Herpes Simplex Virus 1 Counteracts Viperin via Its Virion Host Shutoff Protein UL41

The interferon (IFN)-inducible viperin protein restricts a broad range of viruses. However, whether viperin plays a role during herpes simplex virus 1 (HSV-1) infection is poorly understood. In the present study, it was shown for the first time that wild-type (WT) HSV-1 infection couldn''t induce viperin production, and ectopically expressed viperin inhibited the replication of UL41-null HSV-1 but not WT viruses. The underlying molecular mechanism is that UL41 counteracts viperin''s antiviral activity by reducing its mRNA accumulation.     

The Virion Host Shutoff RNase Plays a Key Role in Blocking the Activation of Protein Kinase R in Cells Infected with Herpes Simplex Virus 1

Earlier studies have shown that active MEK blocks the activation of protein kinase R (PKR), a component of antiviral innate immune responses. In this report we show that the herpes simplex virus 1 virion host shutoff (VHS) RNase protein and MEK (mitogen-activated protein kinase kinase) act cooperatively in blocking the activation of PKR. This conclusion is based on the following. (i) In contrast to viral gene expression in the parental cell line or a cell line expressing a constitutively active MEK, the replication of a VHS mutant is particularly impaired in cells expressing dominant negative MEK. In this cell line PKR is activated by phosphorylation, and the accumulation of several viral proteins is delayed. (ii) In transfected cells, wild-type VHS blocked the activation of PKR, whereas PKR was activated in cells transfected with a mutant VHS or with plasmids encoding the VHS RNase and VP16 and VP22, the two viral proteins that neutralize the RNase activity of VHS. The results suggest that early in infection the VHS RNase degrades RNAs that activate PKR. Coupled with published data, the results suggest that inhibition of activation of PKR or its effect on viral replication is staged early in infection by VHS, postsynthesis of VP16 and VP22 by the γ₁34.5 protein, and very late in infection by the U_S11 protein.     

Relay of Herpes Simplex Virus between Langerhans Cells and Dermal Dendritic Cells in Human Skin

The mechanism by which immunity to Herpes Simplex Virus (HSV) is initiated is not completely defined. HSV initially infects mucosal epidermis prior to entering nerve endings. In mice, epidermal Langerhans cells (LCs) are the first dendritic cells (DCs) to encounter HSV, but it is CD103⁺ dermal DCs that carry viral antigen to lymph nodes for antigen presentation, suggesting DC cross-talk in skin. In this study, we compared topically HSV-1 infected human foreskin explants with biopsies of initial human genital herpes lesions to show LCs are initially infected then emigrate into the dermis. Here, LCs bearing markers of maturation and apoptosis formed large cell clusters with BDCA3⁺ dermal DCs (thought to be equivalent to murine CD103⁺ dermal DCs) and DC-SIGN⁺ DCs/macrophages. HSV-expressing LC fragments were observed inside the dermal DCs/macrophages and the BDCA3⁺ dermal DCs had up-regulated a damaged cell uptake receptor CLEC9A. No other infected epidermal cells interacted with dermal DCs. Correspondingly, LCs isolated from human skin and infected with HSV-1 in vitro also underwent apoptosis and were taken up by similarly isolated BDCA3⁺ dermal DCs and DC-SIGN⁺ cells. Thus, we conclude a viral antigen relay takes place where HSV infected LCs undergo apoptosis and are taken up by dermal DCs for subsequent antigen presentation. This provides a rationale for targeting these cells with mucosal or perhaps intradermal HSV immunization.     

The Herpes Simplex Virus Type 1 Infected Cell Protein 22

As one of the immediate-early(IE)proteins of herpes simplex virus type 1(HSV-1),ICP22 is a multifunctional viral regulator that localizes in the nucleus of infected cells.It is required in experimental animal systems and some nonhuman cell lines,but not in Vero or HEp-2 cells.ICP22 is extensively phosphorylated by viral and cellular kinases and nucleotidylylated by casein kinase Ⅱ.It has been shown to be required for efficient expression of early(E)genes and a subset of late(L)genes.ICP22,in conjunction wit...     

The Herpes Simplex Virus vhs Protein Induces Endoribonucleolytic Cleavage of Target RNAs in Cell Extracts

The herpes simplex virus virion host shutoff (vhs) protein (UL41 gene product) is a component of the HSV virion tegument that triggers shutoff of host protein synthesis and accelerated mRNA degradation during the early stages of HSV infection. Previous studies have demonstrated that extracts from HSV-infected cells and partially purified HSV virions display vhs-dependent RNase activity and that vhs is sufficient to trigger accelerated RNA degradation when expressed as the only HSV protein in an in vitro translation system derived from rabbit reticulocytes. We have used the rabbit reticulocyte translation system to characterize the mode of vhs-induced RNA decay in more detail. We report here that vhs-dependent RNA decay proceeds through endoribonucleolytic cleavage, is not affected by the presence of a 5' cap or a 3' poly(A) tail in the RNA substrate, requires Mg(2+), and occurs in the absence of ribosomes. Intriguingly, sites of preferential initial cleavage were clustered over the 5' quadrant of one RNA substrate that was characterized in detail. The vhs homologue of pseudorabies virus also induced accelerated RNA decay in this in vitro system.     

Proteins Specified by Herpes Simplex Virus XII. The Virion Polypeptides of Type 1 Strains

The polypeptides from purified virions of a herpes simplex 1 (human herpes-virus 1) strain, F1, which had been passaged a limited number of times in cell culture after isolation, formed 33 bands on electrophoretic separation in polyacrylamide gels cross-linked with N, N'-diallyltartardiamide in contrast to a maximum resolution of only 24 to 25 bands in gels cross-linked with N, N'-methylenebisacrylamide. This increase in the number of bands was due chiefly to an improved separation of glycosylated polypeptides from nonglycosylated polypeptides with which they co-electrophoresed on methylenebisacrylamide cross-linked gels. Purified virions of HSV-1 [F1] had a protein/DNA mass ratio of 10.7 +/- 0.96, and based on a DNA molecular mass of 85 x 10(6) to 100 x 10(6) the estimated weight of virion polypeptides ranges from 16.4 to 19.4 x 10(-16) g. The number of molecules of each polypeptide per virion ranged from less than 50 to 1,500. Comparison of the virion polypeptides of two HSV-1 strains with similar isolation and limited passage history with those of four HSV-1 strains with histories of numerous passages outside the human host showed a number of nonrandom variations in virion polypeptides. Thus, although the virion polypeptides of two strains with similar isolation and limited passage history could not be differentiated, strains with extended passage histories differed markedly from each other and from the limited passage strains in the number and electrophoretic mobility of noncapsid polypeptides and notably in those of the envelope.     

Virion Incorporation of the Herpes Simplex Virus Type 1 Tegument Protein VP22 Occurs via Glycoprotein E-Specific Recruitment to the Late Secretory Pathway

The mechanism by which herpesviruses acquire their tegument is not yet clear. One model is that outer tegument proteins are recruited by the cytoplasmic tails of viral glycoproteins. In the case of herpes simplex virus tegument protein VP22, interactions with the glycoproteins gE and gD have been shown. We have previously shown that the C-terminal half of VP22 contains the necessary signal for assembly into the virus. Here, we show that during infection VP22 interacts with gE and gM, as well as its tegument partner VP16. However, by using a range of techniques we were unable to demonstrate VP22 binding to gD. By using pulldown assays, we show that while the cytoplasmic tails of both gE and gM interact with VP22, only gE interacts efficiently with the C-terminal packaging domain of VP22. Furthermore, gE but not gM can recruit VP22 to the Golgi/trans-Golgi network region of the cell in the absence of other virus proteins. To examine the role of the gE-VP22 interaction in infection, we constructed a recombinant virus expressing a mutant VP22 protein with a 14-residue deletion that is unable to bind gE (ΔgEbind). Coimmunoprecipitation assays confirmed that this variant of VP22 was unable to complex with gE. Moreover, VP22 was no longer recruited to its characteristic cytoplasmic trafficking complexes but exhibited a diffuse localization. Importantly, packaging of this variant into virions was abrogated. The mutant virus exhibited poor growth in epithelial cells, similar to the defect we have observed for a VP22 knockout virus. These results suggest that deletion of just 14 residues from the VP22 protein is sufficient to inhibit binding to gE and hence recruitment to the viral envelope and assembly into the virus, resulting in a growth phenotype equivalent to that produced by deleting the entire reading frame.The herpesvirus tegument is the virion compartment located between the DNA-containing capsid and the virus envelope (6). Although it is well defined that the viral capsid assembles in the nucleus (37, 38) and the viral envelope is acquired from cellular membranes (3, 24), the mechanism of tegument protein acquisition is still to be established. At least 20 virus-encoded components are recruited into the herpes simplex virus type 1 (HSV-1) tegument (32), and there is increasing evidence to suggest that subsets of these proteins may be added as assembly progresses along the maturation pathway (28). To ensure efficient incorporation, it is likely that individual tegument proteins are specifically targeted to their cellular site of recruitment. Such targeting could involve interaction with a viral partner, a cellular partner, or both. A clearer understanding of how individual tegument proteins are acquired by newly assembling virions will help to define the herpesvirus assembly pathway.A number of protein-protein interactions between individual tegument proteins (13, 40, 42), and between tegument proteins and glycoproteins (19, 20, 22, 32), have been described that may provide useful insight into the assembly process. In particular, the interaction of tegument proteins with the cytoplasmic tails of virus glycoproteins provides an attractive mechanism for the virion recruitment of at least the outer components of the tegument. In the case of VP22, the homologues from pseudorabies virus (PRV) and HSV-1 have been shown to interact with the cytoplasmic tail of gE (19, 20, 32). However, the role of this interaction in virus infection has not yet been clearly defined and the fact that additional glycoprotein interactions have been described, with gM in the case of PRV and gD in the case of HSV-1, may point to potential redundancy in the mechanism of VP22 packaging (4, 19, 20). In addition, we and others have previously shown that HSV-1 VP22 interacts directly with a second tegument protein, namely, VP16 (13, 33), an interaction that could provide an alternative route for VP22 to enter the virion. In a previous study, we concluded that the region of VP22 containing its VP16 interaction domain was required but not sufficient for optimal VP22 packaging into the assembling virion, with an additional C-terminal determinant also involved (23). We also demonstrated that the same region of VP22 that was required for virion packaging was essential to target the protein to its characteristic cytoplasmic trafficking complexes, suggesting that these specific sites may be the location in the cell for VP22 assembly into the virion (23). Since that study, O''Regan and coworkers have reported that the C-terminal half of HSV-1 VP22 also contains the binding site for gE (32), providing a possible candidate for an additional VP22 binding partner. Furthermore, as HSV-1 VP22 has been shown to bind to gD and PRV VP22 interacts with gM, it is possible that the C terminus of VP22 contains a gD and/or a gM binding site (4, 20).In the present study, we aimed to clarify the molecular mechanism by which VP22 is recruited into the virus particle. We show that HSV-1 VP22 binds efficiently to VP16, gE, and gM in the infected cell, but we cannot detect an interaction with gD. We show that the packaging domain of VP22 binds to the cytoplasmic tail of gE but not gM and that the same region of VP22 is recruited to the secretory pathway by gE in the absence of other virus proteins. Finally, we show that a mutant VP22 protein lacking a 14-residue peptide from its packaging domain is unable to interact with gE during infection, exhibits a different subcellular localization, and fails to assemble into the virus particle. This is the first characterization of a single protein-protein interaction essential for the packaging of an HSV-1 tegument protein.     

Recognition of Herpes Simplex Virus Type 2 Tegument Proteins by CD4 T Cells Infiltrating Human Genital Herpes Lesions

The local cellular immune response to herpes simplex virus (HSV) is important in the control of recurrent HSV infection. The antiviral functions of infiltrating CD4-bearing T cells may include cytotoxicity, inhibition of viral growth, lymphokine secretion, and support of humoral and CD8 responses. The antigens recognized by many HSV-specific CD4 T cells localizing to genital HSV-2 lesions are unknown. T cells recognizing antigens encoded within map units 0.67 to 0.73 of HSV DNA are frequently recovered from herpetic lesions. Expression cloning with this region of DNA now shows that tegument protein VP22 and the viral dUTPase, encoded by genes U_L49 and U_L50, respectively, are T-cell antigens. Separate epitopes in VP22 were defined for T-cell clones from each of three patients. Reactivity with the tegument protein encoded by U_L21 was identified for an additional patient. Three new epitopes were identified in VP16, a tegument protein associated with VP22. Some tegument-specific CD4 T-cell clones exhibited cytotoxic activity against HSV-infected cells. These results suggest that herpes simplex tegument proteins are processed for antigen presentation in vivo and are possible candidate compounds for herpes simplex vaccines.     

The Amino Terminus of Herpes Simplex Virus Type 1 Glycoprotein K (gK) Modulates gB-Mediated Virus-Induced Cell Fusion and Virion Egress

Herpes simplex virus type 1 (HSV-1)-induced cell fusion is mediated by viral glycoproteins and other membrane proteins expressed on infected cell surfaces. Certain mutations in the carboxyl terminus of HSV-1 glycoprotein B (gB) and in the amino terminus of gK cause extensive virus-induced cell fusion. Although gB is known to be a fusogenic glycoprotein, the mechanism by which gK is involved in virus-induced cell fusion remains elusive. To delineate the amino-terminal domains of gK involved in virus-induced cell fusion, the recombinant viruses gKΔ31-47, gKΔ31-68, and gKΔ31-117, expressing gK carrying in-frame deletions spanning the amino terminus of gK immediately after the gK signal sequence (amino acids [aa] 1 to 30), were constructed. Mutant viruses gKΔ31-47 and gKΔ31-117 exhibited a gK-null (ΔgK) phenotype characterized by the formation of very small viral plaques and up to a 2-log reduction in the production of infectious virus in comparison to that for the parental HSV-1(F) wild-type virus. The gKΔ31-68 mutant virus formed substantially larger plaques and produced 1-log-higher titers than the gKΔ31-47 and gKΔ31-117 mutant virions at low multiplicities of infection. Deletion of 28 aa from the carboxyl terminus of gB (gBΔ28syn) caused extensive virus-induced cell fusion. However, the gBΔ28syn mutation was unable to cause virus-induced cell fusion in the presence of the gKΔ31-68 mutation. Transient expression of a peptide composed of the amino-terminal 82 aa of gK (gKa) produced a glycosylated peptide that was efficiently expressed on cell surfaces only after infection with the HSV-1(F), gKΔ31-68, ΔgK, or UL20-null virus. The gKa peptide complemented the gKΔ31-47 and gKΔ31-68 mutant viruses for infectious-virus production and for gKΔ31-68/gBΔ28syn-mediated cell fusion. These data show that the amino terminus of gK modulates gB-mediated virus-induced cell fusion and virion egress.Herpes simplex virus type 1 (HSV-1) specifies at least 11 virally encoded glycoproteins, as well as several nonglycosylated and lipid-anchored membrane-associated proteins, which serve important functions in virion infectivity and virus spread. Although cell-free enveloped virions can efficiently spread viral infection, virions can also spread by causing cell fusion of adjacent cellular membranes. Virus-induced cell fusion, which is caused by viral glycoproteins expressed on infected cell surfaces, enables transmission of virions from one cell to another, avoiding extracellular spaces and exposure of free virions to neutralizing antibodies (reviewed in reference 56). Most mutations that cause extensive virus-induced cell-to-cell fusion (syncytial or syn mutations) have been mapped to at least four regions of the viral genome: the UL20 gene (5, 42, 44); the UL24 gene (37, 58); the UL27 gene, encoding glycoprotein B (gB) (9, 51); and the UL53 gene, coding for gK (7, 15, 35, 53, 54, 57).Increasing evidence suggests that virus-induced cell fusion is mediated by the concerted action of glycoproteins gD, gB, and gH/gL. Recent studies have shown that gD interacts with both gB and gH/gL (1, 2). Binding of gD to its cognate receptors, including Nectin-1, HVEM, and others (12, 29, 48, 59, 60, 62, 63), is thought to trigger conformation changes in gH/gL and gB that cause fusion of the viral envelope with cellular membranes during virus entry and virus-induced cell fusion (32, 34). Transient coexpression of gB, gD, and gH/gL causes cell-to-cell fusion (49, 68). However, this phenomenon does not accurately model viral fusion, because other viral glycoproteins and membrane proteins known to be important for virus-induced cell fusion are not required (6, 14, 31). Specifically, gK and UL20 were shown to be absolutely required for virus-induced cell fusion (21, 46). Moreover, syncytial mutations within gK (7, 15, 35, 53, 54, 57) or UL20 (5, 42, 44) promote extensive virus-induced cell fusion, and viruses lacking gK enter more slowly than wild-type virus into susceptible cells (25). Furthermore, transient coexpression of gK carrying a syncytial mutation with gB, gD, and gH/gL did not enhance cell fusion, while coexpression of the wild-type gK with gB, gD, and gH/gL inhibited cell fusion (3).Glycoproteins gB and gH are highly conserved across all subfamilies of herpesviruses. gB forms a homotrimeric type I integral membrane protein, which is N glycosylated at multiple sites within the polypeptide. An unusual feature of gB is that syncytial mutations that enhance virus-induced cell fusion are located exclusively in the carboxyl terminus of gB, which is predicted to be located intracellularly (51). Single-amino-acid substitutions within two regions of the intracellular cytoplasmic domain of gB were shown to cause syncytium formation and were designated region I (amino acid [aa] positions 816 and 817) and region II (aa positions 853, 854, and 857) (9, 10, 28, 69). Furthermore, deletion of 28 aa from the carboxyl terminus of gB, disrupting the small predicted alpha-helical domain H17b, causes extensive virus-induced cell fusion as well as extensive glycoprotein-mediated cell fusion in the gB, gD, and gH/gL transient-coexpression system (22, 49, 68). The X-ray structure of the ectodomain of gB has been determined and is predicted to assume at least two major conformations, one of which may be necessary for the fusogenic properties of gB. Therefore, perturbation of the carboxyl terminus of gB may alter the conformation of the amino terminus of gB, thus favoring one of the two predicted conformational structures that causes membrane fusion (34).The UL53 (gK) and UL20 genes encode multipass transmembrane proteins of 338 and 222 aa, respectively, which are conserved in all alphaherpesviruses (15, 42, 55). Both proteins have multiple sites where posttranslational modification can occur; however, only gK is posttranslationally modified by N-linked carbohydrate addition (15, 35, 55). The specific membrane topologies of both gK and UL20 protein (UL20p) have been predicted and experimentally confirmed using epitope tags inserted within predicted intracellular and extracellular domains (18, 21, 44). Syncytial mutations in gK map predominantly within extracellular domains of gK and particularly within the amino-terminal portion of gK (domain I) (18), while syncytial mutations of UL20 are located within the amino terminus of UL20p, shown to be located intracellularly (44). A series of recent studies have shown that HSV-1 gK and UL20 functionally and physically interact and that these interactions are necessary for their coordinate intracellular transport and cell surface expression (16, 18, 21, 26, 45). Specifically, direct protein-protein interactions between the amino terminus of HSV-1 UL20 and gK domain III, both of which are localized intracellularly, were recently demonstrated by two-way coimmunoprecipitation experiments (19).According to the most prevalent model for herpesvirus intracellular morphogenesis, capsids initially assemble within the nuclei and acquire a primary envelope by budding into the perinuclear spaces. Subsequently, these virions lose their envelope through fusion with the outer nuclear lamellae. Within the cytoplasm, tegument proteins associate with the viral nucleocapsid and final envelopment occurs by budding of cytoplasmic capsids into specific trans-Golgi network (TGN)-associated membranes (8, 30, 47, 70). Mature virions traffic to cell surfaces, presumably following the cellular secretory pathway (33, 47, 61). In addition to their significant roles in virus-induced cell fusion, gK and UL20 are required for cytoplasmic virion envelopment. Viruses with deletions in either the gK or the UL20 gene are unable to translocate from the cytoplasm to extracellular spaces and accumulated as unenveloped virions in the cytoplasm (5, 15, 20, 21, 26, 35, 36, 38, 44, 55). Current evidence suggests that the functions of gK and UL20 in cytoplasmic virion envelopment and virus-induced cell fusion are carried out by different, genetically separable domains of UL20p. Specifically, UL20 mutations within the amino and carboxyl termini of UL20p allowed cotransport of gK and UL20p to cell surfaces, virus-induced cell fusion, and TGN localization, while effectively inhibiting cytoplasmic virion envelopment (44, 45).In this paper, we demonstrate that the amino terminus of gK expressed as a free peptide of 82 aa (gKa) is transported to infected cell surfaces by viral proteins other than gK or UL20p and facilitates virus-induced cell fusion caused by syncytial mutations in the carboxyl terminus of gB. Thus, functional domains of gK can be genetically separated, as we have shown previously (44, 45), as well as physically separated into different peptide portions that retain functional activities of gK. These results are consistent with the hypothesis that the amino terminus of gK directly or indirectly interacts with and modulates the fusogenic properties of gB.     

Role for Plasmacytoid Dendritic Cells in the Immune Control of Recurrent Human Herpes Simplex Virus Infection

Plasmacytoid dendritic cells (pDC) are an important component of the innate immune response, producing large amounts of alpha interferon in response to viral stimulation in vitro. Under noninflammatory conditions, pDC are not found in the skin and are restricted in location to the blood and lymph nodes. Therefore, their role in mucosal and cutaneous herpes simplex virus (HSV) infection has not been well-defined. In this study we show a role for human pDC in the immune response to HSV infection. First, by confocal microscopy we showed that pDC infiltrate the dermis of recurrent genital herpes simplex lesions at early and late phases, often at the dermo-epidermal junction. We then showed that pDC in vitro are resistant to HSV infection despite expressing the entry receptors CD111, CD112, and HVE-A. Within the lesions, pDC were found closely associated with CD3⁺ lymphocytes and NK cells, especially those which were activated (CD69⁺). Furthermore, these HSV-exposed pDC were able to stimulate virus-specific autologous T-lymphocyte proliferation. We conclude from this work that pDC may contribute to the immune control of recurrent herpes virus infection in vivo.     

The Effect of Helicobacter hepaticus Infection on Immune Responses Specific to Herpes Simplex Virus Type 1 and Characteristics of Dendritic Cells

Infection of mice with Helicobacter hepaticus is common in research colonies, yet little is known about how this persistent infection affects immunologic research. The goal of this study was to determine whether H. hepaticus infection status can modulate immune responses specific to herpes simplex virus type 1 (HSV1) and the phenotypic and functional characteristics of dendritic cells (DC) of mice. We compared virus-specific antibody and T cell-mediated responses in H. hepaticus-infected and noninfected mice that were inoculated intranasally with HSV1. The effect of H. hepaticus on the HSV1-specific antibody and T cell-mediated immune responses in superficial cervical and tracheobronchal lymph nodes (LN) did not reach statistical significance. Surface expression of the maturation-associated markers CD40, CD80, CD86, and MHC II and percentages of IL12p40- and TNFα-producing DC from spleen and colic LN in H. hepaticus-infected mice and noninfected mice were measured in separate experiments. Expression of CD40, CD86, and MHC II and percentages of IL12p40- and TNFα-producing DC from colic LN were decreased in H. hepaticus-infected mice. In contrast, H. hepaticus infection did not reduce the expression of these molecules by splenic DC. Expression of CD40, CD80, CD86, and MHC II on splenic DC from H. hepaticus-infected mice was increased after in vitro lipopolysaccharide stimulation. These results indicate that H. hepaticus infection can influence the results of immunologic assays in mice and support the use of H. hepaticus-free mice in immunologic research.Abbreviations: DC, dendritic cells; HSV1, herpes simplex virus type 1; LN, lymph nodes; MHC II, major histocompatibility complex class II; MHV, mouse hepatitis virus; OVA, ovalbumin peptide SIINFEKL; PE, phycoerythrinHelicobacter hepaticus is a gram-negative, microaerophilic, curved to spiral-shaped bacterium with bipolar, sheathed flagella. H. hepaticus was described for the first time in 1994 as the cause of chronic active hepatitis associated with a high incidence of hepatocellular neoplasms in mice on a long-term toxicology study.³⁹ Since then, H. hepaticus has been identified as a common contaminant of mouse colonies at a variety of research institutions. Although commercial breeders produce H. hepaticus-free animals, many mouse colonies at public and private research institutions still harbor H. hepaticus. A recent survey found H. hepaticus-infected mice in 59% of commercial and academic institutions in Canada, Europe, Asia, Australia, and the United States.³⁵H. hepaticus persistently colonizes the hepatic bile canaliculi and the cecal and colonic mucosa of mice.^9,39 Infection can cause chronic active hepatitis, hepatocellular neoplasms, and typhlocolitis, which vary in severity depending on the strain, age, gender, and immune status of the mouse.^5,9,11,39 In adult immunocompetent mice, H. hepaticus infection is usually asymptomatic. However, immune-dysregulated mice can develop inflammatory bowel disease, which may present as rectal prolapse or diarrhea.¹⁶Mice initiate immune responses against H. hepaticus primarily through its interaction with Toll-like receptor 2 on antigen-presenting cells.²¹ Both systemic and local (at the site of infection) H. hepaticus-specific Th1-type immune responses are induced in immunocompetent mice.^26,40 Systemic antibody and cell-mediated immunity against the bacteria persist for at least 46 wk after experimental inoculation.⁴⁰ Gene expression profiles of cecal tissue of H. hepaticus-infected mice have shown that inflammatory responses differ depending on the mouse strain. For example, A/JCr mice had significant and prolonged expression of the Th1-type cytokines IFNγ and IL12p40 in cecal mucosa, and these expression levels persisted for at least 3 mo after H. hepaticus infection. However, C57BL/6 mice had a lesser elevation of IFNγ gene expression without an effect on IL12p40. IFN γ expression waned by 1 mo after inoculation in C57BL/6 mice.²⁵ In addition, H. hepaticus-specific secretory IgA antibodies are persistently detected in the feces of mice.⁴⁰ How these immune responses in H. hepaticus-infected mice might affect immunologic research is unknown.The goal of this study was to determine whether immune responses to herpes simplex virus type 1 (HSV1) and the phenotypic and functional characteristics of dendritic cells (DC) are altered in H. hepaticus-infected mice. The intranasal HSV1 infection model is used widely to study immune mechanisms in mice. Immunity to HSV1 consists of virus-neutralizing antibodies in the serum and virus-specific T cells in the draining LN. Superficial cervical and mediastinal LN have been described as draining LN for intranasal HSV1 infection.² The response to HSV1 infection peaks at 7 d after infection and leads to clearance of the viral load.² In this study, we compared levels of HSV1-specific antibody and T cell-mediated immune responses between H. hepaticus-infected and noninfected mice.Dendritic cells are important components of the immune system that play a role in antigen processing and presentation. On exposure to foreign antigen, DC mature and express increased levels of major histocompatibility complex class II proteins (MHC II), CD40, CD80, and CD86 on the cell surface. These maturation-associated cell surface markers interact with naive T and B cells to initiate antibody- and cell-mediated immune responses against foreign antigens.²⁷ In addition, mature DC secrete proinflammatory cytokines, including TNFα and IL12p40. These cytokines lead to increased vascular permeability, complement activity, lymphocyte activation, lymphocyte proliferation, and increased antibody production.²⁷ To determine whether infection with H. hepaticus affects characteristics of DC, we measured the expression of the maturation-associated cell surface markers CD40, CD80, CD86, and MHC II and proinflammatory cytokines IL12p40 and TNFα by DC derived from the spleen and colic LN of H. hepaticus-infected and noninfected mice. Our findings indicate that H. hepaticus infection can influence the various aspects of immune responsiveness and, therefore, must be considered as a potential variable in studies in which immune function is a measurable outcome.     

The Pattern of Tegument-Capsid Interaction in the Herpes Simplex Virus Type 1 Virion Is Not Influenced by the Small Hexon-Associated Protein VP26

Examination of the three-dimensional structure of intact herpes simplex virus type 1 (HSV-1) virions had revealed that the icosahedrally symmetrical interaction between the tegument and capsid involves the pentons but not the hexons (Z. H. Zhou, D. H. Chen, J. Jakana, F. J. Rixon, and W. Chiu, J. Virol. 73:3210-3218, 1999). To account for this, we postulated that the presence of the small capsid protein, VP26, on top of the hexons was masking potential binding sites and preventing tegument attachment. We have now tested this hypothesis by determining the structure of virions lacking VP26. Apart from the obvious absence of VP26 from the capsids, the structures of the VP26 minus and wild-type virions were essentially identical. Notably, they showed the same tegument attachment patterns, thereby demonstrating that VP26 is not responsible for the divergent tegument binding properties of pentons and hexons.