scholarly journals Multiple glycosylated forms of the respiratory syncytial virus fusion protein are expressed in virus-infected cells

2002 ◽  
Vol 83 (1) ◽  
pp. 61-66 ◽  
Author(s):  
Helen W. McL. Rixon ◽  
Craig Brown ◽  
Gaie Brown ◽  
Richard J. Sugrue
Keyword(s):  
Respiratory Syncytial Virus ◽  
Fusion Protein ◽  
Vero Cells ◽  
Cross Linking ◽  
Protein Species ◽  
Glycosylation Pattern ◽  
F Protein ◽  
Virus Fusion ◽  
Infected Cells ◽  
Syncytial Virus

Analysis of the respiratory syncytial virus (RSV) fusion (F) protein in RSV-infected Vero cells showed the presence of a single F1 subunit and at least two different forms of the F2 subunit, designated F2a (21 kDa) and F2b (16 kDa), which were collectively referred to as [F2]a/b. Enzymatic deglycosylation of [F2]a/b produced a single 10 kDa product suggesting that [F2]a/b arises from differences in the glycosylation pattern of F2a and F2b. The detection of [F2]a/b was dependent upon the post-translational cleavage of the F protein by furin, since its appearance was prevented in RSV-infected Vero cells treated with the furin inhibitor dec-RVKR-cmk. Analysis by protein cross-linking revealed that the F1 subunit interacted with [F2]a/b, via disulphide bonding, to produce equivalent F protein trimers, which were expressed on the surface of infected cells. Collectively, these data show that multiple F protein species are expressed in RSV-infected cells.

scholarly journals Furin cleavage of the respiratory syncytial virus fusion protein is not a requirement for its transport to the surface of virus-infected cells

2001 ◽  
Vol 82 (6) ◽  
pp. 1375-1386 ◽  
Author(s):  
Richard J. Sugrue ◽  
Craig Brown ◽  
Gaie Brown ◽  
James Aitken ◽  
Helen W. McL. Rixon
Keyword(s):  
Respiratory Syncytial Virus ◽  
Vero Cells ◽  
Intermediate Form ◽  
Furin Cleavage ◽  
F Protein ◽  
Cell Surface Biotinylation ◽  
Inactive Form ◽  
Infected Cells ◽  
Syncytial Virus ◽  
Post Synthesis

The intracellular cleavage of respiratory syncytial virus (RSV) fusion (F) protein by furin was examined. In RSV-infected LoVo cells, which express an inactive form of furin, and in RSV-infected Vero cells treated with the furin inhibitor decanoyl-Arg-Val-Lys-Arg-chloromethyl ketone (dec-RVKR-cmk), the F protein was expressed as a non-cleaved 73 kDa species. In both cases the F protein was initially expressed as an endoglycosidase H (Endo H)-sensitive precursor (F0EHs) which was modified approximately 40 min post-synthesis by the addition of complex carbohydrates to produce the Endo H-resistant form (F0EHr). The size and glycosylation state of F0EHr were identical to a transient intermediate form of non-cleaved F protein which was detected in RSV-infected Vero cells in the absence of inhibitor. Cell surface biotinylation and surface immunofluorescence staining showed that F0EHr was present on the surface of RSV-infected cells. RSV filaments have been shown to be the predominant form of the budding virus that is detected during virus replication. Analysis of the RSV-infected cells using scanning electron microscopy (SEM) showed that, in the presence of dec-RVKR-cmk, virus budding was impaired, producing fewer and much smaller viral filaments than in untreated cells. A comparison of immunofluorescence and SEM data showed that F0EHr was routed to the surface of virus-infected cells but not located in these smaller structures. Our findings suggest that activation of the F protein is required for the efficient formation of RSV filaments.

scholarly journals Cleavage at the Furin Consensus Sequence RAR/KR109 and Presence of the Intervening Peptide of the Respiratory Syncytial Virus Fusion Protein Are Dispensable for Virus Replication in Cell Culture

2002 ◽  
Vol 76 (18) ◽  
pp. 9218-9224 ◽  
Author(s):  
Gert Zimmer ◽  
Karl-Klaus Conzelmann ◽  
Georg Herrler
Keyword(s):  
Cell Culture ◽  
Respiratory Syncytial Virus ◽  
Fusion Protein ◽  
Reverse Genetics ◽  
Consensus Sequence ◽  
Vero Cells ◽  
Proteolytic Processing ◽  
Furin Cleavage ◽  
F Protein ◽  
Syncytial Virus

ABSTRACT Proteolytic processing of the respiratory syncytial virus F (fusion) protein results in the generation of the disulfide-linked subunits F1 and F2 and in the release of pep27, a glycopeptide originally located between the two furin cleavage sites FCS-1 (RKRR136) and FCS-2 (RAR/KR109). We made use of reverse genetics to study the importance of FCS-2 and of pep27 for BRSV replication in cell culture. Replacement of FCS-2 in the F protein of recombinant viruses by either of the sequences NANR109, RANN109 or SANN109, respectively, abolished proteolytic processing at this position, whereas the cleavage of FCS-1 was not affected. All mutants replicated in calf kidney and Vero cells in the absence of exogenous trypsin, although somewhat higher titers of BRSV containing the NANR109 or the RANN109 motif were achieved in the presence of trypsin. The virus mutants showed a reduced cytopathic effect which was lowest in the case of the SANN109 mutant. These findings demonstrate that cleavage at FCS-2 is dispensable for replication of respiratory syncytial virus in cell culture. A deletion mutant containing FCS-1 but lacking FCS-2 and most of pep27 replicated in cell culture as efficiently as the parental virus, indicating that this domain of the F protein is not essential for virus maturation and infectivity.

Vaccination with prefusion-stabilized respiratory syncytial virus fusion protein induces genetically and antigenically diverse antibody responses

Immunity ◽  
2021 ◽  
Author(s):  
Maryam Mukhamedova ◽  
Daniel Wrapp ◽  
Chen-Hsiang Shen ◽  
Morgan S.A. Gilman ◽  
Tracy J. Ruckwardt ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Fusion Protein ◽  
Antibody Responses ◽  
Virus Fusion ◽  
Syncytial Virus

scholarly journals Analysis of the Respiratory Syncytial Virus Fusion Protein Using Monoclonal and Polyclonal Antibodies

1986 ◽  
Vol 67 (3) ◽  
pp. 505-513 ◽  
Author(s):  
E. E. Walsh ◽  
P. J. Cote ◽  
B. F. Fernie ◽  
J. J. Schlesinger ◽  
M. W. Brandriss
Keyword(s):  
Respiratory Syncytial Virus ◽  
Fusion Protein ◽  
Polyclonal Antibodies ◽  
Virus Fusion ◽  
Syncytial Virus

The Design of Vaccines Based on the Shielding of Antigenic Site Ø of a Respiratory Syncytial Virus Fusion Protein Immunogen

2020 ◽  
pp. 2000714
Author(s):  
Steven J. Frey ◽  
Chad Varner ◽  
Ammar Arsiwala ◽  
Michael G. Currier ◽  
Martin L. Moore ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Fusion Protein ◽  
Antigenic Site ◽  
Virus Fusion ◽  
Syncytial Virus

scholarly journals Inferior immunogenicity and efficacy of respiratory syncytial virus fusion protein-based subunit vaccine candidates in aged versus young mice

PLoS ONE ◽  
2017 ◽  
Vol 12 (11) ◽  
pp. e0188708 ◽  
Author(s):  
Corinne Cayatte ◽  
Angie Snell Bennett ◽  
Gaurav Manohar Rajani ◽  
Leigh Hostetler ◽  
Sean K. Maynard ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Fusion Protein ◽  
Subunit Vaccine ◽  
Vaccine Candidates ◽  
Virus Fusion ◽  
Syncytial Virus ◽  
Young Mice

scholarly journals RAGE inhibits human respiratory syncytial virus syncytium formation by interfering with F-protein function

2013 ◽  
Vol 94 (8) ◽  
pp. 1691-1700 ◽  
Author(s):  
Jane Tian ◽  
Kelly Huang ◽  
Subramaniam Krishnan ◽  
Catherine Svabek ◽  
Daniel C. Rowe ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Epithelial Cells ◽  
Syncytium Formation ◽  
Human Respiratory Syncytial Virus ◽  
Host Cells ◽  
Type I ◽  
F Protein ◽  
Viral Spread ◽  
Infected Cells ◽  
Syncytial Virus

Human respiratory syncytial virus (RSV) is a major cause of severe lower respiratory tract infection. Infection is critically dependent on the RSV fusion (F) protein, which mediates fusion between the viral envelope and airway epithelial cells. The F protein is also expressed on infected cells and is responsible for fusion of infected cells with adjacent cells, resulting in the formation of multinucleate syncytia. The receptor for advanced glycation end products (RAGE) is a pattern-recognition receptor that is constitutively highly expressed by type I alveolar epithelial cells. Here, we report that RAGE protected HEK cells from RSV-induced cell death and reduced viral titres in vitro. RAGE appeared to interact directly with the F protein, but, rather than inhibiting RSV entry into host cells, virus replication and budding, membrane-expressed RAGE or soluble RAGE blocked F-protein-mediated syncytium formation and sloughing. These data indicate that RAGE may contribute to protecting the lower airways from RSV by inhibiting the formation of syncytia, viral spread, epithelial damage and airway obstruction.

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