Neuraminidase Is Important for the Initiation of Influenza Virus Infection in Human Airway Epithelium

ABSTRACT Influenza virus neuraminidase (NA) plays an essential role in release and spread of progeny virions, following the intracellular viral replication cycle. To test whether NA could also facilitate virus entry into cell, we infected cultures of human airway epithelium with human and avian influenza viruses in the presence of the NA inhibitor oseltamivir carboxylate. Twenty- to 500-fold less cells became infected in drug-treated versus nontreated cultures (P < 0.0001) 7 h after virus application, indicating that the drug suppressed the initiation of infection. These data demonstrate that viral NA plays a role early in infection, and they provide further rationale for the prophylactic use of NA inhibitors.

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DAS181, a sialidase fusion protein, protects human airway epithelium against influenza virus infection: an in vitro pharmacodynamic analysis

Journal of Antimicrobial Chemotherapy ◽

10.1093/jac/dkp421 ◽

2009 ◽

Vol 65 (2) ◽

pp. 275-284 ◽

Cited By ~ 35

Author(s):

Gallen B. Triana-Baltzer ◽

Maria Babizki ◽

Michael C. W. Chan ◽

Adam C. N. Wong ◽

Laura M. Aschenbrenner ◽

...

Keyword(s):

Influenza Virus ◽

Virus Infection ◽

Fusion Protein ◽

Airway Epithelium ◽

Influenza Virus Infection ◽

Human Airway ◽

Human Airway Epithelium ◽

Pharmacodynamic Analysis

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Membrane-Tethered Mucin 1 is Stimulated by Interferon in Multiple Cell Types and Antagonizes Influenza A Virus Infection in Human Airway Epithelium

10.1101/2021.03.11.434997 ◽

2021 ◽

Cited By ~ 1

Author(s):

Ethan Iverson ◽

Kira Griswold ◽

Daniel Song ◽

Talita B. Gagliardi ◽

Kajal Hamidzadeh ◽

...

Keyword(s):

Influenza Virus ◽

Virus Infection ◽

Airway Epithelium ◽

Influenza A ◽

Influenza Virus Infection ◽

Human Monocyte ◽

Cell Types ◽

Mucin 1 ◽

Human Airway ◽

Human Airway Epithelium

AbstractInfluenza A virus (IAV) causes seasonal epidemics and periodic pandemics, resulting in significant morbidity and mortality in the human population. Tethered mucin 1 (MUC1) is highly expressed in airway epithelium, the primary site of IAV replication, and also by other cell types that influence IAV infection, including macrophages. MUC1 has the potential to influence infection dynamics through physical interactions and/or signaling activity, and recent work suggests MUC1 acts as a releasable decoy receptor and anti-inflammatory molecule during IAV infection. Still, the modulation of MUC1 and its impact during viral pathogenesis remains unclear. Thus, we sought to further investigate the interplay between MUC1 and IAV in an in vitro model of primary human airway epithelium (HAE). Our data indicate that a recombinant IAV hemagglutinin (H3) and H3N2 virus can bind endogenous HAE MUC1. We find that infection of HAE cultures with H1N1 or H3N2 IAV strains does not trigger enhanced MUC1 shedding, but instead stimulates an increase in cell-associated MUC1 protein. We observed a similar increase after stimulation with either type I or type III interferon (IFN); however, inhibition of IFN signaling during H1N1 infection only partially abrogated this increase, indicating multiple soluble factors contribute to MUC1 upregulation during the antiviral response. We expanded these findings and demonstrate that in addition to HAE, primary human monocyte-derived macrophages also upregulate MUC1 protein in response to both IFN treatment and conditioned media from IAV-infected HAE cultures. We then developed HAE genetically depleted for MUC1 to determine its impact on IAV pathogenesis, finding that MUC1 knock-out cultures exhibited enhanced viral growth compared to control cultures. Together, our data support a model whereby MUC1 antagonizes productive uptake of IAV in HAE. Infection then stimulates MUC1 expression on multiple cell types through IFN-dependent and -independent mechanisms that may further impact infection dynamics.Author SummaryThe mucosal surface of the respiratory epithelium is an important site of first contact for viral respiratory pathogens. Large and heavily glycosylated molecules known as tethered mucins extend from the cell surface and may physically restrict access to underlying cells. Recently, one of these tethered mucins, MUC1, has also been shown to influence cell signaling and inflammation. Still, despite its abundance in the airway and multifunctional capability, the role of MUC1 during influenza virus infection in the human respiratory tract remains unclear. Here, we demonstrate that influenza virus directly interacts with MUC1 in a physiologically-relevant model of human airway epithelium and find that MUC1 protein expression is elevated throughout the epithelium and in primary human monocyte-derived macrophages in response to important antiviral signals produced during infection. Using genetically-modified human airway cultures lacking MUC1, we then provide evidence of more efficient influenza virus infection in the absence of this mucin. Our data suggest that MUC1 not only physically restricts influenza virus uptake, but also represents a dynamic component of the host response that acts to further stem viral spread.

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Influenza Virus with Increased pH of Hemagglutinin Activation Has Improved Replication in Cell Culture but at the Cost of Infectivity in Human Airway Epithelium

Journal of Virology ◽

10.1128/jvi.00058-19 ◽

2019 ◽

Vol 93 (17) ◽

Cited By ~ 8

Author(s):

Anika Singanayagam ◽

Maria Zambon ◽

Wendy S. Barclay

Keyword(s):

Influenza Virus ◽

Host Cell ◽

Airway Epithelium ◽

Ph Sensitivity ◽

Influenza Viruses ◽

Ph Stability ◽

Target Cells ◽

Human Airway ◽

Human Airway Epithelium ◽

Virus Survival

ABSTRACT Pandemic H1N1 (pH1N1) influenza virus emerged from swine in 2009 with an adequate capability to infect and transmit between people. In subsequent years, it has circulated as a seasonal virus and evolved further human-adapting mutations. Mutations in the hemagglutinin (HA) stalk that increase pH stability have been associated with human adaptation and airborne transmission of pH1N1 virus. Yet, our understanding of how pH stability impacts virus-host interactions is incomplete. Here, using recombinant viruses with point mutations that alter the pH stability of pH1N1 HA, we found distinct effects on virus phenotypes in different experimental models. Increased pH sensitivity enabled viruses to uncoat in endosomes more efficiently, manifesting as increased replication rate in typical continuous cell cultures under single-cycle conditions. A more acid-labile HA also conferred a small reduction in sensitivity to antiviral therapeutics that act at the pH-sensitive HA fusion step. Conversely, in primary human airway epithelium cultured at the air-liquid interface, increased pH sensitivity attenuated multicycle viral replication by compromising virus survival in the extracellular microenvironment. In a mouse model of influenza pathogenicity, there was an optimum HA activation pH, and viruses with either more- or less-pH-stable HA were less virulent. Opposing pressures inside and outside the host cell that determine pH stability may influence zoonotic potential. The distinct effects that changes in pH stability exert on viral phenotypes underscore the importance of using the most appropriate systems for assessing virus titer and fitness, which has implications for vaccine manufacture, antiviral drug development, and pandemic risk assessment. IMPORTANCE The pH stability of the hemagglutinin surface protein varies between different influenza strains and subtypes and can affect the virus’ ability to replicate and transmit. Here, we demonstrate a delicate balance that the virus strikes within and without the target cell. We show that a pH-stable hemagglutinin enables a human influenza virus to replicate more effectively in human airway cells and mouse lungs by facilitating virus survival in the extracellular environment of the upper respiratory tract. Conversely, after entering target cells, being more pH stable confers a relative disadvantage, resulting in less efficient delivery of the viral genome to the host cell nucleus. Since the balance we describe will be affected differently in different host environments, it may restrict a virus’ ability to cross species. In addition, our findings imply that different influenza viruses may show variation in how well they are controlled by antiviral strategies targeting pH-dependent steps in the virus replication cycle.

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Human and avian influenza viruses target different cell types in cultures of human airway epithelium

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.0308001101 ◽

2004 ◽

Vol 101 (13) ◽

pp. 4620-4624 ◽

Cited By ~ 483

Author(s):

M. N. Matrosovich ◽

T. Y. Matrosovich ◽

T. Gray ◽

N. A. Roberts ◽

H.-D. Klenk

Keyword(s):

Avian Influenza ◽

Airway Epithelium ◽

Cell Types ◽

Influenza Viruses ◽

Avian Influenza Viruses ◽

Human Airway ◽

Human Airway Epithelium ◽

Different Cell Types

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Highly Pathogenic Avian Influenza Virus Infection of Mallards with Homo- and Heterosubtypic Immunity Induced by Low Pathogenic Avian Influenza Viruses

PLoS ONE ◽

10.1371/journal.pone.0006706 ◽

2009 ◽

Vol 4 (8) ◽

pp. e6706 ◽

Cited By ~ 76

Author(s):

Sasan R. Fereidouni ◽

Elke Starick ◽

Martin Beer ◽

Hendrik Wilking ◽

Donata Kalthoff ◽

...

Keyword(s):

Influenza Virus ◽

Avian Influenza ◽

Highly Pathogenic Avian Influenza ◽

Influenza Virus Infection ◽

Influenza Viruses ◽

Avian Influenza Viruses ◽

Highly Pathogenic ◽

Pathogenic Avian Influenza ◽

Heterosubtypic Immunity ◽

Pathogenic Avian Influenza Virus

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Constitutively Expressed IFITM3 Protein in Human Endothelial Cells Poses an Early Infection Block to Human Influenza Viruses

Journal of Virology ◽

10.1128/jvi.01254-16 ◽

2016 ◽

Vol 90 (24) ◽

pp. 11157-11167 ◽

Cited By ~ 15

Author(s):

Xiangjie Sun ◽

Hui Zeng ◽

Amrita Kumar ◽

Jessica A. Belser ◽

Taronna R. Maines ◽

...

Keyword(s):

Endothelial Cells ◽

Influenza Virus ◽

Virus Infection ◽

Avian Influenza ◽

Influenza Virus Infection ◽

Influenza Viruses ◽

Human Influenza ◽

Avian Influenza Viruses ◽

Human Influenza Virus ◽

Pulmonary Endothelial Cells

ABSTRACTA role for pulmonary endothelial cells in the orchestration of cytokine production and leukocyte recruitment during influenza virus infection, leading to severe lung damage, has been recently identified. As the mechanistic pathway for this ability is not fully known, we extended previous studies on influenza virus tropism in cultured human pulmonary endothelial cells. We found that a subset of avian influenza viruses, including potentially pandemic H5N1, H7N9, and H9N2 viruses, could infect human pulmonary endothelial cells (HULEC) with high efficiency compared to human H1N1 or H3N2 viruses. In HULEC, human influenza viruses were capable of binding to host cellular receptors, becoming internalized and initiating hemifusion but failing to uncoat the viral nucleocapsid and to replicate in host nuclei. Unlike numerous cell types, including epithelial cells, we found that pulmonary endothelial cells constitutively express a high level of the restriction protein IFITM3 in endosomal compartments. IFITM3 knockdown by small interfering RNA (siRNA) could partially rescue H1N1 virus infection in HULEC, suggesting IFITM3 proteins were involved in blocking human influenza virus infection in endothelial cells. In contrast, selected avian influenza viruses were able to escape IFITM3 restriction in endothelial cells, possibly by fusing in early endosomes at higher pH or by other, unknown mechanisms. Collectively, our study demonstrates that the human pulmonary endothelium possesses intrinsic immunity to human influenza viruses, in part due to the constitutive expression of IFITM3 proteins. Notably, certain avian influenza viruses have evolved to escape this restriction, possibly contributing to virus-induced pneumonia and severe lung disease in humans.IMPORTANCEAvian influenza viruses, including H5N1 and H7N9, have been associated with severe respiratory disease and fatal outcomes in humans. Although acute respiratory distress syndrome (ARDS) and progressive pulmonary endothelial damage are known to be present during severe human infections, the role of pulmonary endothelial cells in the pathogenesis of avian influenza virus infections is largely unknown. By comparing human seasonal influenza strains to avian influenza viruses, we provide greater insight into the interaction of influenza virus with human pulmonary endothelial cells. We show that human influenza virus infection is blocked during the early stages of virus entry, which is likely due to the relatively high expression of the host antiviral factors IFITMs (interferon-induced transmembrane proteins) located in membrane-bound compartments inside cells. Overall, this study provides a mechanism by which human endothelial cells limit replication of human influenza virus strains, whereas avian influenza viruses overcome these restriction factors in this cell type.

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