CM: Inhibition of herpes simplex virus type 1 immediate-early gene expression by alpha interferon is not VP16 specific. J Virol 1996,70(9):6336-6339.PubMed CentralPubMedGoogle Scholar
- DeLuca NA, Schaffer PA: Activities of herpes simplex virus type 1 (HSV-1) ICP4 genes specifying nonsense peptides. Nucleic Acids Res 1987, 15: 4491-4511.PubMed CentralView ArticlePubMedGoogle Scholar
- Rice SA, Knipe DM: Genetic evidence for two distinct trans-activation functions of the herpes simplex virus a protein ICP27. J Virol 1990, 64: 1704-1715.PubMed CentralPubMedGoogle Scholar
- Long MC, Leong V, Schaffer PA, Spencer CA, Rice SA: ICP22 and the UL13 protein kinase are both required for herpes simplex virus-induced modification of the large subunit of RNA polymerase II. J Virol 1999,73(7):5593-5604.PubMed CentralPubMedGoogle Scholar
- McLean C, Buckmaster A, Hancock D, Buchan A, Fuller A, Minson AC: Monoclonal antibodies to three nonglycosylated antigens of herpes simplex virus type 2. J Gen Virol 1982, 63: 297-305.View ArticlePubMedGoogle Scholar
- Weinheimer SP, Boyd BA, Durham SK, Resnick JL, O'Boyle DR: Deletion of the VP16 open reading frame of herpes simplex virus type 1. J Virol 1992, 66: 258-269.PubMed CentralPubMedGoogle Scholar
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