Herpesvirus Entry Mediator and Nectin-1 Mediate Herpes Simplex Virus 1 Infection of the Murine Cornea

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The cellular proteins nectin-1 and herpesvirus entry mediator (HVEM) can both mediate the entry of herpes simplex virus 1 (HSV-1). We have recently shown how these receptors contribute to infection of skin by investigating HSV-1 entry into murine epidermis. Ex vivo infection studies reveal nectin-1 as the primary receptor in epidermis, whereas HVEM has a more limited role. Although the epidermis represents the outermost layer of skin, the contribution of nectin-1 and HVEM in the underlying dermis is still open. Here, we analyzed the role of each receptor during HSV-1 entry in murine dermal fibroblasts that were deficient in expression of either nectin-1 or HVEM or both receptors. Because infection was not prevented by the absence of either nectin-1 or HVEM, we conclude that they can act as alternative receptors. Although HVEM was found to be highly expressed on fibroblasts, entry was delayed in nectin-1-deficient cells, suggesting that nectin-1 acts as the more efficient receptor. In the absence of both receptors, entry was strongly delayed leading to a much reduced viral spread and virus production. These results suggest an unidentified cellular component that acts as alternate but inefficient receptor for HSV-1 on dermal fibroblasts. Characterization of the cellular entry mechanism suggests that HSV-1 can enter dermal fibroblasts both by direct fusion with the plasma membrane and via endocytic vesicles and that this is not dependent on the presence or absence of nectin-1. Entry was also shown to require dynamin and cholesterol, suggesting comparable entry pathways in keratinocytes and dermal fibroblasts. IMPORTANCEHerpes simplex virus (HSV) is a human pathogen which infects its host via mucosal surfaces or abraded skin. To understand how HSV-1 overcomes the protective barrier of mucosa or skin and reaches its receptors in tissue, it is essential to know which receptors contribute to the entry into individual skin cells. Previously, we have explored the contribution of nectin-1 and herpesvirus entry mediator (HVEM) as receptors for HSV-1 entry into murine epidermis, where keratinocytes form the major cell type. Since the underlying dermis consists primarily of fibroblasts, we have now extended our study of HSV-1 entry to dermal fibroblasts isolated from nectin-1-or HVEM-deficient mice or from mice deficient in both receptors. Our results demonstrate a role for both nectin-1 and HVEM as receptors and suggest a further receptor which appears much less efficient.T o initiate infection, herpes simplex virus 1 (HSV-1) enters its human host via mucosal surfaces or abraded skin. HSV-1 entry into individual cells involves the interaction of several viral glycoproteins with various cell surface receptors (1, 2). The first step during entry is the attachment of virions to glycosaminoglycans, which facilitates the interaction with cellular receptors, leading to the fusion of the viral envelope with a cellular membrane. Fusion can either occur with the plasma membrane or with vesicle membranes after virions are int...

Section: Discussionmentioning

confidence: 93%

“…Since the absence of both nectin-1 and HVEM prevents HSV pathogenesis in the mouse model, nectin-1 and HVEM are reported to be the dominant functional gD receptors in the murine host (11)(12)(13). Using nectin-1-or HVEM-deficient mice, we recently investigated HSV-1 entry into murine epidermis.…”

mentioning

confidence: 99%

Role of Nectin-1 and Herpesvirus Entry Mediator as Cellular Receptors for Herpes Simplex Virus 1 on Primary Murine Dermal Fibroblasts

Petermann

Rahn

Thier

et al. 2015

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“…As noted above, prior studies using intravaginal or intracranial inoculation did not show any effect of knocking out HVEM on HSV-2 disease in mice (10,11). In an ocular scarification infection model using adult mice, Karaba et al demonstrated that disease due to HSV-1 was attenuated in the absence of HVEM expression (12). However, studies using HSV-2 in this model led to results which differ from those with HSV-1, showing no dependence of disease characteristics on HVEM expression (A. H. Karaba, S. J. Kopp, and R. Longnecker, submitted for publication).…”

Section: Discussionmentioning

confidence: 95%

“…Adult nectin-1 KO mice were resistant to encephalitis after direct intracerebral inoculation with HSV-2 (11). Interestingly, compared to that in WT mice, disease was attenuated in both HVEM KO and nectin-1 KO mice after corneal infection with HSV-1 (12).…”

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confidence: 96%

Pathogenesis of Neonatal Herpes Simplex 2 Disease in a Mouse Model Is Dependent on Entry Receptor Expression and Route of Inoculation

Kopp

Karaba

Cohen

et al. 2013

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cHerpes simplex virus (HSV) pathogenesis in mice differs based on availability of the principal entry receptors herpesvirus entry mediator (HVEM) and nectin-1 in a manner dependent upon route of inoculation. After intravaginal or intracranial inoculation of adult mice, nectin-1 is a major mediator of neurologic disease, while the absence of either receptor attenuates disease after ocular infection. We tested the importance of receptor availability and route of infection on disease in mouse models of neonatal HSV. We infected 7-day-old mice lacking neither or one principal HSV receptor or both principal HSV receptors with HSV-2 via a peripheral route (intranasal), via a systemic route (intraperitoneal), or by inoculation directly into the central nervous system (intracranial). Mortality, neurologic disease, and visceral dissemination of virus were significantly attenuated in nectin-1 knockout mice compared with HVEM knockout or wild-type mice after intranasal inoculation. Mice lacking both entry receptors (double-knockout mice) showed no evidence of disease after inoculation by any route. Nectin-1 knockout mice had delayed mortality after intraperitoneal inoculation relative to wild-type and HVEM knockout mice, but virus was able to spread to the brain and viscera in all genotypes except double-knockout mice. Unlike in adult mice, HVEM was sufficient to mediate disease in neonatal mice after direct intracranial inoculation, and the absence of HVEM delayed time to mortality relative to that of wild-type mice. Additionally, in wild-type neonatal mice inoculated intracranially, HSV antigen did not primarily colocalize with NeuNpositive neurons. Our results suggest that differences in receptor expression between adults and newborns may partially explain differences in susceptibility to HSV-2. H erpes simplex virus (HSV) causes neonatal infection in about 1 in 3,200 live births in the United States (1). More than half of infants with neonatal HSV disease have disseminated disease or encephalitis (2), which, despite effective antiviral treatment, results in the deaths of more than 25% of those with disseminated disease and in neurologic morbidity in more than two-thirds of survivors of encephalitis (3). Relative to other populations infected with HSV, newborns have the highest rates of dissemination and central nervous system (CNS) disease (4). Although differences in immune responses from those of adults have been implicated, precise reasons for the increased severity of disease in infants remain unknown (5).Infection of susceptible human and mouse cells by HSV requires binding of the viral glycoprotein gD with one of its cell surface receptors (6, 7). HSV gD binds to three general classes of surface receptors, including herpesvirus entry mediator (HVEM), nectin-1 and Ϫ2, and specific sites in heparan sulfate (7). Of these, HVEM and nectin-1 appear to mediate viral entry most efficiently in both humans and mice (8, 9). The mouse receptors are orthologous to the human receptors, and HSV disease in mice resembles that i...

“…This method is clearly distinct from the natural entry route of HSV, but these studies helped us to understand pathogenesis. Expression of nectin-1 and HVEM was observed in the epithelium of the murine cornea (15,16), and infection studies with HSV-1 in KO mice indicated attenuated disease in the absence of either HVEM or nectin-1 (17). In contrast, HSV-2 does not require HVEM to cause disease in corneal infections, suggesting that the HVEM requirement depends on the serotype (18).…”

mentioning

confidence: 99%

Entry Mechanisms of Herpes Simplex Virus 1 into Murine Epidermis: Involvement of Nectin-1 and Herpesvirus Entry Mediator as Cellular Receptors

Petermann

Thier

Rahn

et al. 2015

Skin keratinocytes represent a primary entry site for herpes simplex virus 1 (HSV-1) in vivo. The cellular proteins nectin-1 and herpesvirus entry mediator (HVEM) act as efficient receptors for both serotypes of HSV and are sufficient for disease development mediated by HSV-2 in mice. How HSV-1 enters skin and whether both nectin-1 and HVEM are involved are not known. We addressed the impact of nectin-1 during entry of HSV-1 into murine epidermis and investigated the putative contribution of HVEM. Using ex vivo infection of murine epidermis, we showed that HSV-1 entered the basal keratinocytes of the epidermis very efficiently. In nectin-1-deficient epidermis, entry was strongly reduced. Almost no entry was observed, however, in nectin-1-deficient keratinocytes grown in culture. This observation correlated with the presence of HVEM on the keratinocyte surface in epidermis and with the lack of HVEM expression in nectin-1-deficient primary keratinocytes. Our results suggest that nectin-1 is the primary receptor in epidermis, while HVEM has a more limited role. For primary murine keratinocytes, on which nectin-1 acts as a single receptor, electron microscopy suggested that HSV-1 can enter both by direct fusion with the plasma membrane and via endocytic vesicles. Thus, we concluded that nectin-1 directs internalization into keratinocytes via alternative pathways. In summary, HSV-1 entry into epidermis was shown to strongly depend on the presence of nectin-1, but the restricted presence of HVEM can potentially replace nectin-1 as a receptor, illustrating the flexibility employed by HSV-1 to efficiently invade tissue in vivo. H erpes simplex viruses (HSV) are ubiquitous human pathogens which can cause a range of diseases, from mild, uncomplicated mucocutaneous lesions to life-threatening infections. HSV-1 is dominantly associated with orofacial infections and encephalitis, whereas HSV-2 more likely causes genital infections. To enter its human host, HSV must come into contact with mucosal surfaces, skin, or the cornea. During initial exposure on mucosa or skin, HSV targets epidermal keratinocytes and establishes a primary infection in the epithelium. Cellular entry of HSV relies on the interaction of several viral glycoproteins with various cell surface receptors (1, 2). The envelope glycoprotein D (gD) is essential for the entry process, and only after gD binding to a receptor is fusion with a cellular membrane induced (3). The major gD receptors mediating entry into mouse and human cells are herpesvirus entry mediator (HVEM) and nectin-1 (4-6). HVEM is a member of the tumor necrosis factor receptor superfamily which can activate either proinflammatory or inhibitory signaling pathways (7), while nectin-1 is an immunoglobulin-like cell adhesion molecule (8). A further gD receptor is 3-O-sulfated heparan sulfate, which may also play a role in HSV-1 entry into various cell types (9, 10).