Spread Of Herpes Simplex Virus Type Research Articles

Olfactory and trigeminal routes of HSV-1 CNS infection with regional microglial heterogeneity.

Herpes simplex virus type 1 (HSV-1) primarily targets the oral and nasal epithelia before establishing latency in the trigeminal ganglion (TG) and other peripheral ganglia. HSV-1 can also infect and become latent in the central nervous system (CNS) independent of latency in the TGs. Recent studies suggest entry to the CNS via two distinct routes: the TG-brainstem connection and olfactory nerve; however, to date, there is no characterization of brain regions targeted during HSV-1 primary infection. Furthermore, the immune response by microglia may also contribute to the heterogeneity between different brain regions. However, the response to HSV-1 by microglia has not been characterized in a region-specific manner. This study investigated the time course of HSV-1 spread within the olfactory epithelium (OE) and CNS following intranasal inoculation and the corresponding macrophage/microglial response in a C57BL/6 mouse model. We found an apical to basal spread of HSV-1 within the OE and underlying tissue accompanied by an inflammatory response of macrophages. OE infection was followed by infection of a small subset of brain regions targeted by the TG in the brainstem and other cranial nerve nuclei, including the vagus and hypoglossal nerve. Furthermore, other brain regions were positive for HSV-1 antigens, such as the locus coeruleus (LC), raphe nucleus (RaN), and hypothalamus while sparing the hippocampus and cortex. Within each brain region, microglia activation also varied widely. These findings provide critical insights into the region-specific dissemination of HSV-1 within the CNS, elucidating potential mechanisms linking viral infection to neurological and neurodegenerative diseases.IMPORTANCEThis study shows how herpes simplex virus type 1 (HSV-1) spreads within the brain after infecting the nasal passages. Our data reveal the distinct pattern of HSV-1 through the brain during a non-encephalitic infection. Furthermore, microglial activation was also temporally and spatially specific, with some regions of the brain having sustained microglial activation even in the absence of viral antigens. Previous reports have identified specific brain regions found to be positive for HSV-1 infection; however, to date, there has not been a concise investigation of the anatomical spread of HSV-1 and the brain regions consistently vulnerable to viral entry and spread. Understanding these region-specific differences in infection and immune response is crucial because it links HSV-1 infection to potential triggers for neurological and neurodegenerative diseases.

The antimicrobial protein RNase 7 directly restricts herpes simplex virus infection of human keratinocytes.

Approximately 22% of moderately to severely affected atopic dermatitis (AD) patients have a history of eczema herpeticum, a disseminated rash primarily caused by herpes simplex virus type 1 (HSV-1). Reduced activity of antimicrobial peptides may contribute to the increased susceptibility of AD patients to HSV-1. We previously demonstrated that the antimicrobial protein RNase 7 limits HSV-1 infection of human keratinocytes by promoting self-DNA sensing. Here, we addressed whether RNase 7 has any effect on HSV-1 infection when infecting keratinocytes without exogenously added costimulatory DNA, and which step(s) of the infection cycle RNase 7 interferes with. We quantified viral gene expression by RT-qPCR and flow cytometry, viral genome replication by qPCR, virucidal effects by plaque titration, and plaque formation and the subcellular localization of incoming HSV-1 particles by microscopy. Recombinant RNase 7 restricted HSV-1 gene expression, genome replication, and plaque formation in human keratinocytes. It decreased HSV-1 immediate-early transcripts independently of the induction of interferon-stimulated genes. Its main effect was on intracellular infection processes and not on extracellular virions or virus binding to cells. RNase 7 reduced the amount of cell-associated capsids and the HSV-1 envelope glycoprotein D at 3 but not at 0.5 h postinfection. Our data show that RNase 7 directly restricts HSV-1 infection of human keratinocytes, possibly by promoting the degradation of incoming HSV-1 particles. This suggests that RNase 7 may limit HSV-1 spread in the skin and that mechanisms that reduce its activity in the lesional skin of AD patients may increase their susceptibility to eczema herpeticum.

OPTN-TBK1 axis and a role for PLK1 in HSV-1 infection.

Herpes simplex virus type 1 (HSV-1) is globally prevalent, with latent infections observed in up to 80% of the population. The virus is known for subverting host defense mechanisms and infiltrating the nervous system to establish latency in peripheral ganglia. Multiple stressors can reactivate the virus, and recurrent herpes has been linked to vision loss and neurodegeneration. Identifying critical host factors that limit the spread of HSV-1 and the subsequent establishment of latent infection holds the potential to drive new intervention strategies for eradicating the virus. Numerous pieces of evidence underscore the significance of Tank-binding kinase 1 (TBK1) in restricting HSV-1. Reports have also suggested that phosphorylation of optineurin (OPTN) by TBK1 is required for triggering OPTN-mediated autophagy for HSV degradation. This report adds new insights into the roles of OPTN and TBK1 in HSV-1 infection and provides proof of a TBK1-independent HSV-1 restriction through OPTN. It confirms that TBK1 activation can be substituted by PLK1 to provide protection against HSV-1. In contrast, the activation of OPTN is likely an indispensable host defense mechanism for optimal defense against HSV-1.

Estimation of Seroprevalence of Anti-Herpes Simplex Virus Type-1 IgG Among Healthy Blood Donors in Sakaka City, Aljouf, Saudi Arabia

Background: Herpes simplex virus type-1 (HSV-1) is a highly infectious neurotropic virus. The data on HSV-1 infection in Saudi Arabia, including the seroprevalence of HSV-1 antibodies, are scarce. Objectives: This is the first study to evaluate the prevalence of anti-HSV-1 immunoglobulin G (IgG) in donated blood in Sakaka, Aljouf, Saudi Arabia. Methods: A total of 300 donated blood samples were collected from the Blood Bank of Prince Mutaib Bin Abdulaziz Hospital in Sakaka. Sensitive and specific enzyme-linked immunosorbent assay (ELISA) was used to detect anti-HSV-1 IgG. A comparison of the age, gender, education, occupation, income, hand hygiene, travel history, and cupping practice of blood donors stratified for the extent of anti-HSV-1 IgG was made. Results: There was a low prevalence of anti-HSV-1 IgG (20%; n = 60/300). Moreover, 50.0% of IgG-positive participants were in the age group of 41 - 45 years, and 81.7% of the participants had a household income of < 10000 SAR (statistically highly significant; P < 0.001*). All the participants performed hand washing with soap before handling food and after using the toilet. Furthermore, IgG-positive participants had a bachelor’s degree (50.0%), were governmental employees (60.0%), were international travelers (50.0%), and practiced cupping (50.0%) with statistically significant associations (P < 0.05*). Conclusions: The current study’s findings support previous reports about the key importance of improving socioeconomic conditions and hygiene measures in reducing the spread of HSV-1. The present study provides an alarm regarding reaching the age of sexual debut without acquiring protective anti-HSV-1 immunoglobulins, consequently becoming more susceptible to acquiring HSV-1 infection through the genital route. These data support the urgent need to develop an effective anti-HSV-1 vaccine.

Role of tear exosomes in the spread of herpes simplex virus type 1 in recurrent herpes simplex keratitis.

Herpes simplex keratitis (HSK) is the most common but serious infectious keratitis with high recurrence. It is predominantly caused by herpes simplex virus type 1 (HSV-1). The spread mechanism of HSV-1 in HSK is not entirely clear. Multiple publications indicate that exosomes participate in the intercellular communication process during viral infections. However, there is rare evidence that HSV-1 spreads in HSK by exosomal pathway. This study aims to investigate the relationship between the spread of HSV-1 and tear exosomes in recurrent HSK. Tear fluids collected from total 59 participants were included in this study. Tear exosomes were isolated by ultracentrifugation, then identified by silver staining and western blot. The size was determined by dynamic light scattering (DLS). The viral biomarkers were identified by western blot. The cellular uptake of exosomes was studied using labelled exosomes. Tear exosomes were indeed enriched in tear fluids. Collected exosomes own normal diameters consistent with related reports. The exosomal biomarkers existed in tear exosomes. Labelled exosomes were successfully taken up by human corneal epithelial cells (HCEC) in large numbers in a short time. After cellular uptake, HSK biomarkers were detectable by western blot in infected cells. Tear exosomes should be the latent sites of HSV-1 in recurrent HSK and might be involved in the spread of HSV-1. Besides, this study verifies HSV-1 genes can be indeed transferred between cells by exosomal pathway, providing new inspiration for the clinical intervention and treatment as well as the drug discovery of recurrent HSK.

Broad-spectrum antiviral diazadispiroalkane core molecules block attachment and cell-to-cell spread of herpesviruses

Regarding the problems with the current available drugs many research studies deal with the class of the dispirotripiperazine (DSTP)-based compounds. These are small molecules consisting of polycyclic saturated ring systems with positively charged nitrogen atoms. These compounds can interact with negatively charged HSPGs and thus block viral attachment. In a previous paper by Adfeldt et al. (2021), we have shown that the diazadispiroalkane derivatives 11826091 and 11826236 exhibit dose-dependent antiviral activity against human cytomegalovirus (HCMV) and pseudorabies virus (PrV). In the present study, these two small molecules are evaluated against two other herpesvirus species, murine cytomegalovirus (MCMV) and herpes simplex virus type 1 (HSV-1), as well as a HCMV clinical isolate. They exhibit potent antiherpetic activity against these herpesviruses with a high selectivity index. The low cytotoxicity was underlined by the LD50 determination in mice. We have shown that inhibition occurs at an early stage of infection. Interestingly, 11826091 and 11826236 reduced immediate early gene expression in HCMV and HSV-1 infected cells in a dose-dependent manner. Both small molecules probably interact electrostatically with sulfated glycosaminoglycans (GAGs) of proteoglycans on target cells resulting in blockage of adsorption sites for herpesvirus glycoprotein. Moreover, both compounds showed significant effects against the cell-associated viral spread of HSV-1 and HCMV. Overall, this study shows that 11826091 and 11826236 represent two promising candidates for a new approach of a broad antiviral therapy.

Alzheimer's Gone Viral: Could Herpes Simplex Virus Type-1 Be Stealing Your Memories?

Alzheimer's disease (AD) is one of the principal causes of disability and morbidity. It is one of the most expensive illnesses. Despite this, there are no significant data regarding its etiology and optimal treatment. This review concentrates on the viral hypothesis of AD. After a comprehensive PubMed literature search, we analyzed the studies associating herpes simplex virus type-1 (HSV1) infection to AD from the previous 10 years. Molecular mechanisms whereby HSV1 induces AD-related pathophysiology, including neuronal production and accumulation of amyloid-beta (amyloid-β), abnormal phosphorylation of tau proteins, impaired calcium homeostasis, and autophagy, are addressed. The virus also imitates the disease in other ways, showing increased neuroinflammation, oxidative stress, synaptic dysfunction, and neuronal apoptosis. Serological studies correlate HSV1 infection with AD and cognitive impairment. A causal link between HSV1 and AD raises the concept of a simple, efficient, and preventive treatment alternative. Anti-viral agents impede brain degeneration by preventing HSV1 spread and its replication, decreasing hyperphosphorylated tau and amyloid-β; thus providing an efficacious treatment for AD. We also mention brown algae, intravenous immunoglobulin (IVIG), and a synthetic drug, BAY57-1293, with anti-viral properties, as options for treating AD. We want to recommend future researchers to look for more affordable, non-invasive, and swifter techniques to identify HSV1 in the brain and assist in the early detection and prevention of AD.

Deletion of the Pseudorabies Virus gE/gI-US9p complex disrupts kinesin KIF1A and KIF5C recruitment during egress, and alters the properties of microtubule-dependent transport in vitro.

During infection of neurons by alphaherpesviruses including Pseudorabies virus (PRV) and Herpes simplex virus type 1 (HSV-1) viral nucleocapsids assemble in the cell nucleus, become enveloped in the cell body then traffic into and down axons to nerve termini for spread to adjacent epithelia. The viral membrane protein US9p and the membrane glycoprotein heterodimer gE/gI play critical roles in anterograde spread of both HSV-1 and PRV, and several models exist to explain their function. Biochemical studies suggest that PRV US9p associates with the kinesin-3 motor KIF1A in a gE/gI-stimulated manner, and the gE/gI-US9p complex has been proposed to recruit KIF1A to PRV for microtubule-mediated anterograde trafficking into or along the axon. However, as loss of gE/gI-US9p essentially abolishes delivery of alphaherpesviruses to the axon it is difficult to determine the microtubule-dependent trafficking properties and motor-composition of Δ(gE/gI−US9p) particles. Alternatively, studies in HSV-1 have suggested that gE/gI and US9p are required for the appearance of virions in the axon because they act upstream, to help assemble enveloped virions in the cell body. We prepared Δ(gE/gI-US9p) mutant, and control parental PRV particles from differentiated cultured neuronal or porcine kidney epithelial cells and quantitated the efficiency of virion assembly, the properties of microtubule-dependent transport and the ability of viral particles to recruit kinesin motors. We find that loss of gE/gI-US9p has no significant effect upon PRV particle assembly but leads to greatly diminished plus end-directed traffic, and enhanced minus end-directed and bidirectional movement along microtubules. PRV particles prepared from infected differentiated mouse CAD neurons were found to be associated with either kinesin KIF1A or kinesin KIF5C, but not both. Loss of gE/gI-US9p resulted in failure to recruit KIF1A and KF5C, but did not affect dynein binding. Unexpectedly, while KIF5C was expressed in undifferentiated and differentiated CAD neurons it was only found associated with PRV particles prepared from differentiated cells.

A synthetic glycosaminoglycan mimetic blocks HSV-1 infection in human iris stromal cells

Herpes simplex virus type-1 (HSV-1) is a significant pathogen that affects vision by targeting multiple regions in the human eye including iris. Using a focused library of synthetic non-saccharide glycosaminoglycan mimetics (NSGMs), we identified sulfated pentagalloylglucoside (SPGG) as a potent inhibitor of HSV-1 entry and cell-to-cell spread in the primary cultures of human iris stromal (HIS) cells isolated from eye donors. Using in vitro β-galactosidase reporter assay and plaque reduction assay, SPGG was found to inhibit HSV-1 entry in a dosage-dependent manner (IC50 ∼6.0 μM). Interestingly, a pronounced inhibition in HSV-1 entry and spread was observed in HIS cells, or a cell line expressing specific gD-receptor, when virions were pre-treated with mimetics suggesting a possible interaction between SPGG and the HSV-1 glycoprotein. To examine the significance of gD-SPGG interaction, HIS cells were pretreated with SPGG, which showed a significant reduction in gD binding. Taken together, our results provide strong evidence of SPGG being a novel viral entry inhibitor against ocular HSV infection.

Extracellular Vesicles in Herpes Viral Spread and Immune Evasion.

Extracellular vesicles (EVs) are involved in numerous processes during infections by both enveloped and non-enveloped viruses. Among them, herpes simplex virus type-1 (HSV-1) modulates secretory pathways, allowing EVs to exit infected cells. Many characteristics regarding the mechanisms of viral spread are still unidentified, and as such, secreted vesicles are promising candidates due to their role in intercellular communications during viral infection. Another relevant role for EVs is to protect virions from the action of neutralizing antibodies, thus increasing their stability within the host during hematogenous spread. Recent studies have suggested the participation of EVs in HSV-1 spread, wherein virion-containing microvesicles (MVs) released by infected cells were endocytosed by naïve cells, leading to a productive infection. This suggests that HSV-1 might use MVs to expand its tropism and evade the host immune response. In this review, we briefly describe the current knowledge about the involvement of EVs in viral infections in general, with a specific focus on recent research into their role in HSV-1 spread. Implications of the autophagic pathway in the biogenesis and secretion of EVs will also be discussed.

Innate defense mechanisms against HSV-1 infection in the target tissues, skin and brain.

Herpes simplex virus type 1 (HSV-1) initiates productive infection in mucocutaneous tissues to cause cold sores and establishes latent infection in the trigeminal ganglia. Under certain circumstances, HSV-1 may cause encephalitis. Here, we compared host innate defenses against HSV-1 in the two clinically relevant tissues, skin and brain, using a unique ex vivo system of organ culture. Upon HSV-1 infection and spread, apoptosis induction was observed in the skin, but not in brain tissues. While the two tissues elicited interferon (IFN-β) response upon HSV1 infection, IFN induction was more robust in the skin compared to the brain. Moreover, antiviral response to exogenous IFNβ treatment was much stronger in the skin compared to brain tissues. This observation was not related to the availability of the IFN receptor on cells' surface. Taken together, our study demonstrates differential innate antiviral responses to HSV-1 infection that may be exploited in future development of selective and tissue-specific anti-viral treatments.

A STING-dependent innate-sensing pathway mediates resistance to corneal HSV-1 infection via upregulation of the antiviral effector tetherin.

Type 1 interferons (IFNs; IFNα/β) mediate immunological host resistance to numerous viral infections, including herpes simplex virus type 1 (HSV-1). The pathways responsible for IFNα/β signaling during the innate immune response to acute HSV-1 infection in the cornea are incompletely understood. Using a murine ocular infection model, we hypothesized that the stimulator of IFN genes (STING) mediates resistance to HSV-1 infection at the ocular surface and preserves the structural integrity of this mucosal site. Viral pathogenesis, tissue pathology, and host immune responses during ocular HSV-1 infection were characterized by plaque assay, esthesiometry, pachymetry, immunohistochemistry, flow cytometry, and small interfering RNA transfection in wild-type C57BL/6 (WT), STING-deficient (STING(-/-)), and IFNα/β receptor-deficient (CD118(-/-)) mice at days 3-5 postinfection. The presence of STING was critical for sustained control of HSV-1 replication in the corneal epithelium and resistance to viral neuroinvasion, but loss of STING had a negligible impact with respect to gross tissue pathology. Auxiliary STING-independent IFNα/β signaling pathways were responsible for maintenance of corneal integrity. Lymphatic vessels, mast cells, and sensory innervation were compromised in CD118(-/-) mice concurrent with increased tissue edema. STING-dependent signaling led to the upregulation of tetherin, a viral restriction factor we identify is important in containing the spread of HSV-1 in vivo.

Synthetic pregnenolone derivatives as antiviral agents against acyclovir-resistant isolates of Herpes Simplex Virus Type 1

The conventional therapy for the management of Herpes Simplex Virus Type 1 (HSV-1) infections mainly comprises acyclovir (ACV) and other nucleoside analogues. A common outcome of this treatment is the emergence of resistant viral strains, principally when immunosuppressed patients are involved. Thus, the development of new antiherpetic compounds remains as a central challenge. In this work we describe the synthesis and the in vitro antiherpetic activity of a new family of steroidal compounds derived from the endogenous hormone pregnenolone. Some of these derivatives showed a remarkable inhibitory effect on HSV-1 spread both on wild type and ACV-resistant strains. The results also show that these compounds seem to interfere with the late steps of the viral cycle.

Zebrafish 3-O-Sulfotransferase-4 Generated Heparan Sulfate Mediates HSV-1 Entry and Spread

Rare modification of heparan sulfate (HS) by glucosaminyl 3-O sulfotransferase (3-OST) isforms generates an entry receptor for herpes simplex virus type-1 (HSV-1). In the zebrafish (ZF) model multiple 3-OST isoforms are differentially expressed. One such isoform is 3-OST-4 which is widely expressed in the central nervous system of ZF. In this report we characterize the role of ZF encoded 3-OST-4 isoform for HSV-1 entry. Expression of ZF 3-OST-4 into resistant Chinese hamster ovary (CHO-K1) cells promoted susceptibility to HSV-1 infection. This entry was 3-O sulfated HS (3-OS HS) dependent as pre-treatment of ZF 3-OST-4 cells with enzyme HS lyases (heparinase II/III) significantly reduced HSV-1 entry. Interestingly, co-expression of ZF 3-OST-4 along with ZF 3-OST-2 which is also expressed in brain rendered cells more susceptible to HSV-1 than 3-OST-4 alone. The role of ZF-3-OST-4 in the spread of HSV-1 was also evaluated as CHO-K1 cells that expressed HSV-1 glycoproteins fused with ZF 3-OST-4 expressing effector CHO-K1 cells. Finally, adding further evidence ZF 3-OST-4 mediated HSV-1 entry was inhibited by anti-3O HS G2 peptide. Taken together our results demonstrate a role for ZF 3-OST-4 in HSV-1 pathogenesis and support the use of ZF as a model to study it.

Different modes of herpes simplex virus type 1 spread in brain and skin tissues

Herpes simplex virus type 1 (HSV-1) initially infects the skin and subsequently spreads to the nervous system. To investigate and compare HSV-1 mode of propagation in the two clinically relevant tissues, we have established ex vivo infection models, using native tissues of mouse and human skin, as well as mouse brain, maintained in organ cultures. HSV-1, which is naturally restricted to the human, infects and spreads in the mouse and human skin tissues in a similar fashion, thus validating the mouse model. The spread of HSV-1 in the skin was concentric to form typical plaques of limited size, predominantly of cytopathic cells. By contrast, HSV-1 spread in the brain tissue was directed along specific neuronal networks with no apparent cytopathic effect. Two additional differences were noted following infection of the skin and brain tissues. First, only a negligible amount of extracellular progeny virus was produced of the infected brain tissues, while substantial quantity of infectious progeny virus was released to the media of the infected skin. Second, antibodies against HSV-1, added following the infection, effectively restricted viral spread in the skin but have no effect on viral spread in the brain tissue. Taken together, these results reveal that HSV-1 spread within the brain tissue mostly by direct transfer from cell to cell, while in the skin the progeny extracellular virus predominates, thus facilitating the infection to new individuals.

Plus-end tracking proteins, CLASPs, and a viral Akt mimic regulate herpesvirus-induced stable microtubule formation and virus spread

Although microtubules (MTs) frequently form highly dynamic networks, subsets of MTs become stabilized in response to environmental cues and function as specialized tracks for vesicle and macromolecular trafficking. MT stabilization is controlled by specialized plus-end tracking proteins (+TIPs) whose accumulation at the MT ends is facilitated by the end-binding protein, EB1, and regulated by various signaling pathways. As cargoes themselves, viruses are dependent on MTs for their intracellular movement. Although many viruses affect MT organization, the potential contribution of MT stabilization by +TIPs to infection remains unknown. Here we show that early in infection of primary human fibroblasts, herpes simplex virus type 1 (HSV-1) disrupts the centrosome, the primary MT organizing center in many cell types. As infection progresses HSV-1 induces the formation of stable MT subsets through inactivation of glycogen synthase kinase 3beta by the viral Ser/Thr kinase, Us3. Stable MT formation is reduced in cells infected with Us3 mutants and those stable MTs that form cluster around the trans-Golgi network. Downstream of glycogen synthase kinase 3beta, cytoplasmic linker-associated proteins (CLASPs), specialized host +TIPs that control MT formation at the trans-Golgi network and cortical capture, are specifically required for virus-induced MT stabilization and HSV-1 spread. Our findings demonstrate the biological importance of +TIPs to viral infection and suggest that HSV-1 has evolved to exploit the trans-Golgi network as an alternate MT organizing center to facilitate virus spread.

Effect of black tea extract on herpes simplex virus-1 infection of cultured cells

BackgroundThe purpose of this investigation was to determine if black tea extract (BTE), consisting primarily of flavanol compounds called theaflavins, could inhibit herpes simplex virus type-1 (HSV-1) infection in cultured A549 (human epithelial) and Vero cells.MethodsThe effect of BTE both on A549 and Vero cultured cells and on HSV-1 was assessed by using phase contrast and fluorescent microscopy, and cell viability and proliferation assays. After establishing the maximum non-cytotoxic concentration of BTE, A549 and Vero cells and HSV-1 virions were treated with varying concentrations of BTE, respectively. A549 and Vero cells were infected with HSV-1 with green fluorescent protein (GFP) insert at the UL46 gene. The effect of infectivity was determined by viral DNA extraction followed by PCR, plaque assays, adsorption assays, and electrophoresis of PCR products.ResultsBTE was not cytotoxic to A549 and Vero cells, as confirmed by cell viability and proliferation assays, in which BTE treated groups paralleled the positive control group. For both cell lines, plaque assays and fluorescent microscopy indicated an inverse relationship between BTE concentration (from 0.14 μM – 1.4 mM) and HSV-1 infectivity. Specifically, PCR and electrophoresis showed a reduction in the viral genome following treatment with BTE. In addition, there was a noticeable decrease in the amount of viral plaques for BTE treated samples in the adsorption assays.ConclusionsBTE consisting primarily of theaflavins is not cytotoxic and can reduce or block the production of infectious HSV-1 virions in cultured A549 and Vero cells, thus inhibiting the infectivity of the virus by interfering in the attachment, penetration and viral DNA replication of HSV-1 particles. These findings indicate that BTE enriched with theaflavins has the potential to be developed as a safe, therapeutic antiviral agent to prevent the spread of HSV-1.

Role of Filopodia in HSV-1 Entry into Zebrafish 3-O-Sulfotransferase-3-Expressing Cells

Heparan sulfate proteoglycans (HSPGs) modified by zebrafish (ZF) encoded glucosaminyl 3-O sulfotransferase-3 (3-OST-3) generate a receptor for herpes simplex virus type-1 (HSV-1) entry and spread. In order to elucidate the mechanism by which HSV-1 enters into ZF-3-OST-3 cells, we investigated the mode of viral entry. Under high resolution scanning electron microscopy (SEM), actin cytoskeleton changes were observed by a dramatic increase in the number of filopodia formed during early interactions of HSV-1 with the target cells. While the increase in number was common among all the infected cells, the highest numbers of filopodia was observed in cells expressing the 3-OST-3 modified form of heparan sulfate (HS) encoded either by human or ZF. The levels of viral infection and filopodia induction were reduced with the actin polymerization inhibitors, Cytochalasin-D and Lantriculin B, suggesting an important role for actin reorganization during ZF-3-OST-3 mediated HSV-1 entry. Supporting an interesting possibility of filopodia usage during HSV-1 spread, pre-treatment of cytochalasin D in ZF-3-OST-3 cells drastically reduced virus glycoprotein induced cell fusion. Taken together, our results provide new evidence on the involvement of filopodia during HSV-1 infection of ZF-3-OST-3 cells and confirm a role for modified heparan sulfate in cytoskeleton rearrangement during HSV-1 entry.

Zebrafish encoded 3-O-sulfotransferase-2 generated heparan sulfate serves as a receptor during HSV-1 entry and spread

Previously we reported the role of zebrafish (ZF) encoded glucosaminyl 3-O-sulfotransferase-3 (3-OST-3) isoform in assisting herpes simplex virus type-1 (HSV-1) entry and spread by generating an entry receptor to HSV-1 envelope glycoprotein D (gD). However, the ability of ZF encoded 3-OST-2 isoform to participate in HSV-1 entry has not been determined although it is predominantly expressed in ZF brain, a prime target for HSV-1 to infect and establish lifelong latency. Here we report the expression cloning of ZF encoded 3-OST-2 isoform and demonstrate HSV-1 entry into resistant Chinese hamster ovary (CHO-K1) cells expressing the clone. Additional significance of ZF encoded 3-OST-2 receptor was demonstrated using medically important isolates of HSV-1. In addition, interference to HSV-1 entry was observed upon co-expression of HSV-1 gD and ZF 3-OST-2. Similarly HSV-1 entry was significantly inhibited by the pre-treatment of cells with enzyme HS lyases (heparinase II/III). Finally, ZF-3-OST-2 expressing CHO-K1 was able to fuse with HSV-1 glycoprotein expressing cells suggesting their role in HSV-1 spread. Taken together our result demonstrates a role for ZF 3-OST-2 in HSV-1 pathogenesis.

Members of 3-O-Sulfotransferases (3-OST) Family: A Valuable Tool from Zebrafish to Humans for Understanding Herpes Simplex Virus Entry

The journey of many viruses to infect cells begins when the virus first binds to cell surface heparan sulfate (HS). The initial step of cell attachment or binding during herpes simplex virus type-1 (HSV-1) entry is mediated by envelope glycoprotein B (gB) and C (gC). The binding is followed by fusion between virus envelope and cell membrane during which HSV-1 glycoprotein D (gD) interacts with a modified form of HS know as 3-O-sulfated heparan sulfate (3-OS HS). The rare modification of 3-O-sulfation on HS chain is governed by enzymes known as 3-O-sulfotransferase (3-OST). Currently, there are seven isoforms of human 3-OSTs that have been identified, and with the exception of 3-OST-1, all other 3-OST isoforms allow HSV-1 entry and spread. Recently, the product of the zebrafish (ZF)-encoded 3-OST-3 was also recognized as a gD receptor, which mediates HSV-1 entry and cell-cell fusion similar to human 3-OST-3. Interestingly, the ZF system expresses multiple isoforms of 3-OST which could be very useful for studying the involvement of HS and 3-OS HS in virus tropism and virus-induced inflammation. In addition, therapeutic targeting of 3-OST generated HS is likely to bring about novel interventions against HSV-1. In this review we have taken a closer look at the potential of both human and ZF encoded 3-OSTs as valuable tools in HSV entry and inflammation studies.