scholarly journals Cleavage of Epstein–Barr virus glycoprotein B is required for full function in cell–cell fusion with both epithelial and B cells

2009 ◽  
Vol 90 (3) ◽  
pp. 591-595 ◽  
Author(s):  
Jessica Sorem ◽  
Richard Longnecker
Keyword(s):  
B Cells ◽  
Cell Fusion ◽  
Epstein Barr Virus ◽  
Glycoprotein B ◽  
Barr Virus ◽  
High Sequence Conservation ◽  
A Cell ◽  
Epstein Barr ◽  
Cleavage Motif ◽  
Cell Cell

Glycoprotein B (gB) homologues within the herpesvirus family display high sequence conservation, and a number of gB homologues contain a cleavage motif R-X-K/R-R recognized by the cellular protease furin. Epstein–Barr virus (EBV) gB contains this motif and cleaved gB is found in EBV virions. To determine the functional significance of this cleavage motif in EBV gB, a deletion mutant (gB Δfurin) was created lacking the motif. This cleavage mutant was expressed well in cell culture but was not cleaved. Experiments examining gB Δfurin in a cell-fusion assay revealed that fusion was reduced by 52 % in epithelial and 28 % in B cells when compared with wild-type EBV gB. This decrease in cell–cell fusion is similar to that observed with multiple alphaherpesvirus gB cleavage mutants and supports a conserved function for cleaved gB.

scholarly journals Compatibility of the gH homologues of Epstein–Barr virus and related lymphocryptoviruses

2007 ◽  
Vol 88 (8) ◽  
pp. 2129-2136 ◽  
Author(s):  
Liguo Wu ◽  
Lindsey M. Hutt-Fletcher
Keyword(s):  
B Cells ◽  
Epithelial Cells ◽  
B Cell ◽  
Cell Fusion ◽  
Epstein Barr Virus ◽  
Common Marmoset ◽  
Barr Virus ◽  
Human B Cells ◽  
The Common ◽  
Epstein Barr

Glycoprotein gH, together with its chaperone gL and a third glycoprotein gB, is essential for cell–cell fusion and virus–cell fusion mediated by herpesviruses. Epstein–Barr virus (EBV), the prototype human lymphocryptovirus, requires a fourth glycoprotein gp42 to support fusion with B cells in addition to epithelial cells. Two other lymphocryptoviruses, the rhesus lymphocryptovirus (Rh-LCV) and the common marmoset lymphocryptovirus (CalHV3), have been sequenced in their entirety and each has a gp42 homologue. Combinations of proteins from EBV, Rh-LCV and CalHV3 were able to mediate fusion of epithelial cells, but, even when complexed with EBV gp42, only Rh-LCV and not CalHV3 proteins were able to mediate fusion with human B cells. CalHV3 gL was also unable to function effectively as a chaperone for EBV or Rh-LCV gH. The Rh-LCV gH homologue supported more fusion than EBV gH with an epithelial cell and supported the highest levels of fusion with a B cell. Chimeric constructs made from Rh-LCV gH and EBV gH that have 85.4 % sequence identity should prove useful for mapping the regions of gH that are of importance to fusion as a whole and to B-cell fusion in particular.

scholarly journals Analysis of Epstein-Barr Virus Glycoprotein B Functional Domains via Linker Insertion Mutagenesis

2008 ◽  
Vol 83 (2) ◽  
pp. 734-747 ◽  
Author(s):  
Jessica J. Reimer ◽  
Marija Backovic ◽  
Charuhas G. Deshpande ◽  
Theodore Jardetzky ◽  
Richard Longnecker
Keyword(s):  
B Cells ◽  
Epstein Barr Virus ◽  
Glycoprotein B ◽  
Functional Domains ◽  
Insertion Mutagenesis ◽  
Fusion Activity ◽  
Wild Type ◽  
Protein Fusion ◽  
Barr Virus ◽  
Epstein Barr

ABSTRACT Epstein-Barr Virus (EBV) glycoprotein B (gB) is essential for viral fusion events with epithelial and B cells. This glycoprotein has been studied extensively in other herpesvirus family members, but functional domains outside of the cytoplasmic tail have not been characterized in EBV gB. In this study, a total of 28 linker insertion mutations were generated throughout the length of gB. In general, the linker insertions did not disrupt intracellular expression and variably altered cell surface expression. Oligomerization was disrupted by insertions located between residues 561 and 620, indicating the location of a potential site of oligomer contacts between EBV gB monomers. In addition, a novel N-glycosylated form of wild-type gB was identified under nonreducing Western blot conditions that likely represents a mature form of the protein. Fusion activity was abolished in all but three variants containing mutations in the N-terminal region (gB30), within the ectodomain (gB421), and in the intracellular C-terminal domain (gB832) of the protein. Fusion activity with variants gB421 and gB832 was comparable to that of the wild type with epithelial and B cells, and only these two mutants, but not gB30, were able to complement gB-null virus and subsequently function in virus entry. The mutant gB30 exhibited a low level of fusion activity with B cells and was unable to complement gB-null virus. The mutations generated here indicate important structural domains, as well as regions important for function in fusion, within EBV gB.

scholarly journals Soluble Epstein-Barr Virus Glycoproteins gH, gL, and gp42 Form a 1:1:1 Stable Complex That Acts Like Soluble gp42 in B-Cell Fusion but Not in Epithelial Cell Fusion

2006 ◽  
Vol 80 (19) ◽  
pp. 9444-9454 ◽  
Author(s):  
Austin N. Kirschner ◽  
Jasmina Omerović ◽  
Boris Popov ◽  
Richard Longnecker ◽  
Theodore S. Jardetzky
Keyword(s):  
B Cells ◽  
Epithelial Cell ◽  
Epithelial Cells ◽  
B Cell ◽  
Membrane Fusion ◽  
Cell Fusion ◽  
Epstein Barr Virus ◽  
Stable Complex ◽  
Barr Virus ◽  
Epstein Barr

ABSTRACT Epstein-Barr virus (EBV) is a herpesvirus that infects cells by fusing its lipid envelope with the target cell membrane. The fusion process requires the actions of viral glycoproteins gH, gL, and gB for entry into epithelial cells and additionally requires gp42 for entry into B cells. To further study the roles of these membrane-associated glycoproteins, purified soluble forms of gp42, gH, and gL were expressed that lack the membrane-spanning regions. The soluble gH/gL protein complex binds to soluble gp42 with high affinity, forming a stable heterotrimer with 1:1:1 stoichiometry, and this complex is not formed by an N-terminally truncated variant of gp42. The effects of adding soluble gp42, gH/gL, and gH/gL/gp42 were examined with a virus-free cell-cell fusion assay. The results demonstrate that, in contrast to gp42, membrane fusion does not proceed with secreted gH/gL. The addition of soluble gH/gL does not inhibit or enhance B-cell or epithelial cell fusion when membrane-bound gH/gL, gB, and gp42 are present. However, the soluble gH/gL/gp42 complex does activate membrane fusion with B cells, similarly to soluble gp42, but it does not inhibit fusion with epithelial cells, as observed for gp42 alone. A gp42 peptide, derived from an N-terminal segment involved in gH/gL interactions, binds to soluble gH/gL and inhibits EBV-mediated epithelial cell fusion, mimicking gp42. These observations reveal distinct functional requirements for gH/gL and gp42 complexes in EBV-mediated membrane fusion.

scholarly journals Mutations of Epstein-Barr Virus gH That Are Differentially Able To Support Fusion with B Cells or Epithelial Cells

2005 ◽  
Vol 79 (17) ◽  
pp. 10923-10930 ◽  
Author(s):  
Liguo Wu ◽  
Corina M. Borza ◽  
Lindsey M. Hutt-Fletcher
Keyword(s):  
B Cells ◽  
Epithelial Cell ◽  
Epithelial Cells ◽  
B Cell ◽  
Cell Fusion ◽  
Epstein Barr Virus ◽  
Common Mechanism ◽  
Barr Virus ◽  
The Core ◽  
Epstein Barr

ABSTRACT The core fusion machinery of all herpesviruses consists of three conserved glycoproteins, gB and gHgL, suggesting a common mechanism for virus cell fusion, but fusion of Epstein-Barr virus (EBV) with B cells and epithelial cells is initiated differently. Fusion with B cells requires a fourth protein, gp42, which complexes with gHgL and interacts with HLA class II, the B-cell coreceptor. Fusion with an epithelial cell does not require gp42 but requires interaction of gHgL with a novel epithelial cell coreceptor. Epithelial cell fusion can be inhibited by gp42 binding to gHgL and by antibodies to gH that fail to block B-cell fusion. This suggests that regions of gHgL initiating fusion with each cell are separable from each other and from regions involved in fusion itself. To address this possibility we mapped the region of gH recognized by a monoclonal antibody to gH that blocks EBV fusion with epithelial cells but not B cells by making a series of chimeras with the gH homolog of rhesus lymphocryptovirus. Proteins with mutations engineered within this region included those that preferentially mediate fusion with B cells, those that preferentially mediate fusion with epithelial cells, and those that mediate fusion with neither cell type. These results support the hypothesis that the core fusion function of gH is the same for B cells and epithelial cells and that it differs only in the way in which it is triggered into a functionally active state.

scholarly journals Membrane Anchoring of Epstein-Barr Virus gp42 Inhibits Fusion with B Cells Even with Increased Flexibility Allowed by Engineered Spacers

mBio ◽  
2015 ◽  
Vol 6 (1) ◽  
Author(s):  
Cynthia L. Rowe ◽  
Jia Chen ◽  
Theodore S. Jardetzky ◽  
Richard Longnecker
Keyword(s):  
B Cells ◽  
B Cell ◽  
Cell Fusion ◽  
Steric Hindrance ◽  
Epstein Barr Virus ◽  
Class Ii ◽  
Hla Class Ii ◽  
Barr Virus ◽  
Membrane Bound ◽  
Epstein Barr

ABSTRACTWe recently described the architecture of the Epstein-Barr virus (EBV) fusion-triggering complex consisting of the EBV B cell receptor human leukocyte antigen (HLA) class II and the EBV-encoded proteins gp42 and gH/gL. The architecture of this structure positioned the main body of gp42, comprising the C-type lectin domain (CTLD), away from the membrane and distant from where the membrane-bound form of gp42 might be tethered. gp42 is a type II membrane glycoprotein, with functional gp42 formed by cleavage near the gp42 amino-terminal transmembrane domain. This cleavage results in an approximately 50-amino-acid unstructured region that is responsible for binding gH/gL with nanomolar affinity. Our previous studies had shown that membrane-bound gp42 is not functional in B cell fusion. To investigate whether we could restore gp42 function by extending it from the membrane, we introduced one, two, and four structured immunoglobulin-like domains from muscle protein titin into a membrane-bound form of gp42 and tested function in binding to gHgL and HLA class II and function in fusion. We hypothesized that cleavage of gp42 generates a soluble functional form that relieves steric hindrance imposed on gHgL by membrane-bound gp42. All of the linker mutants had a dominant-negative effect on gp42 function, indicating that gp42 fusion function could not be restored simply by the addition of one to four titin domains.IMPORTANCEEpstein-Barr virus (EBV) is associated with numerous diseases from benign mononucleosis to Burkitt’s and Hodgkin’s lymphoma, nasopharyngeal and gastric carcinoma, and lymphoproliferative disorders in patients with immune dysfunction resulting from immune suppression. Among the glycoproteins important for fusion, gp42, along with gH/gL, determines EBV tropism between epithelial and B cells. The function of gp42 is dependent on N-terminal cleavage, since membrane-bound gp42 cannot mediate fusion. We further investigated whether insertion of a linker into membrane-bound gp42 would relieve steric hindrance imposed on membrane-bound gp42 and restore fusion function. However, adding one, two, or four structured immunoglobulin-like domains to membrane gp42 did not restore fusion activity, indicating that the architecture and membrane orientation of the B cell fusion-triggering complex of EBV may be easily perturbed and that gp42 cleavage is essential for B cell fusion.

scholarly journals Recurrent Erythema Annulare Centrifugum due to Influenza Type A

2021 ◽  
pp. 134-140
Author(s):  
Luca Ena ◽  
Vittorio Mazzarello ◽  
Marco Ferrari ◽  
Pasquale Ena
Keyword(s):  
Epstein Barr Virus ◽  
Local Treatment ◽  
Type A ◽  
Barr Virus ◽  
Ebv Infection ◽  
Influenza Type ◽  
Erythema Annulare Centrifugum ◽  
A Cell ◽  
Epstein Barr ◽  
Calcipotriol Betamethasone

Erythema annulare centrifugum (EAC) is a rare erythema characterized by erythematous and urticarial papules or annular plaques that enlarges centrifugally. The lesions usually involve the thighs and the legs. Several disorders are occasionally associated with EAC, infections, including mycoses, bacteria, or viruses and drugs have also been regarded as possible causes of this eruption. We present a 42-year-old dark-skinned woman affected by recurrent EAC that appeared secondary to influenza type A (H1N1). Histopathology showed a superficial form of EAC. In our case, a previous cytomegalovirus and Epstein-Barr virus (EBV) infection were identified and no underlying other diseases were found. Clarithromycin with calcipotriol betamethasone treatment was temporarily efficacious. In the last 3 years, the lesions started to appear every 2 weeks and tended to regress with local treatment after a variable period. We believe that the latent cytomegalovirus and the reactivity induced by EBV combined with influenza can determine, in our case, a cell mediate cutaneous immune response, which leads to the peculiar inflammatory disease known as EAC.

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