scholarly journals Characterization of a UL49-Null Mutant: VP22 of Herpes Simplex Virus Type 1 Facilitates Viral Spread in Cultured Cells and the Mouse Cornea

2006 ◽  
Vol 80 (17) ◽  
pp. 8664-8675 ◽  
Author(s):  
Carol Duffy ◽  
Jennifer H. LaVail ◽  
Andrew N. Tauscher ◽  
Elizabeth G. Wills ◽  
John A. Blaho ◽  
...  
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Virus Type ◽  
Herpes Simplex Virus Type ◽  
Cultured Cells ◽  
Wild Type ◽  
Viral Spread ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACT Herpes simplex virus type 1 (HSV-1) virions, like those of all herpesviruses, contain a proteinaceous layer termed the tegument that lies between the nucleocapsid and viral envelope. The HSV-1 tegument is composed of at least 20 different viral proteins of various stoichiometries. VP22, the product of the UL49 gene, is one of the most abundant tegument proteins and is conserved among the alphaherpesviruses. Although a number of interesting biological properties have been attributed to VP22, its role in HSV-1 infection is not well understood. In the present study we have generated both a UL49-null virus and its genetic repair and characterized their growth in both cultured cells and the mouse cornea. While single-step growth analyses indicated that VP22 is dispensable for virus replication at high multiplicities of infection (MOIs), analyses of plaque morphology and intra- and extracellular multistep growth identified a role for VP22 in viral spread during HSV-1 infection at low MOIs. Specifically, VP22 was not required for either virion infectivity or cell-cell spread but was required for accumulation of extracellular virus to wild-type levels. We found that the absence of VP22 also affected virion composition. Intracellular virions generated by the UL49-null virus contained reduced amounts of ICP0 and glycoproteins E and D compared to those generated by the wild-type and UL49-repaired viruses. In addition, viral spread in the mouse cornea was significantly reduced upon infection with the UL49-null virus compared to infection with the wild-type and UL49-repaired viruses, identifying a role for VP22 in viral spread in vivo as well as in vitro.

scholarly journals Suppression of Proinflammatory Cytokine Expression by Herpes Simplex Virus Type 1

2004 ◽  
Vol 78 (11) ◽  
pp. 5883-5890 ◽  
Author(s):  
Trine H. Mogensen ◽  
Jesper Melchjorsen ◽  
Lene Malmgaard ◽  
Antonella Casola ◽  
Søren R. Paludan
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Virus Type ◽  
Proinflammatory Cytokines ◽  
Herpes Simplex Virus Type ◽  
Cytokine Expression ◽  
Wild Type ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACT Viral immune evasion strategies are important for establishment and maintenance of infections. Many viruses are in possession of mechanisms to counteract the antiviral response raised by the infected host. Here we show that a herpes simplex virus type 1 (HSV-1) mutant lacking functional viral protein 16 (VP16)—a tegument protein promoting viral gene expression—induced significantly higher levels of proinflammatory cytokines than wild-type HSV-1. This was observed in several cell lines and primary murine macrophages, as well as in peritoneal cells harvested from mice infected in vivo. The enhanced ability to stimulate cytokine expression in the absence of VP16 was not mediated directly by VP16 but was dependent on the viral immediate-early genes for infected cell protein 4 (ICP4) and ICP27, which are expressed in a VP16-dependent manner during primary HSV infection. The virus appeared to target cellular factors other than interferon-induced double-stranded RNA-activated protein kinase R (PKR), since the virus mutants remained stronger inducers of cytokines in cells stably expressing a dominant-negative mutant form of PKR. Finally, mRNA stability assay revealed a significantly longer half-life for interleukin-6 mRNA after infection with the VP16 mutant than after infection with the wild-type virus. Thus, HSV is able to suppress expression of proinflammatory cytokines by decreasing the stability of mRNAs, thereby potentially impeding the antiviral host response to infection.

scholarly journals A Herpes Simplex Virus Type 1 Latency-Associated Transcript Mutant with Increased Virulence and Reduced Spontaneous Reactivation

1999 ◽  
Vol 73 (2) ◽  
pp. 920-929 ◽  
Author(s):  
Guey-Chuen Perng ◽  
Susan M. Slanina ◽  
Ada Yukht ◽  
Barbara S. Drolet ◽  
William Keleher ◽  
...  
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Virus Type ◽  
Herpes Simplex Virus Type ◽  
Ocular Infection ◽  
Wild Type ◽  
Simplex Virus ◽  
Spontaneous Reactivation ◽  
Hsv 1

ABSTRACT The herpes simplex virus type 1 (HSV-1) latency-associated transcript (LAT) gene is essential for efficient spontaneous reactivation of HSV-1 from latency. We previously reported that insertion of the LAT promoter and just the first 1.5 kb of the 8.3-kb LAT gene into an ectopic location in the virus restored wild-type spontaneous reactivation to a LAT null mutant. This mutant, LAT3.3A (previously designated LAT1.5a), thus showed that the expression of just the first 1.5 kb of LAT is sufficient for wild-type spontaneous reactivation. We also showed that in the context of the entire LAT gene, deletion of LAT nucleotides 76 to 447 (LAT mutantdLAT371) had no effect on spontaneous reactivation or virulence. We report here on a LAT mutant designated LAT2.9A. This mutant is similar to LAT3.3A, except that the ectopic LAT insert contains the same 371-nucleotide deletion found in dLAT371. We found that LAT2.9A had a significantly reduced rate of spontaneous reactivation compared to marker-rescued and wild-type viruses. This was unexpected, since the combined results of dLAT371 and LAT3.3A predicted that spontaneous reactivation of LAT2.9A would be wild type. We also found that LAT2.9A was more virulent than wild-type or marker-rescued viruses after ocular infection of rabbits. This was unexpected, since LAT null mutants and LAT3.3A have wild-type virulence. These results suggest for the first time (i) that regions past the first 1.5 kb of LAT can compensate for deletions in the first 1.5kb of LAT and may therefore play a role in LAT dependent spontaneous reactivation and (ii) that regions of LAT affect viral virulence.

scholarly journals Negative regulation of herpes simplex virus type 1 ICP4 promoter by IE180 protein of pseudorabies virus

2004 ◽  
Vol 85 (8) ◽  
pp. 2125-2130 ◽  
Author(s):  
S. Gómez-Sebastián ◽  
E. Tabarés
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Virus Type ◽  
Herpes Simplex Virus Type ◽  
Negative Regulation ◽  
Wild Type ◽  
Recombinant Viruses ◽  
Simplex Virus ◽  
Hsv 1

Recombinant pseudorabies viruses (PRVs) gIS8 and N1aHTK were constructed by the insertion of a chimeric gene (α4–TK) from herpes simplex virus type 1 (HSV-1) into wild-type PRV. HSV-1 TK expression by these recombinant viruses resulted in enhanced sensitivity to ganciclovir, compared to that of the wild-type PRV, and was similar to the sensitivity shown by HSV-1. Infection with gIS8 or N1aHTK recombinant viruses led to expression of HSV-1 TK mRNA as an immediate–early (IE) gene, observed by downregulation of the HSV-1 α4 promoter. This negative regulation was due to a PRV IE protein, IE180. IE180, however, does not have all the regulatory functions of the infected-cell protein ICP4, as it does not restore the growth of ICP4-deficient HSV-1 mutants.

scholarly journals The Dominant-Negative Herpes Simplex Virus Type 1 (HSV-1) Recombinant CJ83193 Can Serve as an Effective Vaccine against Wild-Type HSV-1 Infection in Mice

2004 ◽  
Vol 78 (11) ◽  
pp. 5756-5765 ◽  
Author(s):  
Hanka Augustinova ◽  
Daniela Hoeller ◽  
Feng Yao
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Virus Type ◽  
Herpes Simplex Virus Type ◽  
Dominant Negative ◽  
Trigeminal Ganglia ◽  
Wild Type ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACT By selectively regulating the expression of the trans-dominant-negative mutant polypeptide UL9-C535C, of herpes simplex virus type 1 (HSV-1) origin binding protein UL9 with the tetracycline repressor (tetR)-mediated gene switch, we recently generated a novel replication-defective and anti-HSV-specific HSV-1 recombinant, CJ83193. The UL9-C535C peptides expressed by CJ83193 can function as a potent intracellular therapy against its own replication, as well as the replication of wild-type HSV-1 and HSV-2 in coinfected cells. In this report, we demonstrate that CJ83193 cannot initiate acute productive infection in corneas of infected mice nor can it reactivate from trigeminal ganglia of mice latently infected by CJ83193 in a mouse ocular model. Given that CJ83193 is capable of expressing the viral α, β, and γ1 genes but little or no γ2 genes, we tested the vaccine potential of CJ83193 against HSV-1 infection in a mouse ocular model. Our studies showed that immunization with CJ83193 significantly reduced the yields of challenge HSV in the eyes and trigeminal ganglia on days 3, 5, and 7 postchallenge. Like in mice immunized with the wild-type HSV-1 strain KOS, immunization of mice with CJ83193 prevents the development of keratitis and encephalitis induced by corneal challenge with wild-type HSV-1 strain mP. Delayed-type hypersensitivity (DTH) assays demonstrate that CJ83193 can elicit durable cell-mediated immunity at the same level as that of wild-type HSV-1 and is more effective than that induced by d27, an HSV-1 ICP27 deletion mutant. Moreover, mice immunized with CJ83193 developed strong, durable HSV-1-neutralizing antibodies at levels at least twofold higher than those induced by d27. The results presented in this report have shed new light on the development of effective HSV viral vaccines that encode a unique safety mechanism capable of inhibiting the mutant's own replication and that of wild-type virus.

scholarly journals Differing Roles of Inner Tegument Proteins pUL36 and pUL37 during Entry of Herpes Simplex Virus Type 1

2008 ◽  
Vol 83 (1) ◽  
pp. 105-116 ◽  
Author(s):  
Ashley P. E. Roberts ◽  
Fernando Abaitua ◽  
Peter O'Hare ◽  
David McNab ◽  
Frazer J. Rixon ◽  
...  
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Virus Type ◽  
Herpes Simplex Virus Type ◽  
Wild Type ◽  
Tegument Proteins ◽  
Infected Cells ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACT Studies with herpes simplex virus type 1 (HSV-1) have shown that secondary envelopment and virus release are blocked in mutants deleted for the tegument protein gene UL36 or UL37, leading to the accumulation of DNA-containing capsids in the cytoplasm of infected cells. The failure to assemble infectious virions has meant that the roles of these genes in the initial stages of infection could not be investigated. To circumvent this, cells infected at a low multiplicity were fused to form syncytia, thereby allowing capsids released from infected nuclei access to uninfected nuclei without having to cross a plasma membrane. Visualization of virus DNA replication showed that a UL37-minus mutant was capable of transmitting infection to all the nuclei within a syncytium as efficiently as the wild-type HSV-1 strain 17+ did, whereas infection by UL36-minus mutants failed to spread. Thus, these inner tegument proteins have differing functions, with pUL36 being essential during both the assembly and uptake stages of infection, while pUL37 is needed for the formation of virions but is not required during the initial stages of infection. Analysis of noninfectious enveloped particles (L-particles) further showed that pUL36 and pUL37 are dependent on each other for incorporation into tegument.

scholarly journals Re-Evaluating Natural Resistance to Herpes Simplex Virus Type 1

2004 ◽  
Vol 78 (18) ◽  
pp. 10086-10095 ◽  
Author(s):  
William P. Halford ◽  
John W. Balliet ◽  
Bryan M. Gebhardt
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Virus Type ◽  
Herpes Simplex Virus Type ◽  
Infection Process ◽  
Natural Resistance ◽  
Viral Spread ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACT It is often stated that individuals of a species can differ significantly in their innate resistance to infection with herpes simplex virus type 1 (HSV-1). Three decades ago Lopez reported that C57BL/6 mice could survive a 5,000-fold-higher inoculum of HSV-1 given intraperitoneally than mice of the A or BALB/c strain (Nature 258:152-153, 1975). Susceptible strains of mice died of encephalitis-like symptoms, suggesting that viral spread to the central nervous system was the cause of death. Although Lopez's study documented that C57BL/6 mice were resistant to the development of HSV-1 encephalitis and mortality, the resistance of C57BL/6 mice to other steps of the HSV-1 infection process was not assessed. The results of the present study extend these observations to clarify the difference between resistance to (i) HSV-1 pathogenesis, (ii) HSV-1 replication, (iii) HSV-1 spread, and (iv) the establishment of latent HSV-1 infection. Although C57BL/6 mice are more resistant to HSV-1 pathogenesis than BALB/c mice, the results of the present study establish that HSV-1 enters, replicates, spreads, and establishes latent infections with virtually identical efficiencies in C57BL/6 and BALB/c mice. These observations raise questions about the validity of the inference that differences in natural resistance are relevant in explaining what differentiates humans with recurrent herpetic disease from the vast majority of asymptomatic carriers of HSV-1 and HSV-2.

scholarly journals The Herpes Simplex Virus Type 1 Regulatory Protein ICP27 Is Required for the Prevention of Apoptosis in Infected Human Cells

1999 ◽  
Vol 73 (4) ◽  
pp. 2803-2813 ◽  
Author(s):  
Martine Aubert ◽  
John A. Blaho
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Virus Type ◽  
Herpes Simplex Virus Type ◽  
Vero Cells ◽  
Human Cells ◽  
Wild Type ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACT The herpes simplex virus type 1 (HSV-1) ICP27 protein is an immediate-early or α protein which is essential for the optimal expression of late genes as well as the synthesis of viral DNA in cultures of Vero cells. Our specific goal was to characterize the replication of a virus incapable of synthesizing ICP27 in cultured human cells. We found that infection with an HSV-1 ICP27 deletion virus of at least three separate strains of human cells did not produce immediate-early or late proteins at the levels observed following wild-type virus infections. Cell morphology, chromatin condensation, and genomic DNA fragmentation measurements demonstrated that the human cells died by apoptosis after infection with the ICP27 deletion virus. These features of the apoptosis were identical to those which occur during wild-type infections of human cells when total protein synthesis has been inhibited. Vero cells infected with the ICP27 deletion virus did not exhibit any of the features of apoptosis. Based on these results, we conclude that while HSV-1 infection likely induced apoptosis in all cells, viral evasion of the response differed among the cells tested in this study.

scholarly journals Anterograde Spread of Herpes Simplex Virus Type 1 Requires Glycoprotein E and Glycoprotein I but Not Us9

2009 ◽  
Vol 83 (17) ◽  
pp. 8315-8326 ◽  
Author(s):  
Helen M. McGraw ◽  
Sita Awasthi ◽  
Jason A. Wojcechowskyj ◽  
Harvey M. Friedman
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Virus Type ◽  
Herpes Simplex Virus Type ◽  
Neuron Culture ◽  
Viral Spread ◽  
Neuron Cell Body ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACT Anterograde neuronal spread (i.e., spread from the neuron cell body toward the axon terminus) is a critical component of the alphaherpesvirus life cycle. Three viral proteins, gE, gI, and Us9, have been implicated in alphaherpesvirus anterograde spread in several animal models and neuron culture systems. We sought to better define the roles of gE, gI, and Us9 in herpes simplex virus type 1 (HSV-1) anterograde spread using a compartmentalized primary neuron culture system. We found that no anterograde spread occurred in the absence of gE or gI, indicating that these proteins are essential for HSV-1 anterograde spread. However, we did detect anterograde spread in the absence of Us9 using two independent Us9-deleted viruses. We confirmed the Us9 finding in different murine models of neuronal spread. We examined viral transport into the optic nerve and spread to the brain after retinal infection; the production of zosteriform disease after flank inoculation; and viral spread to the spinal cord after flank inoculation. In all models, anterograde spread occurred in the absence of Us9, although in some cases at reduced levels. This finding contrasts with gE- and gI-deleted viruses, which displayed no anterograde spread in any animal model. Thus, gE and gI are essential for HSV-1 anterograde spread, while Us9 is dispensable.

scholarly journals Effects of Targeting Herpes Simplex Virus Type 1 gD to the Endoplasmic Reticulum and trans-Golgi Network

1999 ◽  
Vol 73 (11) ◽  
pp. 9515-9520 ◽  
Author(s):  
Alison Whiteley ◽  
Birgitte Bruun ◽  
Tony Minson ◽  
Helena Browne
Keyword(s):  
Endoplasmic Reticulum ◽  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Virus Type ◽  
Herpes Simplex Virus Type ◽  
Wild Type ◽  
Simplex Virus ◽  
Golgi Network ◽  
Hsv 1

ABSTRACT Glycoprotein D (gD) of herpes simplex virus type 1 (HSV-1) was modified to encode targeting signals known to localize proteins to either the endoplasmic reticulum (ER) or the trans-Golgi network. These motifs conferred the predicted targeting properties on gD in transfected cells as judged by immunofluorescence staining, and the exclusion of targeted gD from the cell surface was confirmed by the fact that these molecules exhibited substantially reduced activity in cell-cell fusion assays. Recombinant viruses expressing Golgi-targeted forms of gD grew to wild-type levels in noncomplementing cells, exhibited unaltered particle/infectivity ratios, and were found to contain wild-type levels of gD, whereas a recombinant expressing ER-retained gD was helper cell dependent and, when grown on noncomplementing cells, produced virions of low specific infectivity with greatly reduced levels of gD. These data imply that HSV-1 acquires its final membrane from a post-ER compartment and lend support to the view that the virus undergoes de-envelopment and reenvelopment steps during virus egress.

scholarly journals Emerin Is Hyperphosphorylated and Redistributed in Herpes Simplex Virus Type 1-Infected Cells in a Manner Dependent on both UL34 and US3

2007 ◽  
Vol 81 (19) ◽  
pp. 10792-10803 ◽  
Author(s):  
Natalie Leach ◽  
Susan L. Bjerke ◽  
Desire K. Christensen ◽  
Jacques M. Bouchard ◽  
Fan Mou ◽  
...  
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Virus Type ◽  
Herpes Simplex Virus Type ◽  
Nuclear Lamina ◽  
Wild Type ◽  
Simplex Virus ◽  
In Cells ◽  
Hsv 1

ABSTRACT Cells infected with wild-type herpes simplex virus type 1 (HSV-1) show disruption of the organization of the nuclear lamina that underlies the nuclear envelope. This disruption is reflected in changes in the localization and phosphorylation of lamin proteins. Here, we show that HSV-1 infection causes relocalization of the LEM domain protein emerin. In cells infected with wild-type virus, emerin becomes more mobile in the nuclear membrane, and in cells infected with viruses that fail to express UL34 protein (pUL34) and US3 protein (pUS3), emerin no longer colocalizes with lamins, suggesting that infection causes a loss of connection between emerin and the lamina. Infection causes hyperphosphorylation of emerin in a manner dependent upon both pUL34 and pUS3. Some emerin hyperphosphorylation can be inhibited by the protein kinase Cδ (PKCδ) inhibitor rottlerin. Emerin and pUL34 interact physically, as shown by pull-down and coimmunoprecipitation assays. Emerin expression is not, however, necessary for infection, since virus growth is not impaired in cells derived from emerin-null transgenic mice. The results suggest a model in which pUS3 and PKCδ that has been recruited by pUL34 hyperphosphorylate emerin, leading to disruption of its connections with lamin proteins and contributing to the disruption of the nuclear lamina. Changes in emerin localization, nuclear shape, and lamin organization characteristic of cells infected with wild-type HSV-1 also occur in cells infected with recombinant virus that does not make viral capsids, suggesting that these changes occur independently of capsid envelopment.

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