A potent and protective human neutralizing antibody targeting a novel vulnerable site of Epstein-Barr virus

AbstractEpstein-Barr virus (EBV) is associated with a range of epithelial and B cell malignancies as well as autoimmune disorders, for which there are still no specific treatments or effective vaccines. Here, we isolate EBV gH/gL-specific antibodies from an EBV-infected individual. One antibody, 1D8, efficiently neutralizes EBV infection of two major target cell types, B cells and epithelial cells. In humanized mice, 1D8 provides protection against a high-dose EBV challenge by substantially reducing viral loads and associated tumor burden. Crystal structure analysis reveals that 1D8 binds to a key vulnerable interface between the D-I/D-II domains of the viral gH/gL protein, especially the D-II of the gH, thereby interfering with the gH/gL-mediated membrane fusion and binding to target cells. Overall, we identify a potent and protective neutralizing antibody capable of reducing the EBV load. The novel vulnerable site represents an attractive target that is potentially important for antibody and vaccine intervention against EBV infection.

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A potent and protective human neutralizing antibody targeting a key vulnerable site of Epstein-Barr Virus

10.21203/rs.3.rs-151895/v1 ◽

2021 ◽

Author(s):

Qian-Ying Zhu ◽

Sisi Shan ◽

Jinfang Yu ◽

Si-Ying Peng ◽

Cong Sun ◽

...

Keyword(s):

Epstein Barr Virus ◽

Infected Individual ◽

Neutralizing Antibody ◽

Crystal Structure Analysis ◽

Therapeutic Interventions ◽

Target Cells ◽

High Dose ◽

Barr Virus ◽

Ebv Infection ◽

Epstein Barr

Abstract Epstein-Barr virus (EBV) is associated with a range of epithelial and B cell malignancies as well as autoimmune disorders, for which there are still no specific treatments or effective vaccines. Here, we isolated EBV gH/gL-specific antibodies from an EBV-infected individual. One antibody, 1D8, efficiently neutralized EBV infection of two major target cell types, B cells and epithelial cells. In humanized mice, 1D8 provided strong protection against a high-dose EBV challenge by substantially reducing viral loads and associated tumor burden. Crystal structure analysis revealed that 1D8 binds to a key vulnerable interface between the D-I/D-II domains of the viral gH/gL protein, especially the D-II of the gH, thereby interfering with the gH/gL-mediated membrane fusion and binding to target cells. Overall, we identified a potent neutralizing antibody as a promising candidate for prophylactic and therapeutic interventions against EBV infection. The key vulnerable site also provides insights into the EBV vaccines design.

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The BRRF1 Early Gene of Epstein-Barr Virus Encodes a Transcription Factor That Enhances Induction of Lytic Infection by BRLF1

Journal of Virology ◽

10.1128/jvi.78.10.4983-4992.2004 ◽

2004 ◽

Vol 78 (10) ◽

pp. 4983-4992 ◽

Cited By ~ 41

Author(s):

Gregory K. Hong ◽

Henri-Jacques Delecluse ◽

Henri Gruffat ◽

Thomas E. Morrison ◽

Wen-Hai Feng ◽

...

Keyword(s):

Transcription Factor ◽

Epstein Barr Virus ◽

Cell Types ◽

Early Gene ◽

Barr Virus ◽

Ebv Infection ◽

Early Protein ◽

293 Cells ◽

Latently Infected ◽

Epstein Barr

ABSTRACT The switch from the latent to the lytic form of Epstein-Barr virus (EBV) infection is mediated by expression of the viral immediate-early (IE) proteins, BZLF1 (Z) and BRLF1 (R). An EBV early protein, BRRF1 (Na), is encoded by the opposite strand of the BRLF1 intron, but the function of this nuclear protein in the viral life cycle is unknown. Here we demonstrate that Na enhances the R-mediated induction of lytic EBV infection in 293 cells latently infected with a recombinant EBV (R-KO) defective for the expression of both R and Na. Na also enhances R-induced lytic infections in a gastric carcinoma line (AGS) carrying the R-KO virus, although it has no effect in a Burkitt lymphoma line (BL-30) stably infected with the same mutant virus. We show that Na is a transcription factor that increases the ability of R to activate Z expression from the R-KO viral genome in 293 cells and that Na by itself activates the Z promoter (Zp) in EBV-negative cells. Na activation of Zp requires a CRE motif (ZII), and a consensus CRE motif is sufficient to transfer Na responsiveness to the heterologous E1b promoter. Furthermore, we show that Na enhances the transactivator function of a Gal4-c-Jun fusion protein but does not increase the transactivator function of other transcription factors (including ATF-1, ATF-2, and CREB) known to bind CRE motifs. Na expression in cells results in increased levels of a hyperphosphorylated form of c-Jun, suggesting a mechanism by which Na activates c-Jun. Our results indicate that Na is a transcription factor that activates the EBV Zp IE promoter through its effects on c-Jun and suggest that Na cooperates with BRLF1 to induce the lytic form of EBV infection in certain cell types.

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Impaired Epstein-Barr virus–specific CD8+ T-cell function in X-linked lymphoproliferative disease is restricted to SLAM family–positive B-cell targets

Blood ◽

10.1182/blood-2009-09-238832 ◽

2010 ◽

Vol 116 (17) ◽

pp. 3249-3257 ◽

Cited By ~ 79

Author(s):

Andrew D. Hislop ◽

Umaimainthan Palendira ◽

Alison M. Leese ◽

Peter D. Arkwright ◽

Pierre S. Rohrlich ◽

...

Keyword(s):

T Cell ◽

B Cell ◽

Epstein Barr Virus ◽

Cd8 T Cell ◽

Lymphoproliferative Disease ◽

Target Cells ◽

Barr Virus ◽

Ebv Infection ◽

Slam Family ◽

Epstein Barr

Abstract X-linked lymphoproliferative disease (XLP) is a condition associated with mutations in the signaling lymphocytic activation molecule (SLAM)–associated protein (SAP; SH2D1A). SAP functions as an adaptor, binding to and recruiting signaling molecules to SLAM family receptors expressed on T and natural killer cells. XLP is associated with extreme sensitivity to primary Epstein-Barr virus (EBV) infection, often leading to a lethal infectious mononucleosis. To investigate EBV-specific immunity in XLP patients, we studied 5 individuals who had survived EBV infection and found CD8+ T-cell responses numerically comparable with healthy donors. However, further investigation of in vitro–derived CD8+ T-cell clones established from 2 of these donors showed they efficiently recognized SLAM ligand–negative target cells expressing EBV antigens, but showed impaired recognition of EBV-transformed, SLAM ligand–positive, lymphoblastoid cell lines (LCLs). Importantly, LCL recognition was restored when interactions between the SLAM receptors CD244 and natural killer–, T-, and B-cell antigen (NTBA) and their ligands on LCLs were blocked. We propose that XLP patients' particular sensitivity to EBV, and not to other viruses, reflects at least in part EBV's strict tropism for B lymphocytes and the often inability of the CD8+ T-cell response to contain the primary infection of SLAM ligand–expressing target cells.

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Kinetics of Epstein-Barr Virus (EBV) Neutralizing and Virus-Specific Antibodies after Primary Infection with EBV

Clinical and Vaccine Immunology ◽

10.1128/cvi.00674-15 ◽

2016 ◽

Vol 23 (4) ◽

pp. 363-369 ◽

Cited By ~ 18

Author(s):

Wei Bu ◽

Gregory M. Hayes ◽

Hui Liu ◽

Lorraine Gemmell ◽

David O. Schmeling ◽

...

Keyword(s):

Infectious Mononucleosis ◽

Neutralizing Antibodies ◽

Primary Infection ◽

Epstein Barr Virus ◽

Neutralizing Antibody ◽

Enzyme Linked Immunosorbent Assay ◽

Barr Virus ◽

Ebv Infection ◽

Epstein Barr ◽

Kinetics Of

ABSTRACTProspective studies of antibodies to multiple Epstein-Barr virus (EBV) proteins and EBV neutralizing antibodies in the same individuals before, during, and after primary EBV infection have not been reported. We studied antibody responses to EBV in college students who acquired primary EBV infection during prospective surveillance and correlated the kinetics of antibody response with the severity of disease. Neutralizing antibodies and enzyme-linked immunosorbent assay (ELISA) antibodies to gp350, the major target of neutralizing antibody, reached peak levels at medians of 179 and 333 days after the onset of symptoms of infectious mononucleosis, respectively. No clear correlation was found between the severity of the symptoms of infectious mononucleosis and the peak levels of antibody to individual viral proteins or to neutralizing antibody. In summary, we found that titers of neutralizing antibody and antibodies to multiple EBV proteins increase over many months after primary infection with EBV.

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A primary nasopharyngeal three-dimensional air-liquid interface cell culture model of the pseudostratified epithelium reveals differential donor- and cell type-specific susceptibility to Epstein-Barr virus infection

PLoS Pathogens ◽

10.1371/journal.ppat.1009041 ◽

2021 ◽

Vol 17 (4) ◽

pp. e1009041

Author(s):

Phillip Ziegler ◽

Yarong Tian ◽

Yulong Bai ◽

Sanna Abrahamsson ◽

Alan Bäckerholm ◽

...

Keyword(s):

Differentially Expressed Genes ◽

Epstein Barr Virus ◽

Cell Types ◽

Lytic Infection ◽

Cell Type ◽

Barr Virus ◽

Ebv Infection ◽

Pseudostratified Epithelium ◽

Epstein Barr ◽

Air Liquid Interface

Epstein-Barr virus (EBV) is a ubiquitous γ-herpesvirus with latent and lytic cycles. EBV replicates in the stratified epithelium but the nasopharynx is also composed of pseudostratified epithelium with distinct cell types. Latent infection is associated with nasopharyngeal carcinoma (NPC). Here, we show with nasopharyngeal conditionally reprogrammed cells cultured at the air-liquid interface that pseudostratified epithelial cells are susceptible to EBV infection. Donors varied in susceptibility to de novo EBV infection, but susceptible cultures also displayed differences with respect to pathogenesis. The cultures from one donor yielded lytic infection but cells from two other donors were positive for EBV-encoded EBERs and negative for other lytic infection markers. All cultures stained positive for the pseudostratified markers CK7, MUC5AC, α-tubulin in cilia, and the EBV epithelial cell receptor Ephrin receptor A2. To define EBV transcriptional programs by cell type and to elucidate latent/lytic infection-differential changes, we performed single cell RNA-sequencing on one EBV-infected culture that resulted in alignment with many EBV transcripts. EBV transcripts represented a small portion of the total transcriptome (~0.17%). All cell types in the pseudostratified epithelium had detectable EBV transcripts with suprabasal cells showing the highest number of reads aligning to many EBV genes. Several restriction factors (IRF1, MX1, STAT1, C18orf25) known to limit lytic infection were expressed at lower levels in the lytic subcluster. A third of the differentially-expressed genes in NPC tumors compared to an uninfected pseudostratified ALI culture overlapped with the differentially-expressed genes in the latent subcluster. A third of these commonly perturbed genes were specific to EBV infection and changed in the same direction. Collectively, these findings suggest that the pseudostratified epithelium could harbor EBV infection and that the pseudostratified infection model mirrors many of the transcriptional changes imposed by EBV infection in NPC.

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Epstein-Barr Virus and Systemic Autoimmune Diseases

Frontiers in Immunology ◽

10.3389/fimmu.2020.587380 ◽

2021 ◽

Vol 11 ◽

Author(s):

Gunnar Houen ◽

Nicole Hartwig Trier

Keyword(s):

B Cells ◽

Epithelial Cells ◽

Autoimmune Diseases ◽

Epstein Barr Virus ◽

Cell Types ◽

Human Beings ◽

Systemic Autoimmune Diseases ◽

Barr Virus ◽

Ebv Infection ◽

Epstein Barr

Epstein-Barr Virus (EBV) is an extremely successful human herpes virus, which infects essentially all human beings at some time during their life span. EBV infection and the associated immune response results in production of antibodies (seroconversion), which occurs mainly during the first years of life, but may also happen during adolescence or later in life. Infection of adolescents can result in infectious mononucleosis, an acute serious condition characterized by massive lymphocytosis. Transmission of EBV mainly occurs through saliva but can rarely be spread through semen or blood, e.g. through organ transplantations and blood transfusions. EBV transmission through oral secretions results in infection of epithelial cells of the oropharynx. From the epithelial cells EBV can infect B cells, which are the major reservoir for the virus, but other cell types may also become infected. As a result, EBV can shuttle between different cell types, mainly B cells and epithelial cells. Moreover, since the virus can switch between a latent and a lytic life cycle, EBV has the ability to cause chronic relapsing/reactivating infections. Chronic or recurrent EBV infection of epithelial cells has been linked to systemic lupus erythematosus and Sjögren’s syndrome, whereas chronic/recurrent infection of B cells has been associated with rheumatoid arthritis, multiple sclerosis and other diseases. Accordingly, since EBV can shuttle between epithelial cells and B cells, the systemic autoimmune diseases often occur as overlapping syndromes with symptoms and characteristic autoantibodies (e.g. antinuclear antibodies and rheumatoid factors) reflecting epithelial and/or B cell infection.

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A three-dimensional Air-Liquid Interface Culture Model for the Study of Epstein-Barr virus Infection in the Nasopharynx

10.1101/2020.08.31.272096 ◽

2020 ◽

Cited By ~ 1

Author(s):

Phillip Ziegler ◽

Yulong Bai ◽

Yarong Tian ◽

Sanna Abrahamsson ◽

Anthony Green ◽

...

Keyword(s):

Epstein Barr Virus ◽

Three Dimensional ◽

Cell Types ◽

Liquid Interface ◽

Epstein Barr Virus Infection ◽

Barr Virus ◽

Ebv Infection ◽

Transcription Profiles ◽

Epstein Barr ◽

Air Liquid Interface

ABSTRACTEpstein-Barr virus (EBV) infection is ubiquitous in humans and is associated with the cancer, nasopharyngeal carcinoma. EBV replicates in the differentiated layers of stratified keratinocytes but whether the other cell types of the airway epithelium are susceptible to EBV is unknown. Here, we demonstrate with primary nasopharyngeal cells grown at the air-liquid interface that the pseudostratified epithelium can be susceptible to EBV infection and we report that susceptible cell types with distinct EBV transcription profiles can be identified by single-cell RNA-sequencing. Although EBV infection in the nasopharynx has evaded detection in asymptomatic carriers, these findings demonstrate that EBV latent and lytic infection can occur in the cells of the nasopharyngeal epithelium.

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R9AP is a functional receptor for Epstein-Barr virus infection in both epithelial cells and B cells

10.21203/rs.3.rs-113641/v1 ◽

2020 ◽

Author(s):

Mu-Sheng Zeng ◽

li yan ◽

Hua Zhang ◽

Xiao-Dong Dong ◽

Cong Sun ◽

...

Keyword(s):

B Cells ◽

Epithelial Cells ◽

Epstein Barr Virus ◽

Human Tumor ◽

Neutralizing Antibody ◽

Epstein Barr Virus Infection ◽

Barr Virus ◽

Ebv Infection ◽

Epstein Barr

Abstract Epstein-Barr virus (EBV), also known as the first human tumor virus, is linked to about 200,000 new cancer cases and millions of non-malignant diseases every year. EBV infects both human epithelial cells and B cells. Several virally encoded glycoproteins define tropism and mediate a complicated entry process. Here, we show that in both epithelial cells and B cells, R9AP silencing or genetic knockout significantly inhibits EBV infection, whereas R9AP overexpression promotes EBV infection, establishing R9AP as an essential entry receptor for EBV. Mechanistically, R9AP directly binds to EBV glycoproteins gH/gL to mediate membrane fusion. Importantly, the interaction of R9AP with gH/gL is inhibited by the highly potent, competitive gH/gL neutralizing antibody AMMO1 that blocks EBV infection of both epithelial cells and B cells. Furthermore, a R9AP peptide encompassing the gH/gL binding site inhibits EBV infection in vitro and reduces viral load in EBV infected humanized mice. Altogether, we propose R9AP as the first characterized receptor for EBV infection common to epithelial cells and B cells and a potential target for intervention.

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