scholarly journals HveA (Herpesvirus Entry Mediator A), a Coreceptor for Herpes Simplex Virus Entry, also Participates in Virus-Induced Cell Fusion

1998 ◽  
Vol 72 (7) ◽  
pp. 5802-5810 ◽  
Author(s):  
Tracy Terry-Allison ◽  
Rebecca I. Montgomery ◽  
J. Charles Whitbeck ◽  
Ruliang Xu ◽  
Gary H. Cohen ◽  
...  
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Cell Fusion ◽  
Viral Entry ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Simplex Virus ◽  
Herpesvirus Entry Mediator ◽  
Hsv 1 ◽  
Cell Cell

ABSTRACT The purpose of this study was to determine whether a cell surface protein that can serve as coreceptor for herpes simplex virus type 1 (HSV-1) entry, herpesvirus entry mediator (previously designated HVEM but renamed HveA), also mediates HSV-1-induced cell-cell fusion. We found that transfection of DNA from KOS-804, a previously described HSV-1 syncytial (Syn) strain whose Syn mutation was mapped to an amino acid substitution in gK, induced numerous large syncytia on HveA-expressing Chinese hamster ovary cells (CHO-HVEM12) but not on control cells (CHO-C8). Antibodies specific for gD as well as for HveA were effective inhibitors of KOS-804-induced fusion, consistent with previously described direct interactions between gD and HveA. Since mutations in gD determine the ability of HSV-1 to utilize HveA for entry, we examined whether the form of virally expressed gD also influenced the ability of HveA to mediate fusion. We produced a recombinant virus carrying the KOS-804 Syn mutation and the KOS-Rid1 gD mutation, which significantly reduces viral entry via HveA, and designated it KOS-SR1. KOS-SR1 DNA had a markedly reduced ability to induce syncytia on CHO-HVEM12 cells and a somewhat enhanced ability to induce syncytia on CHO-C8 cells. These results support previous findings concerning the relative abilities of KOS and KOS-Rid1 to infect CHO-HVEM12 and CHO-C8 cells. Thus, HveA mediates cell-cell fusion as well as viral entry and both activities of HveA are contingent upon the form of gD expressed by the virus.

scholarly journals Structure-Function Analysis of Herpes Simplex Virus Type 1 gD and gH-gL: Clues from gDgH Chimeras

2003 ◽  
Vol 77 (12) ◽  
pp. 6731-6742 ◽  
Author(s):  
Tina M. Cairns ◽  
Richard S. B. Milne ◽  
Manuel Ponce-de-Leon ◽  
Deanna K. Tobin ◽  
Gary H. Cohen ◽  
...  
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Cell Surface ◽  
Cell Fusion ◽  
Virus Type ◽  
Herpes Simplex Virus Type ◽  
Simplex Virus ◽  
Hsv 1 ◽  
Cell Cell

ABSTRACT In alphaherpesviruses, glycoprotein B (gB), gD, gH, and gL are essential for virus entry. A replication-competent gL-null pseudorabies virus (PrV) (B. G. Klupp and T. C. Mettenleiter, J. Virol. 73:3014-3022, 1999) was shown to express a gDgH hybrid protein that could replace gD, gH, and gL in cell-cell fusion and null virus complementation assays. To study this phenomenon in herpes simplex virus type 1 (HSV-1), we constructed four gDgH chimeras, joining the first 308 gD amino acids to various gH N-terminal truncations. The chimeras were named for the first amino acid of gH at which each was truncated: 22, 259, 388, and 432. All chimeras were immunoprecipitated with both gD and gH antibodies to conformational epitopes. Normally, transport of gH to the cell surface requires gH-gL complex formation. Chimera 22 contains full-length gH fused to gD308. Unlike PrV gDgH, chimera 22 required gL for transport to the surface of transfected Vero cells. Interestingly, although chimera 259 failed to reach the cell surface, chimeras 388 and 432 exhibited gL-independent transport. To examine gD and gH domain function, each chimera was tested in cell-cell fusion and null virus complementation assays. Unlike PrV gDgH, none of the HSV-1 chimeras substituted for gL for fusion. Only chimera 22 was able to replace gH for fusion and could also replace either gH or gD in the complementation assay. Surprisingly, this chimera performed very poorly as a substitute for gD in the fusion assay despite its ability to complement gD-null virus and bind HSV entry receptors (HveA and nectin-1). Chimeras 388 and 432, which contain the same portion of gD as that in chimera 22, substituted for gD for fusion at 25 to 50% of wild-type levels. However, these chimeras functioned poorly in gD-null virus complementation assays. The results highlight the fact that these two functional assays are measuring two related but distinct processes.

scholarly journals Herpes Simplex Virus Glycoprotein K, but Not Its Syncytial Allele, Inhibits Cell-Cell Fusion Mediated by the Four Fusogenic Glycoproteins, gD, gB, gH, and gL

2003 ◽  
Vol 77 (12) ◽  
pp. 6836-6844 ◽  
Author(s):  
Elisa Avitabile ◽  
Giulia Lombardi ◽  
Gabriella Campadelli-Fiume
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Cell Fusion ◽  
Vero Cells ◽  
Giant Cells ◽  
Glycoprotein K ◽  
Herpes Simplex Virus Glycoprotein ◽  
Simplex Virus ◽  
Hsv 1 ◽  
Cell Cell

ABSTRACT A Myc epitope was inserted at residue 283 of herpes simplex virus type 1 (HSV-1) glycoprotein K (gK), a position previously shown not to interfere with gK activity. The Myc-tagged gK localized predominantly to the endoplasmic reticulum, both in uninfected and in HSV-infected cells. gK, coexpressed with the four HSV fusogenic glycoproteins, gD, gB, gH, and gL, inhibited cell-cell fusion. The effect was partially dose dependent and was observed both in baby hamster kidney (BHK) and in Vero cells, indicating that the antifusion activity of gK may be cell line independent. The antifusion activity of gK did not require viral proteins other than the four fusogenic glycoproteins. A syncytial (syn) allele of gK (syn-gK) carrying the A40V substitution present in HSV-1(MP) did not block fusion to the extent seen with the wild-type (wt) gK, indicating that the syn mutation ablated, at least in part, the antifusogenic activity of wt gK. We conclude that gK is part of the mechanism whereby HSV negatively regulates its own fusion activity. Its effect accounts for the notion that cells infected with wt HSV do not fuse with adjacent, uninfected cells into multinucleated giant cells or syncytia. gK may also function to preclude fusion between virion envelope and the virion-encasing vesicles during virus transport to the extracellular compartment, thus preventing nucleocapsid de-envelopment in the cytoplasm.

scholarly journals Insertion Mutations in Herpes Simplex Virus 1 Glycoprotein H Reduce Cell Surface Expression, Slow the Rate of Cell Fusion, or Abrogate Functions in Cell Fusion and Viral Entry

2009 ◽  
Vol 84 (4) ◽  
pp. 2038-2046 ◽  
Author(s):  
Julia O. Jackson ◽  
Erick Lin ◽  
Patricia G. Spear ◽  
Richard Longnecker
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Cell Surface ◽  
Cell Fusion ◽  
Viral Entry ◽  
Cytoplasmic Tail ◽  
Target Cells ◽  
Simplex Virus ◽  
Insertion Mutations ◽  
Hsv 1

ABSTRACT Of the four required herpes simplex virus (HSV) entry glycoproteins, the precise role of gH-gL in fusion remains the most elusive. The heterodimer gH-gL has been proposed to mediate hemifusion after the interaction of another required glycoprotein, gD, with a receptor. To identify functional domains of HSV-1 gH, we generated 22 randomized linker-insertion mutants. Analyses of 22 gH mutants revealed that gH is relatively tolerant of insertion mutations, as 15 of 22 mutants permitted normal processing and transport of gH-gL to the cell surface. gH mutants that were not expressed well at the cell surface did not function in fusion or viral entry. The screening of gH mutants for function revealed the following: (i) for wild-type gH and some gH mutants, fusion with nectin-1-expressing target cells occurred more rapidly than with herpesvirus entry mediator (HVEM)-expressing target cells; (ii) some gH mutants reduced the rate of cell fusion without abrogating fusion completely, indicating that gH may play a role in governing the kinetics of fusion and may be responsible for a rate-limiting first stage in HSV-1 fusion; and (iii) only one gH mutant, located within the short cytoplasmic tail, completely abrogated function, indicating that the gH cytoplasmic tail is crucial for cell fusion and viral infectivity.

scholarly journals Engineered Disulfide Bonds in Herpes Simplex Virus Type 1 gD Separate Receptor Binding from Fusion Initiation and Viral Entry

2007 ◽  
Vol 82 (2) ◽  
pp. 700-709 ◽  
Author(s):  
Eric Lazear ◽  
Andrea Carfi ◽  
J. Charles Whitbeck ◽  
Tina M. Cairns ◽  
Claude Krummenacher ◽  
...  
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Cell Fusion ◽  
Receptor Binding ◽  
Disulfide Bond ◽  
Viral Entry ◽  
Disulfide Bonds ◽  
C Terminus ◽  
Simplex Virus ◽  
Cell Cell

ABSTRACT Glycoprotein D (gD) is the receptor binding protein of herpes simplex virus (HSV) and binds to at least two distinct protein receptors, herpesvirus entry mediator (HVEM) and nectin-1. While both receptor binding regions are found within the first 234 amino acids, a crystal structure shows that the C terminus of the gD ectodomain normally occludes the receptor binding sites. Receptor binding must therefore displace the C terminus, and this conformational change is postulated to be required for inducing fusion via gB and gH/gL. When cysteine residues are introduced at positions 37 and 302 of gD, a disulfide bond is formed that stabilizes the C terminus and prevents binding to either receptor. We speculated that if disulfide bonds were engineered further upstream, receptor binding might be separated from the induction of fusion. To test this, we made five additional double cysteine mutants, each potentially introducing a disulfide bond between the ectodomain C terminus and the core of the gD ectodomain. The two mutants predicted to impose the greatest constraint were unable to bind receptors or mediate cell-cell fusion. However, the three mutants with the most flexible C terminus bound well to both HVEM and nectin-1. Two of these mutants were impaired in cell-cell fusion and null-virus complementation. Importantly, a third mutant in this group was nonfunctional in both assays. This mutant clearly separates the role of gD in triggering fusion from its role in receptor binding. Based upon the properties of the panel of mutants we conclude that fusion requires greater flexibility of the gD ectodomain C terminus than does receptor binding.

scholarly journals Regulation of Herpes Simplex Virus Glycoprotein-Induced Cascade of Events Governing Cell-Cell Fusion

2016 ◽  
Vol 90 (23) ◽  
pp. 10535-10544 ◽  
Author(s):  
Doina Atanasiu ◽  
Wan Ting Saw ◽  
Roselyn J. Eisenberg ◽  
Gary H. Cohen
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Cell Fusion ◽  
Critical Role ◽  
Fusion Process ◽  
Simplex Virus ◽  
The Impact ◽  
Hsv 1 ◽  
Cell Cell

ABSTRACTReceptor-dependent herpes simplex virus (HSV)-induced cell-cell fusion requires glycoproteins gD, gH/gL, and gB. Our current model posits that during fusion, receptor-activated conformational changes in gD activate gH/gL, which subsequently triggers the transformation of the prefusion form of gB into a fusogenic state. To examine the role of each glycoprotein in receptor-dependent cell-cell fusion, we took advantage of our discovery that fusion by wild-type herpes simplex virus 2 (HSV-2) glycoproteins occurs twice as fast as that achieved by HSV-1 glycoproteins. By sequentially swapping each glycoprotein between the two serotypes, we established that fusion speed was governed by gH/gL, with gH being the main contributor. While the mutant forms of gB fuse at distinct rates that are dictated by their molecular structure, these restrictions can be overcome by gH/gL of HSV-2 (gH2/gL2), thereby enhancing their activity. We also found that deregulated forms of gD of HSV-1 (gD1) and gH2/gL2can alter the fusogenic potential of gB, promoting cell fusion in the absence of a cellular receptor, and that deregulated forms of gB can drive the fusion machinery to even higher levels. Low pH enhanced fusion by affecting the structure of both gB and gH/gL mutants. Together, our data highlight the complexity of the fusion machinery, the impact of the activation state of each glycoprotein on the fusion process, and the critical role of gH/gL in regulating HSV-induced fusion.IMPORTANCECell-cell fusion mediated by HSV glycoproteins requires gD, gH/gL, gB, and a gD receptor. Here, we show that fusion by wild-type HSV-2 glycoproteins occurs twice as fast as that achieved by HSV-1 glycoproteins. By sequentially swapping each glycoprotein between the two serotypes, we found that the fusion process was controlled by gH/gL. Restrictions imposed on the gB structure by mutations could be overcome by gH2/gL2, enhancing the activity of the mutants. Under low-pH conditions or when using deregulated forms of gD1and gH2/gL2, the fusogenic potential of gB could only be increased in the absence of receptor, underlining the exquisite regulation that occurs in the presence of receptor. Our data highlight the complexity of the fusion machinery, the impact of the activation state of each glycoprotein on the fusion process, and the critical role of gH/gL in regulating HSV-induced fusion.

scholarly journals Differential Effects on Cell Fusion Activity of Mutations in Herpes Simplex Virus 1 Glycoprotein B (gB) Dependent on Whether a gD Receptor or a gB Receptor Is Overexpressed

2009 ◽  
Vol 83 (15) ◽  
pp. 7384-7390 ◽  
Author(s):  
Qing Fan ◽  
Erick Lin ◽  
Takeshi Satoh ◽  
Hisashi Arase ◽  
Patricia G. Spear
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Cell Fusion ◽  
Viral Entry ◽  
Selective Reduction ◽  
Glycoprotein B ◽  
Fusion Activity ◽  
Differential Effects ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACT Glycoprotein B (gB) of herpes simplex virus (HSV) is one of four glycoproteins essential for viral entry and cell fusion. Recently, paired immunoglobulin-like type 2 receptor (PILRα) was identified as a receptor for HSV type 1 (HSV-1) gB. Both PILRα and a gD receptor were shown to participate in HSV-1 entry into certain cell types. The purpose of this study was to determine whether insertional mutations in gB had differential effects on its function with PILRα and the gD receptor, nectin-1. Previously described gB mutants and additional newly characterized mutants were used in this study. We found that insertional mutations near the N terminus and C terminus of gB and especially in the central region of the ectodomain reduced cell fusion activity when PILRα was overexpressed much more than when nectin-1 was overexpressed. Most of the insertions reduced the binding of gB to PILRα, for at least some forms of gB, but this reduction did not necessarily correlate with the selective reduction in cell fusion activity with PILRα. These results suggest that the regions targeted by the relevant mutations are critical for functional activity with PILRα. They also suggest that, although both the binding of gB to a gB receptor and the binding of gD to a gD receptor may be required for HSV-induced cell fusion, the two receptor-binding activities may have unequal weights in triggering fusogenic activity, depending on the ratios of gB and gD receptors or other factors.

scholarly journals Contribution of Endocytic Motifs in the Cytoplasmic Tail of Herpes Simplex Virus Type 1 Glycoprotein B to Virus Replication and Cell-Cell Fusion

2007 ◽  
Vol 81 (24) ◽  
pp. 13889-13903 ◽  
Author(s):  
Igor Beitia Ortiz de Zarate ◽  
Lilia Cantero-Aguilar ◽  
Magalie Longo ◽  
Clarisse Berlioz-Torrent ◽  
Flore Rozenberg
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Cell Surface ◽  
Cell Fusion ◽  
Virus Replication ◽  
Herpes Simplex Virus Type ◽  
Wild Type ◽  
Simplex Virus ◽  
Cell Cell

ABSTRACT The use of endocytic pathways by viral glycoproteins is thought to play various functions during viral infection. We previously showed in transfection assays that herpes simplex virus type 1 (HSV-1) glycoprotein B (gB) is transported from the cell surface back to the trans-Golgi network (TGN) and that two motifs of gB cytoplasmic tail, YTQV and LL, function distinctly in this process. To investigate the role of each of these gB trafficking signals in HSV-1 infection, we constructed recombinant viruses in which each motif was rendered nonfunctional by alanine mutagenesis. In infected cells, wild-type gB was internalized from the cell surface and concentrated in the TGN. Disruption of YTQV abolished internalization of gB during infection, whereas disruption of LL induced accumulation of internalized gB in early recycling endosomes and impaired its return to the TGN. The growth of both recombinants was moderately diminished. Moreover, the fusion phenotype of cells infected with the gB recombinants differed from that of cells infected with the wild-type virus. Cells infected with the YTQV-mutated virus displayed reduced cell-cell fusion, whereas giant syncytia were observed in cells infected with the LL-mutated virus. Furthermore, blocking gB internalization or impairing gB recycling to the cell surface, using drugs or a transdominant negative form of Rab11, significantly reduced cell-cell fusion. These results favor a role for endocytosis in virus replication and suggest that gB intracellular trafficking is involved in the regulation of cell-cell fusion.

scholarly journals Herpes Simplex Virus 1 Latency and the Kinetics of Reactivation Are Regulated by a Complex Network of Interactions between the Herpesvirus Entry Mediator, Its Ligands (gD, BTLA, LIGHT, and CD160), and the Latency-Associated Transcript

2018 ◽  
Vol 92 (24) ◽  
Author(s):  
Shaohui Wang ◽  
Alexander V. Ljubimov ◽  
Ling Jin ◽  
Klaus Pfeffer ◽  
Mitchell Kronenberg ◽  
...  
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Trigeminal Ganglia ◽  
Wild Type ◽  
Herpes Simplex Virus 1 ◽  
Latently Infected ◽  
Simplex Virus ◽  
Kinetics Of ◽  
Herpesvirus Entry Mediator ◽  
Hsv 1

ABSTRACTRecently, we reported that the herpesvirus entry mediator (HVEM; also called TNFRSF14 or CD270) is upregulated by the latency-associated transcript (LAT) of herpes simplex virus 1 (HSV-1) and that the absence of HVEM affects latency reactivation but not primary infection in ocularly infected mice. gD has been shown to bind to HVEM. LIGHT (TNFSF14), CD160, and BTLA (B- and T-lymphocyte attenuator) also interact with HVEM and can interfere with HSV gD binding. It was not known if LIGHT, CD160, or BTLA affected the level of latency reactivation in the trigeminal ganglia (TG) of latently infected mice. To address this issue, we ocularly infected LIGHT−/−, CD160−/−, and BTLA−/−mice with LAT(+) and LAT(−) viruses, using similarly infected wild-type (WT) and HVEM−/−mice as controls. The amount of latency, as determined by the levels of gB DNA in the TG of the LIGHT−/−, CD160−/−, and BTLA−/−mice infected with either LAT(+) or LAT(−) viruses, was lower than that in WT mice infected with LAT(+) virus and was similar in WT mice infected with LAT(−) virus. The levels of LAT RNA in HVEM−/−, LIGHT−/−, CD160−/−, and BTLA−/−mice infected with LAT(+) virus were similar and were lower than the levels of LAT RNA in WT mice. However, LIGHT−/−, CD160−/−, and BTLA−/−mice, independent of the presence of LAT, had levels of reactivation similar to those of WT mice infected with LAT(+) virus. Faster reactivation correlated with the upregulation of HVEM transcript. The LIGHT−/−, CD160−/−, and BTLA−/−mice had higher levels of HVEM expression, and this, along with the absence of BTLA, LIGHT, or CD160, may contribute to faster reactivation, while the absence of each molecule, independent of LAT, may have contributed to lower latency. This study suggests that, in the absence of competition with gD for binding to HVEM, LAT RNA is important for WT levels of latency but not for WT levels of reactivation.IMPORTANCEThe effects of BTLA, LIGHT, and CD160 on latency reactivation are not known. We show here that in BTLA, LIGHT, or CD160 null mice, latency is reduced; however, HVEM expression is upregulated compared to that of WT mice, and this upregulation is associated with higher reactivation that is independent of LAT but dependent on gD expression. Thus, one of the mechanisms by which BTLA, LIGHT, and CD160 null mice enhance reactivation appears to be the increased expression of HVEM in the presence of gD. Thus, our results suggest that blockade of HVEM-LIGHT-BTLA-CD160 contributes to reduced HSV-1 latency and reactivation.

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