scholarly journals Reversible disruption of XPO1-mediated nuclear export inhibits respiratory syncytial virus (RSV) replication

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Cynthia Mathew ◽  
Sharon Tamir ◽  
Ralph A. Tripp ◽  
Reena Ghildyal
Keyword(s):  
Respiratory Syncytial Virus ◽  
Nuclear Export ◽  
Cell Cycle Progression ◽  
Virus Assembly ◽  
The Elderly ◽  
M Protein ◽  
Structural Protein ◽  
Syncytial Virus ◽  
Tract Disease ◽  
High Risk Infants

AbstractRespiratory syncytial virus (RSV) is the primary cause of serious lower respiratory tract disease in infants, young children, the elderly and immunocompromised individuals. Therapy for RSV infections is limited to high risk infants and there are no safe and efficacious vaccines. Matrix (M) protein is a major RSV structural protein with a key role in virus assembly. Interestingly, M is localised to the nucleus early in infection and its export into the cytoplasm by the nuclear exporter, exportin-1 (XPO1) is essential for RSV assembly. We have shown previously that chemical inhibition of XPO1 function results in reduced RSV replication. In this study, we have investigated the anti-RSV efficacy of Selective Inhibitor of Nuclear Export (SINE) compounds, KPT-335 and KPT-185. Our data shows that therapeutic administration of the SINE compounds results in reduced RSV titre in human respiratory epithelial cell culture. Within 24 h of treatment, RSV replication and XPO1 expression was reduced, M protein was partially retained in the nucleus, and cell cycle progression was delayed. Notably, the effect of SINE compounds was reversible within 24 h after their removal. Our data show that reversible inhibition of XPO1 can disrupt RSV replication by affecting downstream pathways regulated by the nuclear exporter.

scholarly journals The Respiratory Syncytial Virus Matrix Protein Possesses a Crm1-Mediated Nuclear Export Mechanism

2009 ◽  
Vol 83 (11) ◽  
pp. 5353-5362 ◽  
Author(s):  
Reena Ghildyal ◽  
Adeline Ho ◽  
Manisha Dias ◽  
Lydia Soegiyono ◽  
Phillip G. Bardin ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Nuclear Export ◽  
Matrix Protein ◽  
Virus Assembly ◽  
Nuclear Export Signal ◽  
M Protein ◽  
Nuclear Accumulation ◽  
Infected Cells ◽  
Syncytial Virus ◽  
In Cells

ABSTRACT The respiratory syncytial virus (RSV) matrix (M) protein is localized in the nucleus of infected cells early in infection but is mostly cytoplasmic late in infection. We have previously shown that M localizes in the nucleus through the action of the importin β1 nuclear import receptor. Here, we establish for the first time that M's ability to shuttle to the cytoplasm is due to the action of the nuclear export receptor Crm1, as shown in infected cells, and in cells transfected to express green fluorescent protein (GFP)-M fusion proteins. Specific inhibition of Crm1-mediated nuclear export by leptomycin B increased M nuclear accumulation. Analysis of truncated and point-mutated M derivatives indicated that Crm1-dependent nuclear export of M is attributable to a nuclear export signal (NES) within residues 194 to 206. Importantly, inhibition of M nuclear export resulted in reduced virus production, and a recombinant RSV carrying a mutated NES could not be rescued by reverse genetics. That this is likely to be due to the inability of a nuclear export deficient M to localize to regions of virus assembly is indicated by the fact that a nuclear-export-deficient GFP-M fails to localize to regions of virus assembly when expressed in cells infected with wild-type RSV. Together, our data suggest that Crm1-dependent nuclear export of M is central to RSV infection, representing the first report of such a mechanism for a paramyxovirus M protein and with important implications for related paramyxoviruses.

scholarly journals Characterization of recombinant influenza A virus as a vector expressing respiratory syncytial virus fusion protein epitopes

2014 ◽  
Vol 95 (9) ◽  
pp. 1886-1891 ◽  
Author(s):  
Peirui Zhang ◽  
Hongjing Gu ◽  
Chengrong Bian ◽  
Na Liu ◽  
Zhiwei Li ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Respiratory Syncytial Virus ◽  
Cell Line ◽  
Nuclear Export ◽  
Influenza A ◽  
Haemagglutination Inhibition ◽  
The Elderly ◽  
Ns Gene ◽  
Syncytial Virus

Respiratory syncytial virus (RSV) is the most common cause of respiratory infection in infants and the elderly, and no vaccine against this virus has yet been licensed. Here, we report a recombinant PR8 influenza virus with the RSV fusion (F) protein epitopes of the subgroup A gene inserted into the influenza virus non-structural (NS) gene (rFlu/RSV/F) that was generated as an RSV vaccine candidate. The rescued viruses were assessed by microscopy and Western blotting. The proper expression of NS1, the NS gene product, and the nuclear export protein (NEP) of rFlu/RSV/F was also investigated using an immunofluorescent assay. The rescued virus replicated well in the MDCK kidney cell line, A549 lung adenocarcinoma cell line and CNE-2Z nasopharyngeal carcinoma cell line. BALB/c mice immunized intranasally with rFlu/RSV/F had specific haemagglutination inhibition antibody responses against the PR8 influenza virus and RSV neutralization test proteins. Furthermore, intranasal immunization with rFlu/RSV/F elicited T helper type 1-dominant cytokine profiles against the RSV strain A2 virus. Taken together, our findings suggested that rFlu/RSV/F was immunogenic in vivo and warrants further development as a promising candidate vaccine.

scholarly journals Dual Proinflammatory and Antiviral Properties of Pulmonary Eosinophils in Respiratory Syncytial Virus Vaccine-Enhanced Disease

2014 ◽  
Vol 89 (3) ◽  
pp. 1564-1578 ◽  
Author(s):  
Yung-Chang Su ◽  
Dijana Townsend ◽  
Lara J. Herrero ◽  
Ali Zaid ◽  
Michael S. Rolph ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
The Elderly ◽  
Pulmonary Eosinophilia ◽  
Recurrent Wheezing ◽  
Interleukin 5 ◽  
Lower Respiratory Tract Disease ◽  
Syncytial Virus ◽  
Tract Disease ◽  
Deficient Mice

ABSTRACTHuman respiratory syncytial virus (RSV) is a major cause of morbidity and severe lower respiratory tract disease in the elderly and very young, with some infants developing bronchiolitis, recurrent wheezing, and asthma following infection. Previous studies in humans and animal models have shown that vaccination with formalin-inactivated RSV (FI-RSV) leads to prominent airway eosinophilic inflammation following RSV challenge; however, the roles of pulmonary eosinophilia in the antiviral response and in disease pathogenesis are inadequately understood.In vivostudies in mice with eotaxin and/or interleukin 5 (IL-5) deficiency showed that FI-RSV vaccination did not lead to enhanced pulmonary disease, where following challenge there were reduced pulmonary eosinophilia, inflammation, Th2-type cytokine responses, and altered chemokine (TARC and CCL17) responses. In contrast to wild-type mice, RSV was recovered at high titers from the lungs of eotaxin- and/or IL-5-deficient mice. Adoptive transfer of eosinophils to FI-RSV-immunized eotaxin- and IL-5-deficient (double-deficient) mice challenged with RSV was associated with potent viral clearance that was mediated at least partly through nitric oxide. These studies show that pulmonary eosinophilia has dual outcomes: one linked to RSV-induced airway inflammation and pulmonary pathology and one with innate features that contribute to a reduction in the viral load.IMPORTANCEThis study is critical to understanding the mechanisms attributable to RSV vaccine-enhanced disease. This study addresses the hypothesis that IL-5 and eotaxin are critical in pulmonary eosinophil response related to FI-RSV vaccine-enhanced disease. The findings suggest that in addition to mediating tissue pathology, eosinophils within a Th2 environment also have antiviral activity.

scholarly journals A Decade of Respiratory Syncytial Virus Epidemiology and Prophylaxis: Translating Evidence into Everyday Clinical Practice

2011 ◽  
Vol 18 (2) ◽  
pp. e10-e19 ◽  
Author(s):  
Bosco A Paes ◽  
Ian Mitchell ◽  
Anna Banerji ◽  
Krista L Lanctôt ◽  
Joanne M Langley
Keyword(s):  
Respiratory Syncytial Virus ◽  
High Risk ◽  
Health Care Professionals ◽  
Burden Of Illness ◽  
Scientific Evidence ◽  
Rsv Infection ◽  
Syncytial Virus ◽  
Tract Disease ◽  
High Risk Infants ◽  
Long Term Impact

Respiratory syncytial virus (RSV) is a common infection in infancy, with nearly all children affected by two years of age. Approximately 0.5% to 2.0% of all children are hospitalized with lower respiratory tract disease, of which 50% to 90% have bronchiolitis and 5% to 40% have pneumonia. Morbidity and mortality are highest in children with nosocomial infection and in those with underlying medical illnesses such as cardiac and chronic lung disease. Aboriginal children residing in remote northern regions are specifically considered to be at high risk for hospitalization due to RSV infection. Thorough hand washing and health education are the principal strategies in primary prevention. In the absence of a vaccine, palivizumab prophylaxis is currently the best intervention to reduce the burden of illness and RSV-related hospitalization in high-risk children. Health care professionals should provide palivizumab prophylaxis cost effectively in accordance with recommendations issued by pediatric societies and national advisory bodies.The present article reviews the epidemiology of RSV infection and the short- and long-term impact of disease in high-risk infants and special populations. Prevention strategies and treatment are discussed based on the existing scientific evidence, and future challenges in the management of RSV infection are addressed.

scholarly journals Verdinexor (KPT-335), a Selective Inhibitor of Nuclear Export, Reduces Respiratory Syncytial Virus Replication In Vitro

2018 ◽  
Vol 93 (4) ◽  
Author(s):  
Patricia A. Jorquera ◽  
Cynthia Mathew ◽  
Jennifer Pickens ◽  
Colin Williams ◽  
Jasmina M. Luczo ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Young Children ◽  
Nuclear Export ◽  
Symptomatic Relief ◽  
Antiviral Drugs ◽  
M Protein ◽  
Nuclear Accumulation ◽  
Syncytial Virus ◽  
Infants And Young Children

ABSTRACT Respiratory syncytial virus (RSV) is a leading cause of hospitalization of infants and young children, causing considerable respiratory disease and repeat infections that may lead to chronic respiratory conditions such as asthma, wheezing, and bronchitis. RSV causes ∼34 million new episodes of lower respiratory tract illness (LRTI) in children younger than 5 years of age, with >3 million hospitalizations due to severe RSV-associated LRTI. The standard of care is limited to symptomatic relief as there are no approved vaccines and few effective antiviral drugs; thus, a safe and efficacious RSV therapeutic is needed. Therapeutic targeting of host proteins hijacked by RSV to facilitate replication is a promising antiviral strategy as targeting the host reduces the likelihood of developing drug resistance. The nuclear export of the RSV M protein, mediated by the nuclear export protein exportin 1 (XPO1), is crucial for RSV assembly and budding. Inhibition of RSV M protein export by leptomycin B correlated with reduced RSV replication in vitro. In this study, we evaluated the anti-RSV efficacy of Verdinexor (KPT-335), a small molecule designed to reversibly inhibit XPO1-mediated nuclear export. KPT-335 inhibited XPO1-mediated transport and reduced RSV replication in vitro. KPT-335 was effective against RSV A and B strains and reduced viral replication following prophylactic or therapeutic administration. Inhibition of RSV replication by KPT-335 was due to a combined effect of reduced XPO1 expression, disruption of the nuclear export of RSV M protein, and inactivation of the NF-κB signaling pathway. IMPORTANCE RSV is an important cause of LRTI in infants and young children for which there are no suitable antiviral drugs offered. We evaluated the efficacy of KPT-335 as an anti-RSV drug and show that KPT-335 inhibits XPO1-mediated nuclear export, leading to nuclear accumulation of RSV M protein and reduction in RSV levels. KPT-335 treatment also resulted in inhibition of proinflammatory pathways, which has important implications for its effectiveness in vivo.

scholarly journals The Morphology and Assembly of Respiratory Syncytial Virus Revealed by Cryo-Electron Tomography

Viruses ◽  
2018 ◽  
Vol 10 (8) ◽  
pp. 446 ◽  
Author(s):  
Zunlong Ke ◽  
Rebecca Dillard ◽  
Tatiana Chirkova ◽  
Fredrick Leon ◽  
Christopher Stobart ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Electron Tomography ◽  
Rna Virus ◽  
3D Structure ◽  
Three Dimensional ◽  
The Elderly ◽  
Cryo Electron Tomography ◽  
Infected Cells ◽  
Syncytial Virus ◽  
Tract Disease

Human respiratory syncytial virus (RSV) is the leading cause of lower respiratory tract disease in young children. With repeat infections throughout life, it can also cause substantial disease in the elderly and in adults with compromised cardiac, pulmonary and immune systems. RSV is a pleomorphic enveloped RNA virus in the Pneumoviridae family. Recently, the three-dimensional (3D) structure of purified RSV particles has been elucidated, revealing three distinct morphological categories: spherical, asymmetric, and filamentous. However, the native 3D structure of RSV particles associated with or released from infected cells has yet to be investigated. In this study, we have established an optimized system for studying RSV structure by imaging RSV-infected cells on transmission electron microscopy (TEM) grids by cryo-electron tomography (cryo-ET). Our results demonstrate that RSV is filamentous across several virus strains and cell lines by cryo-ET, cryo-immuno EM, and thin section TEM techniques. The viral filament length varies from 0.5 to 12 μm and the average filament diameter is approximately 130 nm. Taking advantage of the whole cell tomography technique, we have resolved various stages of RSV assembly. Collectively, our results can facilitate the understanding of viral morphogenesis in RSV and other pleomorphic enveloped viruses.

scholarly journals Development of Luciferase Immunoprecipitation Systems (LIPS) Assay to Detect IgG Antibodies against Human Respiratory Syncytial Virus G-Glycoprotein

Vaccines ◽  
2019 ◽  
Vol 7 (1) ◽  
pp. 16 ◽  
Author(s):  
Roberta Crim ◽  
Sangeeta Kumari ◽  
Priyanka Jayanti ◽  
Susette Audet ◽  
Ashwin Kulkarni ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Measles Virus ◽  
Vaccine Development ◽  
The Elderly ◽  
Igg Antibodies ◽  
Serum Samples ◽  
Genes Encoding ◽  
Syncytial Virus ◽  
Tract Disease ◽  
Paired Serum

Respiratory syncytial virus (RSV) causes severe lower respiratory tract disease in infants and the elderly. Although there is no licensed vaccine, RSV-F and -G glycoproteins are targets for vaccine development and therapeutics. We developed an assay that can detect anti-RSV-G IgG antibodies, either as a biomarker of natural exposure or immunization. RSV genes encoding native and mutated G (mG) proteins from subgroups A and B strains were cloned, expressed as luciferase-tagged proteins, and tested individually to detect anti-RSV-G specific IgG antibodies using a high-throughput luciferase immunoprecipitation system (LIPS-G). RSV monoclonal antibodies and polyclonal antisera specifically bound in the LIPS-GA and/or -GB assays; whereas anti-RSV-F and -N, and antisera against measles virus or human metapneumovirus did not bind. Anti-RSV-GA and -GB IgG responses detected in mice infected intranasally with RSV-A or -B strains were subtype specific. Subtype specific anti-RSV-GA or -GB IgG responses were also detected using paired serum samples from infants while human adolescent serum samples reacted in both LIPS-GA and -GB assays, reflecting a broader experience.

scholarly journals Association of Respiratory Syncytial Virus M Protein with Viral Nucleocapsids Is Mediated by the M2-1 Protein

2008 ◽  
Vol 82 (17) ◽  
pp. 8863-8870 ◽  
Author(s):  
Dongsheng Li ◽  
David A. Jans ◽  
Phillip G. Bardin ◽  
Jayesh Meanger ◽  
John Mills ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Virus Assembly ◽  
M Protein ◽  
Viral Transcription ◽  
Nucleocapsid Proteins ◽  
Cytoplasmic Inclusions ◽  
The Matrix ◽  
A Cell ◽  
Syncytial Virus ◽  
P Proteins

ABSTRACT Cytoplasmic inclusions in respiratory syncytial virus-infected cells comprising viral nucleocapsid proteins (L, N, P, and M2-1) and the viral genome are sites of viral transcription. Although not believed to be necessary for transcription, the matrix (M) protein is also present in these inclusions, and we have previously shown that M inhibits viral transcription. In this study, we have investigated the mechanisms for the association of the M protein with cytoplasmic inclusions. Our data demonstrate for the first time that the M protein associates with cytoplasmic inclusions via an interaction with the M2-1 protein. The M protein colocalizes with M2-1 in the cytoplasm of cells expressing only the M and M2-1 proteins and directly interacts with M2-1 in a cell-free binding assay. Using a cotransfection system, we confirmed that the N and P proteins are sufficient to form cytoplasmic inclusions and that M2-1 localizes to these inclusions; additionally, we show that M associates with cytoplasmic inclusions only in the presence of the M2-1 protein. Using truncated mutants, we show that the N-terminal 110 amino acids of M mediate the interaction with M2-1 and the subsequent association with nucleocapsids. The interaction of M2-1 with M and, in particular, the N-terminal region of M may represent a target for novel antivirals that block the association of M with nucleocapsids, thereby inhibiting virus assembly.

scholarly journals Considerations for a Respiratory Syncytial Virus Vaccine Targeting an Elderly Population

Vaccines ◽  
2021 ◽  
Vol 9 (6) ◽  
pp. 624
Author(s):  
Laura M. Stephens ◽  
Steven M. Varga
Keyword(s):  
Respiratory Syncytial Virus ◽  
Disease Burden ◽  
Elderly Population ◽  
Vaccine Development ◽  
The Elderly ◽  
The United States ◽  
Susceptible Population ◽  
Age Related ◽  
Syncytial Virus ◽  
Tract Infections

Respiratory syncytial virus (RSV) is most commonly associated with acute lower respiratory tract infections in infants and children. However, RSV also causes a high disease burden in the elderly that is often under recognized. Adults >65 years of age account for an estimated 80,000 RSV-associated hospitalizations and 14,000 deaths in the United States annually. RSV infection in aged individuals can result in more severe disease symptoms including pneumonia and bronchiolitis. Given the large disease burden caused by RSV in the aged, this population remains an important target for vaccine development. Aging results in lowered immune responsiveness characterized by impairments in both innate and adaptive immunity. This immune senescence poses a challenge when developing a vaccine targeting elderly individuals. An RSV vaccine tailored towards an elderly population will need to maximize the immune response elicited in order to overcome age-related defects in the immune system. In this article, we review the hurdles that must be overcome to successfully develop an RSV vaccine for use in the elderly, and discuss the vaccine candidates currently being tested in this highly susceptible population.

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