scholarly journals Influenza A viruses use multivalent sialic acid clusters for cell binding and receptor activation

2020 ◽  
Vol 16 (7) ◽  
pp. e1008656
Author(s):  
Christian Sieben ◽  
Erdinc Sezgin ◽  
Christian Eggeling ◽  
Suliana Manley
Keyword(s):  
Sialic Acid ◽  
Influenza A ◽  
Receptor Activation ◽  
Cell Binding ◽  
Influenza A Viruses

scholarly journals Influenza A viruses use multivalent sialic acid clusters for cell binding and receptor activation

2018 ◽  
Author(s):  
Christian Sieben ◽  
Erdinc Sezgin ◽  
Christian Eggeling ◽  
Suliana Manley
Keyword(s):  
Plasma Membrane ◽  
Sialic Acid ◽  
Influenza A ◽  
Super Resolution ◽  
Receptor Activation ◽  
Cell Binding ◽  
Virus Binding ◽  
Super Resolution Microscopy ◽  
Functional Receptor ◽  
Lateral Organization

AbstractInfluenza A virus (IAV) binds its host cell using the major viral surface protein hemagglutinin (HA). HA recognizes sialic acid, a plasma membrane glycan that functions as the specific primary attachment factor (AF). Since sialic acid alone cannot fulfill a signaling function, the virus needs to activate downstream factors to trigger endocytic uptake. Recently, the epidermal growth factor receptor (EGFR), a member of the receptor-tyrosine kinase family, was shown to be activated by and transmit IAV entry signals. However, how IAV engages and activates EGFR remains largely unclear.We used multicolor super-resolution microscopy to study the lateral organization of both IAV attachment factors and its functional receptor at the scale of the IAV particle. Intriguingly, quantitative cluster analysis revealed that AF and EGFR are organized in partially overlapping submicrometer clusters in the apical plasma membrane of A549 cells. Within AF domains, which are distinct from microvilli, the local AF concentration, a parameter that directly influences virus-cell binding, reaches on average 10-fold the background concentration and tends to increase towards the cluster center, thereby representing a multivalent virus-binding platform. Using our experimentally measured cluster characteristics, we simulated virus diffusion on a membrane, revealing that the distinct mobility pattern of IAVs is dominated by the local AF concentration, consistent with live cell single-virus tracking data. In contrast to AF, EGFR resides in clusters of rather low molecular density. Virus binding activates EGFR, but interestingly, this process occurs without a major lateral EGFR redistribution, instead relying on activation of pre-formed clusters, which we show are long-lived.Taken together, our results provide a quantitative understanding of the initial steps of influenza virus infection. Co-clustering of AF and EGFR permit a cooperative effect of binding and signaling at specific platforms, and thus we relate their spatial organization to their functional role during virus-cell binding and receptor activation.Author SummaryThe plasma membrane is the major interface between a cell and its environment. It is a complex and dynamic organelle that needs to protect as a barrier but also process subtle signals into and out of the cell. For IAV, an enveloped virus, it represents a major obstacle that it needs to overcome during infection as well as the site for the assembly of progeny virus particles. However, the organisation of the plasma membrane in particular the sites of virus interaction at the scale of an infecting particle (length scales < 100 nm) remains largely unknown.Sialic acids serve as IAV attachment factors but are not able to transmit signals across the plasma membrane. Receptor tyrosine kinases were identified to be activated upon virus binding and serve as functional receptor. How IAV engages and activates its functional receptors still remains speculative. Here we use super resolution microscopy to study the lateral organization as well as the functional relationship of plasma membrane-bound molecules involved in IAV infection. We find that molecules are organized in submicrometer nanodomains and, in combination with virus diffusion simulations, present a mechanistic view for how IAV first engages with AFs in the plasma membrane to then engage and trigger entry-associated membrane receptors.

scholarly journals N-Glycolylneuraminic Acid in Animal Models for Human Influenza A Virus

Viruses ◽  
2021 ◽  
Vol 13 (5) ◽  
pp. 815
Author(s):  
Cindy M. Spruit ◽  
Nikoloz Nemanichvili ◽  
Masatoshi Okamatsu ◽  
Hiromu Takematsu ◽  
Geert-Jan Boons ◽  
...  
Keyword(s):  
Sialic Acid ◽  
Animal Models ◽  
Influenza A ◽  
Influenza Virus Infection ◽  
Influenza Viruses ◽  
Upper Respiratory Tract ◽  
Human Influenza ◽  
Influenza A Viruses ◽  
Sialic Acid Content ◽  
Acetylneuraminic Acid

The first step in influenza virus infection is the binding of hemagglutinin to sialic acid-containing glycans present on the cell surface. Over 50 different sialic acid modifications are known, of which N-acetylneuraminic acid (Neu5Ac) and N-glycolylneuraminic acid (Neu5Gc) are the two main species. Animal models with α2,6 linked Neu5Ac in the upper respiratory tract, similar to humans, are preferred to enable and mimic infection with unadapted human influenza A viruses. Animal models that are currently most often used to study human influenza are mice and ferrets. Additionally, guinea pigs, cotton rats, Syrian hamsters, tree shrews, domestic swine, and non-human primates (macaques and marmosets) are discussed. The presence of NeuGc and the distribution of sialic acid linkages in the most commonly used models is summarized and experimentally determined. We also evaluated the role of Neu5Gc in infection using Neu5Gc binding viruses and cytidine monophosphate-N-acetylneuraminic acid hydroxylase (CMAH)-/- knockout mice, which lack Neu5Gc and concluded that Neu5Gc is unlikely to be a decoy receptor. This article provides a base for choosing an appropriate animal model. Although mice are one of the most favored models, they are hardly naturally susceptible to infection with human influenza viruses, possibly because they express mainly α2,3 linked sialic acids with both Neu5Ac and Neu5Gc modifications. We suggest using ferrets, which resemble humans closely in the sialic acid content, both in the linkages and the lack of Neu5Gc, lung organization, susceptibility, and disease pathogenesis.

scholarly journals Characterization of the Sialic Acid Binding Activity of Influenza A Viruses Using Soluble Variants of the H7 and H9 Hemagglutinins

PLoS ONE ◽  
2014 ◽  
Vol 9 (2) ◽  
pp. e89529 ◽  
Author(s):  
Anne-Kathrin Sauer ◽  
Chi-Hui Liang ◽  
Jürgen Stech ◽  
Ben Peeters ◽  
Pascale Quéré ◽  
...  
Keyword(s):  
Sialic Acid ◽  
Influenza A ◽  
Binding Activity ◽  
Influenza A Viruses ◽  
Sialic Acid Binding

scholarly journals Influenza Viruses in Mice: Deep Sequencing Analysis of Serial Passage and Effects of Sialic Acid Structural Variation

2019 ◽  
Vol 93 (23) ◽  
Author(s):  
Brian R. Wasik ◽  
Ian E. H. Voorhees ◽  
Karen N. Barnard ◽  
Brynn K. Alford-Lawrence ◽  
Wendy S. Weichert ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Sialic Acid ◽  
Influenza A Virus ◽  
Deep Sequencing ◽  
Influenza A ◽  
Natural Host ◽  
H1n1 Pandemic ◽  
Influenza A Viruses ◽  
Canine Influenza Virus ◽  
Canine Influenza

ABSTRACT Influenza A viruses have regularly jumped to new host species to cause epidemics or pandemics, an evolutionary process that involves variation in the viral traits necessary to overcome host barriers and facilitate transmission. Mice are not a natural host for influenza virus but are frequently used as models in studies of pathogenesis, often after multiple passages to achieve higher viral titers that result in clinical disease such as weight loss or death. Here, we examine the processes of influenza A virus infection and evolution in mice by comparing single nucleotide variations of a human H1N1 pandemic virus, a seasonal H3N2 virus, and an H3N2 canine influenza virus during experimental passage. We also compared replication and sequence variation in wild-type mice expressing N-glycolylneuraminic acid (Neu5Gc) with those seen in mice expressing only N-acetylneuraminic acid (Neu5Ac). Viruses derived from plasmids were propagated in MDCK cells and then passaged in mice up to four times. Full-genome deep sequencing of the plasmids, cultured viruses, and viruses from mice at various passages revealed only small numbers of mutational changes. The H3N2 canine influenza virus showed increases in frequency of sporadic mutations in the PB2, PA, and NA segments. The H1N1 pandemic virus grew well in mice, and while it exhibited the maintenance of some minority mutations, there was no clear evidence for adaptive evolution. The H3N2 seasonal virus did not establish in the mice. Finally, there were no clear sequence differences associated with the presence or absence of Neu5Gc. IMPORTANCE Mice are commonly used as a model to study the growth and virulence of influenza A viruses in mammals but are not a natural host and have distinct sialic acid receptor profiles compared to humans. Using experimental infections with different subtypes of influenza A virus derived from different hosts, we found that evolution of influenza A virus in mice did not necessarily proceed through the linear accumulation of host-adaptive mutations, that there was variation in the patterns of mutations detected in each repetition, and that the mutation dynamics depended on the virus examined. In addition, variation in the viral receptor, sialic acid, did not affect influenza virus evolution in this model. Overall, our results show that while mice provide a useful animal model for influenza virus pathology, host passage evolution will vary depending on the specific virus tested.

scholarly journals Sialic Acid Species as a Determinant of the Host Range of Influenza A Viruses

2000 ◽  
Vol 74 (24) ◽  
pp. 11825-11831 ◽  
Author(s):  
Yasuo Suzuki ◽  
Toshihiro Ito ◽  
Takashi Suzuki ◽  
Robert E. Holland ◽  
Thomas M. Chambers ◽  
...  
Keyword(s):  
Sialic Acid ◽  
Viral Replication ◽  
Influenza A ◽  
Lectin Binding ◽  
Influenza Viruses ◽  
Influenza A Viruses ◽  
Acetylneuraminic Acid ◽  
Human Viruses ◽  
Immunohistochemical Analyses

ABSTRACT The distribution of sialic acid (SA) species varies among animal species, but the biological role of this variation is largely unknown. Influenza viruses differ in their ability to recognize SA-galactose (Gal) linkages, depending on the animal hosts from which they are isolated. For example, human viruses preferentially recognize SA linked to Gal by the α2,6(SAα2,6Gal) linkage, while equine viruses favor SAα2,3Gal. However, whether a difference in relative abundance of specific SA species (N-acetylneuraminic acid [NeuAc] andN-glycolylneuraminic acid [NeuGc]) among different animals affects the replicative potential of influenza viruses is uncertain. We therefore examined the requirement for the hemagglutinin (HA) for support of viral replication in horses, using viruses whose HAs differ in receptor specificity. A virus with an HA recognizing NeuAcα2,6Gal but not NeuAcα2,3Gal or NeuGcα2,3Gal failed to replicate in horses, while one with an HA recognizing the NeuGcα2,3Gal moiety replicated in horses. Furthermore, biochemical and immunohistochemical analyses and a lectin-binding assay demonstrated the abundance of the NeuGcα2,3Gal moiety in epithelial cells of horse trachea, indicating that recognition of this moiety is critical for viral replication in horses. Thus, these results provide evidence of a biological effect of different SA species in different animals.

scholarly journals Cross-Reactive Neuraminidase-Inhibiting Antibodies Elicited by Immunization with Recombinant Neuraminidase Proteins of H5N1 and Pandemic H1N1 Influenza A Viruses

2015 ◽  
Vol 89 (14) ◽  
pp. 7224-7234 ◽  
Author(s):  
Wen-Chun Liu ◽  
Chia-Ying Lin ◽  
Yung-Ta Tsou ◽  
Jia-Tsrong Jan ◽  
Suh-Chin Wu
Keyword(s):  
Influenza Virus ◽  
Sialic Acid ◽  
Influenza Vaccine ◽  
Influenza A ◽  
Expression System ◽  
Influenza Viruses ◽  
Pandemic H1n1 ◽  
Influenza A Viruses ◽  
Homologous Virus ◽  
Universal Influenza Vaccine

ABSTRACTNeuraminidase (NA), an influenza virus envelope glycoprotein, removes sialic acid from receptors for virus release from infected cells. For this study, we used a baculovirus-insect cell expression system to construct and purify recombinant NA (rNA) proteins of H5N1 (A/Vietnam/1203/2004) and pandemic H1N1 (pH1N1) (A/Texas/05/2009) influenza viruses. BALB/c mice immunized with these proteins had high titers of NA-specific IgG and NA-inhibiting (NI) antibodies against H5N1, pH1N1, H3N2, and H7N9 viruses. H5N1 rNA immunization resulted in higher quantities of NA-specific antibody-secreting B cells against H5N1 and heterologous pH1N1 viruses in the spleen. H5N1 rNA and pH1N1 rNA immunizations both provided complete protection against homologous virus challenges, with H5N1 rNA immunization providing better protection against pH1N1 virus challenges. Cross-reactive NI antibodies were further dissected via pH1N1 rNA protein immunizations with I149V (NA with a change of Ile to Val at position 149), N344Y, and I365T/S366N NA mutations. The I365T/S366N mutation of pH1N1 rNA enhanced cross-reactive NI antibodies against H5N1, H3N2, and H7N9 viruses. It is our hope that these findings provide useful information for the development of an NA-based universal influenza vaccine.IMPORTANCENeuraminidase (NA) is an influenza virus enzymatic protein that cleaves sialic acid linkages on infected cell surfaces, thus facilitating viral release and contributing to viral transmission and mucus infection. In currently available inactivated or live, attenuated influenza vaccines based on the antigenic content of hemagglutinin proteins, vaccine efficacy can be contributed partly through NA-elicited immune responses. We investigated the NA immunity of different recombinant NA (rNA) proteins associated with pH1N1 and H5N1 viruses. Our results indicate that H5N1 rNA immunization induced more potent cross-protective immunity than pH1N1 rNA immunization, and three mutated residues, I149V, I365T, and S366N, near the NA enzyme active site(s) are linked to enhanced cross-reactive NA-inhibiting antibodies against heterologous and heterosubtypic influenza A viruses. These findings provide useful information for the development of an NA-based universal influenza vaccine.

scholarly journals Influenza binds phosphorylated glycans from human lung

2019 ◽  
Vol 5 (2) ◽  
pp. eaav2554 ◽  
Author(s):  
Lauren Byrd-Leotis ◽  
Nan Jia ◽  
Sucharita Dutta ◽  
Jessica F. Trost ◽  
Chao Gao ◽  
...  
Keyword(s):  
Sialic Acid ◽  
Influenza A ◽  
Human Lung ◽  
Species Specificity ◽  
Influenza Viruses ◽  
Surface Markers ◽  
Influenza A Viruses ◽  
Synthetic Receptor ◽  
Glycan Receptors ◽  
Sialic Acid Linkage

Influenza A viruses can bind sialic acid–terminating glycan receptors, and species specificity is often correlated with sialic acid linkage with avian strains recognizing α2,3-linked sialylated glycans and mammalian strains preferring α2,6-linked sialylated glycans. These paradigms derive primarily from studies involving erythrocyte agglutination, binding to synthetic receptor analogs or binding to undefined surface markers on cells or tissues. Here, we present the first examination of the N-glycome of the human lung for identifying natural receptors for a range of avian and mammalian influenza viruses. We found that the human lung contains many α2,3- and α2,6-linked sialylated glycan determinants bound by virus, but all viruses also bound to phosphorylated, nonsialylated glycans.

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