Deletion of a Predicted β-Sheet Domain within the Amino Terminus of Herpes Simplex Virus Glycoprotein K Conserved among Alphaherpesviruses Prevents Virus Entry into Neuronal Axons

Jambunathan, Nithya; Charles, Anu Susan; Subramanian, R.; Saied, Ahmad; Naderi, Misagh; Rider, Paul J. F.; Bryliński, Michał; Chouljenko, Vladimir N.; Kousoulas, Konstantin G.

doi:10.1128/jvi.02468-15

Cited by 26 publications

(34 citation statements)

References 84 publications

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“…To support the results from the PLA, two-way coimmunoprecipitation assays were performed using anti-gB and anti-Akt-1 monoclonal antibodies, and the presence of gB and Akt-1 in immunoblots of infected SK-N-SH cell lysates and in immunoprecipitates was detected with either anti-gB or anti-Akt-1 antibody. Similar amounts of gB were detected in either McKrae-or gKΔ31-68 mutant-infected lysates, with gB appearing as a two major protein species migrating with apparent molecular masses of 130 and 120 kDa, most likely representing the high-mannose precursor and the fully glycosylated species, respectively, as we have reported previously (54,81,82). Similar amounts of Akt-1(S473) were detected in both McKrae-and gKΔ31-68 mutant-infected SK-N-SH cell McKrae gKΔ31-68 (100 PFU) for 1 h at 4°C.…”

Section: Resultssupporting

confidence: 78%

“…A miltefosine concentration of 50 M and higher was toxic to the SK-N-SH cells, while at concentrations of up to 30 M there were no apparent toxic effects observed. Briefly, PLA reveals the deposition of capsids into the cytoplasm of infected cells by detecting the interaction of the HSV-1 UL37 protein with dynein, while cell surface-bound virions are detected via the interaction of gD with nectin-1 (81). Both viruses attached to cell surfaces equally well; however, at the maximum concentration of miltefosine tested (30 M, since 100 M miltefosine was toxic to SK-N-SH cells), McKrae wild-type virus entry was drastically inhibited, while gKΔ31-68 virus entry was unaffected (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…An MOI of 10 was used for SK-N-SH cells. Virus entry was detected using PLA following 1 h of infection in the case of SK-N-SH cells, as described previously (81), Briefly, PLA detects the deposition of capsids into the cytoplasm of infected cells by detecting the interaction of the HSV-1 UL37 protein with dynein, while cell surface-bound virions are detected via the interaction of gD with nectin-1 (81). Similar numbers of McKrae and gKΔ31-68 virions were attached to SK-N-SH cells, as detected by PLA (Fig.…”

Section: Resultsmentioning

confidence: 99%

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The Amino Terminus of Herpes Simplex Virus 1 Glycoprotein K (gK) Is Required for gB Binding to Akt, Release of Intracellular Calcium, and Fusion of the Viral Envelope with Plasma Membranes

Musarrat

Jambunathan

Rider

et al. 2018

J Virol

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Previously, we have shown that the amino terminus of glycoprotein K (gK) binds to the amino terminus of gB and that deletion of the amino-terminal 38 amino acids of gK prevents herpes simplex virus 1 (HSV-1) infection of mouse trigeminal ganglia after ocular infection and virus entry into neuronal axons. Recently, it has been shown that gB binds to Akt during virus entry and induces Akt phosphorylation and intracellular calcium release. Proximity ligation and two-way immunoprecipitation assays using monoclonal antibodies against gB and Akt-1 phosphorylated at S473 [Akt-1(S473)] confirmed that HSV-1(McKrae) gB interacted with Akt-1(S473) during virus entry into human neuroblastoma (SK-N-SH) cells and induced the release of intracellular calcium. In contrast, the gB specified by HSV-1(McKrae) gKΔ31-68, lacking the amino-terminal 38 amino acids of gK, failed to interact with Akt-1(S473) and induce intracellular calcium release. The Akt inhibitor miltefosine inhibited the entry of McKrae but not the gKΔ31-68 mutant into SK-N-SH cells. Importantly, the entry of the gKΔ31-68 mutant but not McKrae into SK-N-SH cells treated with the endocytosis inhibitors pitstop-2 and dynasore hydrate was significantly inhibited, indicating that McKrae gKΔ31-68 entered via endocytosis. These results suggest that the amino terminus of gK functions to regulate the fusion of the viral envelope with cellular plasma membranes. HSV-1 glycoprotein B (gB) functions in the fusion of the viral envelope with cellular membranes during virus entry. Herein, we show that a deletion in the amino terminus of glycoprotein K (gK) inhibits gB binding to Akt-1(S473), the release of intracellular calcium, and virus entry via fusion of the viral envelope with cellular plasma membranes.

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Section: Resultssupporting

confidence: 78%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

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The Amino Terminus of Herpes Simplex Virus 1 Glycoprotein K (gK) Is Required for gB Binding to Akt, Release of Intracellular Calcium, and Fusion of the Viral Envelope with Plasma Membranes

Musarrat

Jambunathan

Rider

et al. 2018

J Virol

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“…Herpesviruses are double-stranded DNA, enveloped viruses with intricate envelopes that contain at least a dozen proteins (2). Herpesvirus entry is a complex process that requires three conserved proteins plus additional nonconserved glycoproteins specific to individual herpesviruses (3)(4)(5) and could be further modulated by viral and host proteins (6)(7)(8)(9).…”

mentioning

confidence: 99%

Characterization of Vesicular Stomatitis Virus Pseudotypes Bearing Essential Entry Glycoproteins gB, gD, gH, and gL of Herpes Simplex Virus 1

Rogalin

Heldwein

2016

J Virol

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Herpes simplex viruses (HSVs) are unusual in that unlike most enveloped viruses, they require at least four entry glycoproteins, gB, gD, gH, and gL, for entry into target cells in addition to a cellular receptor for gD. The dissection of the herpes simplex virus 1 (HSV-1) entry mechanism is complicated by the presence of more than a dozen proteins on the viral envelope. To investigate HSV-1 entry requirements in a simplified system, we generated vesicular stomatitis virus (VSV) virions pseudotyped with HSV-1 essential entry glycoproteins gB, gD, gH, and gL but lacking the native VSV fusogen G. These virions, referred to here as VSV⌬G-BHLD virions, infected a cell line expressing a gD receptor, demonstrating for the first time that the four essential entry glycoproteins of HSV-1 are not only required but also sufficient for cell entry. To our knowledge, this is the first time the VSV pseudotyping system has been successfully extended beyond two proteins. Entry of pseudotyped virions required a gD receptor and was inhibited by HSV-1 specific anti-gB or anti-gH/gL neutralizing antibodies, which suggests that membrane fusion during the entry of the pseudotyped virions shares common requirements with the membrane fusion involved in HSV-1 entry and HSV-1-mediated syncytium formation. The HSV pseudotyping system established in this study presents a novel tool for systematic exploration of the HSV entry and membrane fusion mechanisms. IMPORTANCEHerpes simplex viruses (HSVs) are human pathogens that can cause cold sores, genital herpes, and blindness. No vaccines or preventatives are available. HSV entry into cells-a prerequisite for a successful infection-is a complex process that involves multiple viral and host proteins and occurs by different routes. Detailed mechanistic knowledge of the HSV entry is important for understanding its pathogenesis and would benefit antiviral and vaccine development, yet the presence of more than a dozen proteins on the viral envelope complicates the dissection of the HSV entry mechanisms. In this study, we generated heterologous virions displaying the four essential entry proteins of HSV-1 and showed that they are capable of cell entry and, like HSV-1, require all four entry glycoproteins along with a gD receptor. This HSV pseudotyping system pioneered in this work opens doors for future systematic exploration of the herpesvirus entry mechanisms.T o enter living cells to replicate, viruses must overcome the barrier of the cellular membrane. Enveloped viruses accomplish this task by facilitating the merger of their envelope with a target cell membrane, during which capsids are delivered into the cytosol and infection ensues. Entry is initiated by binding of a virus to an appropriate receptor on the surface of the host cell and is catalyzed by a virus-encoded membrane fusogen. In most enveloped viruses, the receptor binding and the fusogenic functions are executed by a single protein (1).Herpesviruses are double-stranded DNA, enveloped viruses with intricate envelopes that contain...

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“…The HSV homolog of ORF39 is UL20, which also associates with gK. Moreover, recent studies that used an HSV gKΔ31–68 mutant demonstrated that gK is implicated in neurotropism [78]. Whether VZV gK has similar roles as a neurotropic factor needs to be investigated.…”

Section: Glycoproteins That Function In Virion Attachmentmentioning

confidence: 99%

Varicella-Zoster Virus Glycoproteins: Entry, Replication, and Pathogenesis

Oliver

Yang

Arvin

2016

Curr Clin Micro Rpt

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Varicella-zoster virus (VZV), an alphaherpesvirus that causes chicken pox (varicella) and shingles (herpes zoster), is a medically important pathogen that causes considerable morbidity and, on occasion, mortality in immunocompromised patients. Herpes zoster can afflict the elderly with a debilitating condition, postherpetic neuralgia, triggering severe, untreatable pain for months or years. The lipid envelope of VZV, similar to all herpesviruses, contains numerous glycoproteins required for replication and pathogenesis. Purpose of Review To summarize the current knowledge about VZV glycoproteins and their roles in cell entry, replication and pathogenesis. Recent Findings The functions for some VZV glycoproteins are known, such as gB, gH and gL in membrane fusion, cell-cell fusion regulation, and receptor binding properties. However, the molecular mechanisms that trigger or mediate VZV glycoproteins remains poorly understood. Summary VZV glycoproteins are central to successful replication but their modus operandi during replication and pathogenesis remain elusive requiring further mechanistic based studies.

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Deletion of a Predicted β-Sheet Domain within the Amino Terminus of Herpes Simplex Virus Glycoprotein K Conserved among Alphaherpesviruses Prevents Virus Entry into Neuronal Axons

Cited by 26 publications

References 84 publications

The Amino Terminus of Herpes Simplex Virus 1 Glycoprotein K (gK) Is Required for gB Binding to Akt, Release of Intracellular Calcium, and Fusion of the Viral Envelope with Plasma Membranes

The Amino Terminus of Herpes Simplex Virus 1 Glycoprotein K (gK) Is Required for gB Binding to Akt, Release of Intracellular Calcium, and Fusion of the Viral Envelope with Plasma Membranes

Characterization of Vesicular Stomatitis Virus Pseudotypes Bearing Essential Entry Glycoproteins gB, gD, gH, and gL of Herpes Simplex Virus 1

Varicella-Zoster Virus Glycoproteins: Entry, Replication, and Pathogenesis

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