scholarly journals Functional Analysis of the Fusion and Attachment Glycoproteins of Mojiang Henipavirus

Viruses ◽  
2021 ◽  
Vol 13 (3) ◽  
pp. 517
Author(s):  
Sofia Cheliout Da Silva ◽  
Lianying Yan ◽  
Ha V. Dang ◽  
Kai Xu ◽  
Jonathan H. Epstein ◽  
...  
Keyword(s):  
Functional Analysis ◽  
Cell Fusion ◽  
Sequence Data ◽  
Nipah Virus ◽  
Hendra Virus ◽  
Soluble Form ◽  
Fusion Activity ◽  
Low Levels ◽  
Entry Receptor ◽  
Functional Defects

Mojiang virus (MojV) is the first henipavirus identified in a rodent and known only by sequence data, whereas all other henipaviruses have been isolated from bats (Hendra virus, Nipah virus, Cedar virus) or discovered by sequence data from material of bat origin (Ghana virus). Ephrin-B2 and -B3 are entry receptors for Hendra and Nipah viruses, but Cedar virus can utilize human ephrin-B1, -B2, -A2 and -A5 and mouse ephrin-A1. However, the entry receptor for MojV remains unknown, and its species tropism is not well characterized. Here, we utilized recombinant full-length and soluble forms of the MojV fusion (F) and attachment (G) glycoproteins in membrane fusion and receptor tropism studies. MojV F and G were functionally competent and mediated cell–cell fusion in primate and rattine cells, albeit with low levels and slow fusion kinetics. Although a relative instability of the pre-fusion conformation of a soluble form of MojV F was observed, MojV F displayed significantly greater fusion activity when heterotypically paired with Ghana virus G. An exhaustive investigation of A- and B-class ephrins indicated that none serve as a primary receptor for MojV. The MojV cell fusion phenotype is therefore likely the result of receptor restriction rather than functional defects in recombinant MojV F and G glycoproteins.

scholarly journals Potent Neutralization of Hendra and Nipah Viruses by Human Monoclonal Antibodies

2006 ◽  
Vol 80 (2) ◽  
pp. 891-899 ◽  
Author(s):  
Zhongyu Zhu ◽  
Antony S. Dimitrov ◽  
Katharine N. Bossart ◽  
Gary Crameri ◽  
Kimberly A. Bishop ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Cell Fusion ◽  
Hendra Virus ◽  
Soluble Form ◽  
Enzyme Linked Immunosorbent Assay ◽  
Human Antibody ◽  
Emerging Viruses ◽  
Human Monoclonal Antibodies ◽  
Initial Attempt ◽  
Fusion Event

ABSTRACT Hendra virus (HeV) and Nipah virus (NiV) are closely related emerging viruses comprising the Henipavirus genus of the Paramyxovirinae. Each has a broad species tropism and can cause disease with high mortality in both animal and human hosts. These viruses infect cells by a pH-independent membrane fusion event mediated by their attachment (G) and fusion (F) envelope glycoproteins (Envs). Seven Fabs, m101 to -7, were selected for their significant binding to a soluble form of Hendra G (sG) which was used as the antigen for panning of a large naïve human antibody library. The selected Fabs inhibited, to various degrees, cell fusion mediated by the HeV or NiV Envs and virus infection. The conversion of the most potent neutralizer of infectious HeV, Fab m101, to immunoglobulin G1 (IgG1) significantly increased its cell fusion inhibitory activity: the 50% inhibitory concentration was decreased more than 10-fold to approximately 1 μg/ml. The IgG1 m101 was also exceptionally potent in neutralizing infectious HeV; complete (100%) neutralization was achieved with 12.5 μg/ml, and 98% neutralization required only 1.6 μg/ml. The inhibition of fusion and infection correlated with binding of the Fabs to full-length G as measured by immunoprecipitation and less with binding to sG as measured by enzyme-linked immunosorbent assay and Biacore. m101 and m102 competed with the ephrin-B2, which we recently identified as a functional receptor for both HeV and NiV, indicating a possible mechanism of neutralization by these antibodies. The m101, m102, and m103 antibodies competed with each other, suggesting that they bind to overlapping epitopes which are distinct from the epitopes of m106 and m107. In an initial attempt to localize the epitopes of m101 and m102, we measured their binding to a panel of 11 G alanine-scanning mutants and identified two mutants, P185A and Q191 K192A, which significantly decreased binding to m101 and one, G183, which decreased binding of m102 to G. These results suggest that m101 to -7 are specific for HeV or NiV or both and exhibit various neutralizing activities; they are the first human monoclonal antibodies identified against these viruses and could be used for treatment, prophylaxis, and diagnosis and as research reagents and could aid in the development of vaccines.

scholarly journals Structural and functional analyses reveal promiscuous and species specific use of ephrin receptors by Cedar virus

2019 ◽  
Vol 116 (41) ◽  
pp. 20707-20715 ◽  
Author(s):  
Eric D. Laing ◽  
Chanakha K. Navaratnarajah ◽  
Sofia Cheliout Da Silva ◽  
Stephanie R. Petzing ◽  
Yan Xu ◽  
...  
Keyword(s):  
Receptor Binding ◽  
Contact Region ◽  
Nipah Virus ◽  
Hendra Virus ◽  
Receptor Binding Site ◽  
Sequence Identity ◽  
Ephrin Receptors ◽  
Species Specific ◽  
Functional Analyses ◽  
Entry Receptor

Cedar virus (CedV) is a bat-borne henipavirus related to Nipah virus (NiV) and Hendra virus (HeV), zoonotic agents of fatal human disease. CedV receptor-binding protein (G) shares only ∼30% sequence identity with those of NiV and HeV, although they can all use ephrin-B2 as an entry receptor. We demonstrate that CedV also enters cells through additional B- and A-class ephrins (ephrin-B1, ephrin-A2, and ephrin-A5) and report the crystal structure of the CedV G ectodomain alone and in complex with ephrin-B1 or ephrin-B2. The CedV G receptor-binding site is structurally distinct from other henipaviruses, underlying its capability to accommodate additional ephrin receptors. We also show that CedV can enter cells through mouse ephrin-A1 but not human ephrin-A1, which differ by 1 residue in the key contact region. This is evidence of species specific ephrin receptor usage by a henipavirus, and implicates additional ephrin receptors in potential zoonotic transmission.

scholarly journals Novel Functions of Hendra Virus G N-Glycans and Comparisons to Nipah Virus

2015 ◽  
Vol 89 (14) ◽  
pp. 7235-7247 ◽  
Author(s):  
Birgit G. Bradel-Tretheway ◽  
Qian Liu ◽  
Jacquelyn A. Stone ◽  
Samantha McInally ◽  
Hector C. Aguilar
Keyword(s):  
Membrane Fusion ◽  
Cell Fusion ◽  
Immune Evasion ◽  
Viral Entry ◽  
Nipah Virus ◽  
Hendra Virus ◽  
Viral Attachment ◽  
Glycoprotein G ◽  
Protein Functions ◽  
Cell Cell

ABSTRACTHendra virus (HeV) and Nipah virus (NiV) are reportedly the most deadly pathogens within theParamyxoviridaefamily. These two viruses bind the cellular entry receptors ephrin B2 and/or ephrin B3 via the viral attachment glycoprotein G, and the concerted efforts of G and the viral fusion glycoprotein F result in membrane fusion. Membrane fusion is essential for viral entry into host cells and for cell-cell fusion, a hallmark of the disease pathobiology. HeV G is heavily N-glycosylated, but the functions of the N-glycans remain unknown. We disrupted eight predicted N-glycosylation sites in HeV G by conservative mutations (Asn to Gln) and found that six out of eight sites were actually glycosylated (G2 to G7); one in the stalk (G2) and five in the globular head domain (G3 to G7). We then tested the roles of individual and combined HeV G N-glycan mutants and found functions in the modulation of shielding against neutralizing antibodies, intracellular transport, G-F interactions, cell-cell fusion, and viral entry. Between the highly conserved HeV and NiV G glycoproteins, similar trends in the effects of N-glycans on protein functions were observed, with differences in the levels at which some N-glycan mutants affected such functions. While the N-glycan in the stalk domain (G2) had roles that were highly conserved between HeV and NiV G, individual N-glycans in the head affected the levels of several protein functions differently. Our findings are discussed in the context of their contributions to our understanding of HeV and NiV pathogenesis and immune responses.IMPORTANCEViral envelope glycoproteins are important for viral pathogenicity and immune evasion. N-glycan shielding is one mechanism by which immune evasion can be achieved. In paramyxoviruses, viral attachment and membrane fusion are governed by the close interaction of the attachment proteins H/HN/G and the fusion protein F. In this study, we show that the attachment glycoprotein G of Hendra virus (HeV), a deadly paramyxovirus, is N-glycosylated at six sites (G2 to G7) and that most of these sites have important roles in viral entry, cell-cell fusion, G-F interactions, G oligomerization, and immune evasion. Overall, we found that the N-glycan in the stalk domain (G2) had roles that were very conserved between HeV G and the closely related Nipah virus G, whereas individual N-glycans in the head quantitatively modulated several protein functions differently between the two viruses.

scholarly journals Exceptionally Potent Cross-Reactive Neutralization of Nipah and Hendra Viruses by a Human Monoclonal Antibody

2008 ◽  
Vol 197 (6) ◽  
pp. 846-853 ◽  
Author(s):  
Zhongyu Zhu ◽  
Katharine N Bossart ◽  
Kimberly A Bishop ◽  
Gary Crameri ◽  
Antony S Dimitrov ◽  
...  
Keyword(s):  
Therapeutic Agent ◽  
Human Monoclonal Antibody ◽  
Nipah Virus ◽  
Random Mutagenesis ◽  
Hendra Virus ◽  
Soluble Form ◽  
Antibody Library ◽  
Human Monoclonal Antibodies ◽  
Glycoprotein G ◽  
Better Than

Abstract We have previously identified neutralizing human monoclonal antibodies against Nipah virus (NiV) and Hendra virus (HeV) by panning a large nonimmune antibody library against a soluble form of the HeV attachment-envelope glycoprotein G (sGHeV). One of these antibodies, m102, which exhibited the highest level of cross-reactive neutralization of both NiV and HeV G, was affinity maturated by light-chain shuffling combined with random mutagenesis of its heavy-chain variable domain and panning against sGHeV. One of the selected antibody Fab clones, m102.4, had affinity of binding to sGHeV that was equal to or higher than that of the other Fabs; it was converted to IgG1 and tested against infectious NiV and HeV. It exhibited exceptionally potent and cross-reactive inhibitory activity with 50% inhibitory concentrations below 0.04 and 0.6 μg/mL, respectively. The virus-neutralizing activity correlated with the binding affinity of the antibody to sGHeV and sGNiV. m102.4 bound a soluble form of NiV G (sGNiV) better than it bound sGHeV, and it neutralized NiV better than HeV, despite being originally selected against sGHeV. These results suggest that m102.4 has potential as a therapeutic agent for the treatment of diseases caused by henipaviruses. It could be also used for prophylaxis and diagnosis, and as a research reagent

scholarly journals Acquisition of Cell–Cell Fusion Activity by Amino Acid Substitutions in Spike Protein Determines the Infectivity of a Coronavirus in Cultured Cells

PLoS ONE ◽  
2009 ◽  
Vol 4 (7) ◽  
pp. e6130 ◽  
Author(s):  
Yoshiyuki Yamada ◽  
Xiao Bo Liu ◽  
Shou Guo Fang ◽  
Felicia P. L. Tay ◽  
Ding Xiang Liu
Keyword(s):  
Amino Acid ◽  
Cell Fusion ◽  
Cultured Cells ◽  
Spike Protein ◽  
Amino Acid Substitutions ◽  
Fusion Activity ◽  
Cell Cell

scholarly journals Generation of Genetically RGD σ1-Modified Oncolytic Reovirus That Enhances JAM-A-Independent Infection of Tumor Cells

2020 ◽  
Vol 94 (23) ◽  
Author(s):  
Takahiro Kawagishi ◽  
Yuta Kanai ◽  
Ryotaro Nouda ◽  
Ichika Fukui ◽  
Jeffery A. Nurdin ◽  
...  
Keyword(s):  
Cell Lines ◽  
Cancer Cell ◽  
Cell Attachment ◽  
Genetically Modified ◽  
Cancer Cell Lines ◽  
Oncolytic Viruses ◽  
Peptide Sequence ◽  
Attachment Protein ◽  
Low Levels ◽  
Entry Receptor

ABSTRACT Mammalian reovirus (MRV) strain type 3 Dearing (T3D) is a naturally occurring oncolytic virus that has been developed as a potential cancer therapeutic. However, MRV treatment cannot be applied to cancer cells expressing low levels of junctional adhesion molecule A (JAM-A), which is the entry receptor of MRV. In this study, we developed a reverse genetics system for MRV strain T3D-L, which showed high oncolytic potency. To modify the cell tropism of MRV, an arginine–glycine–aspartic acid (RGD) peptide with an affinity to integrin was inserted at the C terminus or loop structures of the viral cell attachment protein σ1. The recombinant RGD σ1-modified viruses induced remarkable cell lysis in human cancer cell lines with marginal JAM-A expression and in JAM-A knockout cancer cell lines generated by a CRISPR/Cas9 system. Pretreatment of cells with anti-integrin antibody decreased cell death caused by the RGD σ1-modified virus, suggesting the infection to the cells was via a specific interaction with integrin αV. By using mouse models, we assessed virulence of the RGD σ1-modified viruses in vivo. This system will open new avenues for the use of genetically modified oncolytic MRV for use as a cancer therapy. IMPORTANCE Oncolytic viruses kill tumors without affecting normal cells. A variety of oncolytic viruses are used as cancer therapeutics. Mammalian reovirus (MRV), which belongs to the genus Orthoreovirus, family Reoviridae, is one such natural oncolytic virus. The anticancer effects of MRV are being evaluated in clinical trials. Unlike other oncolytic viruses, MRV has not been genetically modified for use as a cancer therapeutic in clinical trials. Here, we used a reverse genetic approach to introduce an integrin-affinity peptide sequence into the MRV cell attachment protein σ1 to alter the natural tropism of the virus. The recombinant viruses were able to infect cancer cell lines expressing very low levels of the MRV entry receptor, junctional adhesion molecule A (JAM-A), and cause tumor cell death while maintaining its original tropism via JAM-A. This is a novel report of a genetically modified oncolytic MRV by introducing a peptide sequence into σ1.

scholarly journals Variations in Disparate Regions of the Murine Coronavirus Spike Protein Impact the Initiation of Membrane Fusion

2001 ◽  
Vol 75 (6) ◽  
pp. 2792-2802 ◽  
Author(s):  
Dawn K. Krueger ◽  
Sean M. Kelly ◽  
Daniel N. Lewicki ◽  
Rosanna Ruffolo ◽  
Thomas M. Gallagher
Keyword(s):  
Tissue Culture ◽  
Membrane Fusion ◽  
Cultured Cells ◽  
Hepatitis Virus ◽  
Fusion Reaction ◽  
Spike Protein ◽  
Soluble Form ◽  
Fusion Activity ◽  
Murine Hepatitis Virus

ABSTRACT The prototype JHM strain of murine hepatitis virus (MHV) is an enveloped, RNA-containing coronavirus that has been selected in vivo for extreme neurovirulence. This virus encodes spike (S) glycoproteins that are extraordinarily effective mediators of intercellular membrane fusion, unique in their ability to initiate fusion even without prior interaction with the primary MHV receptor, a murine carcinoembryonic antigen-related cell adhesion molecule (CEACAM). In considering the possible role of this hyperactive membrane fusion activity in neurovirulence, we discovered that the growth of JHM in tissue culture selected for variants that had lost murine CEACAM-independent fusion activity. Among the collection of variants, mutations were identified in regions encoding both the receptor-binding (S1) and fusion-inducing (S2) subunits of the spike protein. Each mutation was separately introduced into cDNA encoding the prototype JHM spike, and the set of cDNAs was expressed using vaccinia virus vectors. The variant spikes were similar to that of JHM in their assembly into oligomers, their proteolysis into S1 and S2 cleavage products, their transport to cell surfaces, and their affinity for a soluble form of murine CEACAM. However, these tissue culture-adapted spikes were significantly stabilized as S1-S2 heteromers, and their entirely CEACAM-dependent fusion activity was delayed or reduced relative to prototype JHM spikes. The mutations that we have identified therefore point to regions of the S protein that specifically regulate the membrane fusion reaction. We suggest that cultured cells, unlike certain in vivo environments, select for S proteins with delayed, CEACAM-dependent fusion activities that may increase the likelihood of virus internalization prior to the irreversible uncoating process.

scholarly journals C-Terminal Tyrosine Residues Modulate the Fusion Activity of the Hendra Virus Fusion Protein

Biochemistry ◽  
2011 ◽  
Vol 50 (6) ◽  
pp. 945-952 ◽  
Author(s):  
Andreea Popa ◽  
Cara Teresia Pager ◽  
Rebecca Ellis Dutch
Keyword(s):  
Fusion Protein ◽  
Hendra Virus ◽  
Fusion Activity ◽  
Tyrosine Residues ◽  
Virus Fusion

scholarly journals Chloroquine Administration Does Not Prevent Nipah Virus Infection and Disease in Ferrets

2009 ◽  
Vol 83 (22) ◽  
pp. 11979-11982 ◽  
Author(s):  
Jackie Pallister ◽  
Deborah Middleton ◽  
Gary Crameri ◽  
Manabu Yamada ◽  
Reuben Klein ◽  
...  
Keyword(s):  
Virus Infection ◽  
Nipah Virus ◽  
Disease Outbreaks ◽  
Mortality Rates ◽  
Hendra Virus ◽  
Malaria Therapy ◽  
Sporadic Disease

ABSTRACT Hendra virus and Nipah virus, two zoonotic paramyxoviruses in the genus Henipavirus, have recently emerged and continue to cause sporadic disease outbreaks in humans and animals. Mortality rates of up to 75% have been reported in humans, but there are presently no clinically licensed therapeutics for treating henipavirus-induced disease. A recent report indicated that chloroquine, used in malaria therapy for over 70 years, prevented infection with Nipah virus in vitro. Chloroquine was assessed using a ferret model of lethal Nipah virus infection and found to be ineffective against Nipah virus infection in vivo.

Sign in / Sign up

Export Citation Format

Share Document