scholarly journals Recombinant bovine respiratory syncytial virus with deletions of the G or SH genes: G and F proteins bind heparin

2001 ◽  
Vol 82 (3) ◽  
pp. 631-640 ◽  
Author(s):  
Axel Karger ◽  
Ulrike Schmidt ◽  
Ursula J. Buchholz
Keyword(s):  
Respiratory Syncytial Virus ◽  
Reverse Genetics ◽  
Chondroitin Sulphate ◽  
Binding Activity ◽  
Bovine Respiratory Syncytial Virus ◽  
Deletion Mutants ◽  
Heparin Binding ◽  
Virus Attachment ◽  
Glycoprotein G ◽  
Syncytial Virus

Bovine respiratory syncytial virus (BRSV) encodes three transmembrane envelope glycoproteins, namely the small hydrophobic (SH) protein, the attachment glycoprotein (G) and the fusion glycoprotein (F). The BRSV reverse genetics system has been used to generate viable recombinant BRSV lacking either the G gene or the SH gene or both genes. The deletion mutants were fully competent for multicycle growth in cell culture, proving that, of the BRSV glycoprotein genes, the SH and G genes are non-essential. Virus morphogenesis was not impaired by either of the deletions. The deletion mutants were used to study the role of the F glycoprotein and the contributions of SH and G with respect to virus attachment. Attachment mediated by the F protein alone could be blocked by soluble heparin, but not by chondroitin sulphate. Heparin affinity chromatography revealed that both the BRSV G and F glycoproteins have heparin-binding activity, with the affinity of the F glycoprotein being significantly lower than that of G. Therefore, the roles of the BRSV glycoproteins in virus attachment and receptor binding have to be reconsidered.

scholarly journals Features of the Human Antibody Response against the Respiratory Syncytial Virus Surface Glycoprotein G

Vaccines ◽  
2020 ◽  
Vol 8 (2) ◽  
pp. 337
Author(s):  
Kristina Borochova ◽  
Katarzyna Niespodziana ◽  
Katarina Stenberg Hammar ◽  
Marianne van Hage ◽  
Gunilla Hedlin ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Surface Glycoprotein ◽  
Trigger Factor ◽  
Human Antibody ◽  
Th2 Response ◽  
Virus Surface ◽  
Virus Attachment ◽  
Peptide Epitopes ◽  
Glycoprotein G ◽  
Syncytial Virus

Respiratory syncytial virus (RSV) infections are a major cause of serious respiratory disease in infants. RSV occurs as two major subgroups A and B, which mainly differ regarding the surface glycoprotein G. The G protein is important for virus attachment and G-specific antibodies can protect against infection. We expressed the surface-exposed part of A2 strain-derived G (A2-G) in baculovirus-infected insect cells and synthesized overlapping peptides spanning complete A2-G. The investigation of the natural IgG response of adult subjects during a period of one year showed that IgG antibodies (i) recognize G significantly stronger than the fusion protein F0, (ii) target mainly non-conformational, sequential peptide epitopes from the exposed conserved region but also buried peptides, and (iii) exhibit a scattered but constant recognition profile during the observation period. The IgG subclass reactivity profile (IgG1 > IgG2 > IgG4 = IgG3) was indicative of a mixed Th1/Th2 response. Two strongly RSV-neutralizing sera including the 1st WHO standard contained high IgG anti-G levels. G-specific IgG increased strongly in children after wheezing attacks suggesting RSV as trigger factor. Our study shows that RSV G and G-derived peptides are useful for serological diagnosis of RSV-triggered exacerbations of respiratory diseases and underlines the importance of G for development of RSV-neutralizing vaccines.

scholarly journals Functional Analysis of the 60-Nucleotide Duplication in the Respiratory Syncytial Virus Buenos Aires Strain Attachment Glycoprotein

2015 ◽  
Vol 89 (16) ◽  
pp. 8258-8266 ◽  
Author(s):  
Anne L. Hotard ◽  
Elizabeth Laikhter ◽  
Kelsie Brooks ◽  
Tina V. Hartert ◽  
Martin L. Moore
Keyword(s):  
Respiratory Syncytial Virus ◽  
Buenos Aires ◽  
Virus Attachment ◽  
Recombinant Viruses ◽  
Fitness Advantage ◽  
Virus Life Cycle ◽  
Glycoprotein G ◽  
Syncytial Virus ◽  
Different Strains

ABSTRACTThere are two subgroups of respiratory syncytial virus (RSV), A and B, and within each subgroup, isolates are further divided into clades. Several years ago, multiple subgroup B isolates which contained a duplication of 60 nucleotides in the glycoprotein (G) gene were described. These isolates were given a new clade designation of BA based on the site of isolation, Buenos Aires, Argentina. BA RSV strains have since become the predominant circulating clade of RSV B viruses. We hypothesized that the duplicated region in G serves to enhance the function of G in the virus life cycle. We generated recombinant viruses that express a consensus BA G gene or a consensus BA G gene lacking the duplication (GΔdup). We determined that the duplicated region functions during virus attachment to cells. Additionally, we showed thatin vitro, the virus containing the duplication has a fitness advantage compared to the virus without the duplication. Our data demonstrate that the duplicated region in the BA strain G protein augments virus attachment and fitness.IMPORTANCERespiratory syncytial virus (RSV) is an important pathogen for infants for which there is no vaccine. Different strains of RSV circulate from year to year, and the predominating strains change over time. Subgroup B RSV strains with a duplication in the attachment glycoprotein (G) emerged and then became the dominant B genotype. We found that a recombinant virus harboring the duplication bound more efficiently to cells and was more fit than a recombinant strain lacking the duplication. Our work advances a mechanism for an important natural RSV mutation.

scholarly journals The role of the light chain in the structure and binding activity of two cattle antibodies that neutralize bovine respiratory syncytial virus

2019 ◽  
Vol 112 ◽  
pp. 123-130 ◽  
Author(s):  
Jingshan Ren ◽  
Joanne E. Nettleship ◽  
Gemma Harris ◽  
William Mwangi ◽  
Nahid Rhaman ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Light Chain ◽  
Binding Activity ◽  
Bovine Respiratory Syncytial Virus ◽  
Syncytial Virus

scholarly journals Bovine Respiratory Syncytial Virus Nonstructural Proteins NS1 and NS2 Cooperatively Antagonize Alpha/Beta Interferon-Induced Antiviral Response

2000 ◽  
Vol 74 (18) ◽  
pp. 8234-8242 ◽  
Author(s):  
Jörg Schlender ◽  
Birgit Bossert ◽  
Ursula Buchholz ◽  
Karl-Klaus Conzelmann
Keyword(s):  
Respiratory Syncytial Virus ◽  
Vero Cells ◽  
Bovine Respiratory Syncytial Virus ◽  
Deletion Mutants ◽  
Nonstructural Proteins ◽  
Antiviral Responses ◽  
Fold Reduction ◽  
Mdbk Cells ◽  
Syncytial Virus ◽  
In Cells

ABSTRACT The functions of bovine respiratory syncytial virus (BRSV) nonstructural proteins NS1 and NS2 were studied by generation and analysis of recombinant BRSV carrying single and double gene deletions. Whereas in MDBK cells the lack of either or both NS genes resulted in a 5,000- to 10,000-fold reduction of virus titers, in Vero cells a moderate (10-fold) reduction was observed. Interestingly, cell culture supernatants from infected MDBK cells were able to restrain the growth of NS deletion mutants in Vero cells, suggesting the involvement of NS proteins in escape from cytokine-mediated host cell responses. The responsible factors in MDBK supernatants were identified as type I interferons by neutralization of the inhibitory effect with antibodies blocking the alpha interferon (IFN-α) receptor. Treatment of cells with recombinant universal IFN-α A/D or IFN-β revealed severe inhibition of single and double deletion mutants, whereas growth of full-length BRSV was not greatly affected. Surprisingly, all NS deletion mutants were equally repressed, indicating an obligatory cooperation of NS1 and NS2 in antagonizing IFN-mediated antiviral mechanisms. To verify this finding, we generated recombinant rabies virus (rRV) expressing either NS1 or NS2 and determined their IFN sensitivity. In cells coinfected with NS1- and NS2-expressing rRVs, virus replication was resistant to doses of IFN which caused a 1,000-fold reduction of replication in cells infected with wild-type RV or with each of the NS-expressing rRVs alone. Thus, BRSV NS proteins have the potential to cooperatively protect an unrelated virus from IFN-α/β mediated antiviral responses. Interestingly, BRSV NS proteins provided a more pronounced resistance to IFN in the bovine cell line MDBK than in cell lines of other origins, suggesting adaptation to host-specific antiviral responses. The findings described have a major impact on the design of live recombinant BRSV and HRSV vaccines.

scholarly journals Characterization of an Experimental Vaccine for Bovine Respiratory Syncytial Virus

2014 ◽  
Vol 21 (7) ◽  
pp. 997-1004 ◽  
Author(s):  
Sara Hägglund ◽  
Kefei Hu ◽  
Krister Blodörn ◽  
Boby Makabi-Panzu ◽  
Anne-Laure Gaillard ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Vaccine Development ◽  
Cellular Membrane ◽  
Bovine Respiratory Syncytial Virus ◽  
Major Protein ◽  
Igg Antibody ◽  
Glycoprotein G ◽  
Syncytial Virus ◽  
Nucleoprotein N

ABSTRACTBovine respiratory syncytial virus (BRSV) and human respiratory syncytial virus (HRSV) are major causes of respiratory disease in calves and children, respectively, and are priorities for vaccine development. We previously demonstrated that an experimental vaccine, BRSV-immunostimulating complex (ISCOM), is effective in calves with maternal antibodies. The present study focuses on the antigenic characterization of this vaccine for the design of new-generation subunit vaccines. The results of our study confirmed the presence of membrane glycoprotein (G), fusion glycoprotein (F), and nucleoprotein (N) proteins in the ISCOMs, and this knowledge was extended by the identification of matrix (M), M2-1, phosphoprotein (P), small hydrophobic protein (SH) and of cellular membrane proteins, such as the integrins αVβ1, αVβ3, and α3β1. The quantity of the major protein F was 4- to 5-fold greater than that of N (∼77 μg versus ∼17 μg/calf dose), whereas G, M, M2-1, P, and SH were likely present in smaller amounts. The polymerase (L), M2-2, nonstructural 1 (NS1), and NS2 proteins were not detected, suggesting that they are not essential for protection. Sera from the BRSV-ISCOM-immunized calves contained high titers of IgG antibody specific for F, G, N, and SH. Antibody responses against M and P were not detected; however, this does not exclude their role in protective T-cell responses. The absence of immunopathological effects of the cellular proteins, such as integrins, needs to be further confirmed, and their possible contribution to adjuvant functions requires elucidation. This work suggests that a combination of several surface and internal proteins should be included in subunit RSV vaccines and identifies absent proteins as potential candidates for differentiating infected from vaccinated animals.

scholarly journals Mucosal Immunization with Live Recombinant Bovine Respiratory Syncytial Virus (BRSV) and Recombinant BRSV Lacking the Envelope Glycoprotein G Protects against Challenge with Wild-Type BRSV

2002 ◽  
Vol 76 (23) ◽  
pp. 12355-12359 ◽  
Author(s):  
Ulrike Schmidt ◽  
Jörg Beyer ◽  
Ulf Polster ◽  
Laurel J. Gershwin ◽  
Ursula J. Buchholz
Keyword(s):  
Respiratory Syncytial Virus ◽  
Neutralizing Antibodies ◽  
Protective Efficacy ◽  
Bovine Respiratory Syncytial Virus ◽  
Upper Respiratory Tract ◽  
Mucosal Immunization ◽  
Recombinant Viruses ◽  
Glycoprotein G ◽  
Upper Respiratory ◽  
Syncytial Virus

ABSTRACT Recombinant bovine respiratory syncytial virus (rBRSV) and an rBRSV deletion mutant lacking the G gene (rBRSVΔG) were characterized in calves with respect to replication competence, attenuation, and protective efficacy as live-attenuated BRSV vaccines. Both recombinant viruses were safe and induced protection against a BRSV challenge infection. rBRSV replicated efficiently in the upper respiratory tract. Intranasal immunization with rBRSVΔG led to infection but not to mucosal virus replication. Neutralizing antibodies were induced by rBRSV and rBRSVΔG. Thus, the BRSV attachment glycoprotein G seems to be dispensable in vaccinating calves against BRSV.

scholarly journals Identification of a Linear Heparin Binding Domain for Human Respiratory Syncytial Virus Attachment Glycoprotein G

1999 ◽  
Vol 73 (8) ◽  
pp. 6610-6617 ◽  
Author(s):  
Steven A. Feldman ◽  
R. Michael Hendry ◽  
Judy A. Beeler
Keyword(s):  
Respiratory Syncytial Virus ◽  
Vero Cells ◽  
Parental Cell ◽  
Binding Domain ◽  
Parental Cell Line ◽  
Virus Infectivity ◽  
Heparin Binding ◽  
Virus Attachment ◽  
Heparin Binding Domain ◽  
Syncytial Virus

ABSTRACT Respiratory syncytial virus (RSV) is the leading cause of lower respiratory tract disease in infants and young children worldwide. Infection is mediated, in part, by an initial interaction between attachment protein (G) and a highly sulfated heparin-like glycosaminoglycan (Gag) located on the cell surface. Synthetic overlapping peptides derived from consensus sequences of the G protein ectodomain from both RSV subgroups A and B were tested by heparin-agarose affinity chromatography for their abilities to bind heparin. This evaluation identified a single linear heparin binding domain (HBD) for RSV subgroup A (184A→T198) and B (183K→K197). The binding of these peptides to Vero cells was inhibited by heparin. Peptide binding to two CHO cell mutants (pgsD-677 and pgsA-745) deficient in heparan sulfate or total Gag synthesis was decreased 50% versus the parental cell line, CHO-K1, and decreased an average of 87% in the presence of heparin. The RSV-G HBD peptides were also able to inhibit homologous and heterologous virus infectivity of Vero cells. These results indicate that the sequence184A/183K→198T/K197for RSV subgroups A and B, respectively, defines an important determinant of RSV-G interactions with heparin.

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