Competitive Cooperation of Hemagglutinin and Neuraminidase during Influenza A Virus Entry

The hemagglutinin (HA) and neuraminidase (NA) of influenza A virus possess antagonistic activities on interaction with sialic acid (SA), which is the receptor for virus attachment. HA binds SA through its receptor-binding sites, while NA is a receptor-destroying enzyme by removing SAs. The function of HA during virus entry has been extensively investigated, however, examination of NA has long been focused to its role in the exit of progeny virus from infected cells, and the role of NA in the entry process is still under-appreciated. This review summarizes the current understanding of the roles of HA and NA in relation to each other during virus entry.

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Neurotropic influenza A virus infection causes prion protein misfolding into infectious prions in neuroblastoma cells

Scientific Reports ◽

10.1038/s41598-021-89586-6 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Hideyuki Hara ◽

Junji Chida ◽

Keiji Uchiyama ◽

Agriani Dini Pasiana ◽

Etsuhisa Takahashi ◽

...

Keyword(s):

Prion Protein ◽

Influenza A Virus ◽

Protein Misfolding ◽

Influenza A ◽

Prion Diseases ◽

Neuroblastoma Cells ◽

Cellular Prion Protein ◽

Infected Cells ◽

Hit And Run

AbstractMisfolding of the cellular prion protein, PrPC, into the amyloidogenic isoform, PrPSc, which forms infectious protein aggregates, the so-called prions, is a key pathogenic event in prion diseases. No pathogens other than prions have been identified to induce misfolding of PrPC into PrPSc and propagate infectious prions in infected cells. Here, we found that infection with a neurotropic influenza A virus strain (IAV/WSN) caused misfolding of PrPC into PrPSc and generated infectious prions in mouse neuroblastoma cells through a hit-and-run mechanism. The structural and biochemical characteristics of IAV/WSN-induced PrPSc were different from those of RML and 22L laboratory prions-evoked PrPSc, and the pathogenicity of IAV/WSN-induced prions were also different from that of RML and 22L prions, suggesting IAV/WSN-specific formation of PrPSc and infectious prions. Our current results may open a new avenue for the role of viral infection in misfolding of PrPC into PrPSc and formation of infectious prions.

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Host Lipid Rafts Play a Major Role in Binding and Endocytosis of Influenza A Virus

Viruses ◽

10.3390/v10110650 ◽

2018 ◽

Vol 10 (11) ◽

pp. 650 ◽

Cited By ~ 17

Author(s):

Dileep Verma ◽

Dinesh Gupta ◽

Sunil Lal

Keyword(s):

Lipid Rafts ◽

Influenza A Virus ◽

Lipid Raft ◽

Influenza A ◽

Virus Entry ◽

Critical Role ◽

Ganglioside Gm1 ◽

Raft Fraction ◽

Infected Cells ◽

Host Cell Surface

Influenza still remains one of the most challenging diseases, posing a significant threat to public health. Host lipid rafts play a critical role in influenza A virus (IAV) assembly and budding, however, their role in polyvalent IAV host binding and endocytosis had remained elusive until now. In the present study, we observed co-localization of IAV with a lipid raft marker ganglioside, GM1, on the host surface. Further, we isolated the lipid raft micro-domains from IAV infected cells and detected IAV protein in the raft fraction. Finally, raft disruption using Methyl-β-Cyclodextrin revealed significant reduction in IAV host binding, suggesting utilization of host rafts for polyvalent binding on the host cell surface. In addition to this, cyclodextrin mediated inhibition of raft-dependent endocytosis showed significantly reduced IAV internalization. Interestingly, exposure of cells to cyclodextrin two hours post-IAV binding showed no such reduction in IAV entry, indicating use of raft-dependent endocytosis for host entry. In summary, this study demonstrates that host lipid rafts are selected by IAV as a host attachment factors for multivalent binding, and IAV utilizes these micro-domains to exploit raft-dependent endocytosis for host internalization, a virus entry route previously unknown for IAV.

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Substrate Binding by the Second Sialic Acid-Binding Site of Influenza A Virus N1 Neuraminidase Contributes to Enzymatic Activity

Journal of Virology ◽

10.1128/jvi.01243-18 ◽

2018 ◽

Vol 92 (20) ◽

Cited By ~ 9

Author(s):

Wenjuan Du ◽

Meiling Dai ◽

Zeshi Li ◽

Geert-Jan Boons ◽

Ben Peeters ◽

...

Keyword(s):

Sialic Acid ◽

Enzymatic Activity ◽

Binding Site ◽

Influenza A Virus ◽

Influenza A ◽

Substrate Binding ◽

Host Tropism ◽

Contact Residues ◽

Human Viruses

ABSTRACTThe influenza A virus (IAV) neuraminidase (NA) protein plays an essential role in the release of virus particles from cells and decoy receptors. The NA enzymatic activity presumably needs to match the activity of the IAV hemagglutinin (HA) attachment protein and the host sialic acid (SIA) receptor repertoire. We analyzed the enzymatic activities of N1 NA proteins derived from avian (H5N1) and human (H1N1) IAVs and analyzed the role of the second SIA-binding site, located adjacent to the conserved catalytic site, therein. SIA contact residues in the second SIA-binding site of NA are highly conserved in avian, but not human, IAVs. All N1 proteins preferred cleaving α2,3- over α2,6-linked SIAs even when their corresponding HA proteins displayed a strict preference for α2,6-linked SIAs, indicating that the specificity of the NA protein does not need to fully match that of the corresponding HA protein. NA activity was affected by substitutions in the second SIA-binding site that are observed in avian and human IAVs, at least when multivalent rather than monovalent substrates were used. These mutations included both SIA contact residues and residues that do not directly interact with SIA in all three loops of the second SIA-binding site. Substrate binding via the second SIA-binding site enhanced the catalytic activity of N1. Mutation of the second SIA-binding site was also shown to affect virus replicationin vitro. Our results indicate an important role for the N1 second SIA-binding site in binding to and cleavage of multivalent substrates.IMPORTANCEAvian and human influenza A viruses (IAVs) preferentially bind α2,3- and α2,6-linked sialic acids (SIAs), respectively. A functional balance between the hemagglutinin (HA) attachment and neuraminidase (NA) proteins is thought to be important for host tropism. What this balance entails at the molecular level is, however, not well understood. We now show that N1 proteins of both avian and human viruses prefer cleaving avian- over human-type receptors although human viruses were relatively better in cleavage of the human-type receptors. In addition, we show that substitutions at different positions in the second SIA-binding site found in NA proteins of human IAVs have a profound effect on binding and cleavage of multivalent, but not monovalent, receptors and affect virus replication. Our results indicate that the HA-NA balance can be tuned via modification of substrate binding via this site and suggest an important role of the second SIA-binding site in host tropism.

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Attachment of bovine parvovirus to sialic acids on bovine cell membranes

Journal of General Virology ◽

10.1099/vir.0.79899-0 ◽

2004 ◽

Vol 85 (8) ◽

pp. 2199-2207 ◽

Cited By ~ 21

Author(s):

F. Brent Johnson ◽

Laura B. Fenn ◽

Thomas J. Owens ◽

Laura J. Faucheux ◽

Shawn D. Blackburn

Keyword(s):

Sialic Acid ◽

Binding Sites ◽

Influenza A ◽

Cell Membranes ◽

Lung Fibroblasts ◽

Glycophorin A ◽

Kidney Cells ◽

Virus Binding ◽

Virus Attachment ◽

Bovine Parvovirus

Although it has previously been shown that bovine parvovirus (BPV) attaches to the sialated glycoprotein glycophorin A on erythrocytes, the nature of virus-binding moieties on mammalian nucleated cells is less clear. Buffalo lung fibroblasts (Bu), primary bovine embryonic kidney cells, Madin–Darby bovine kidney cells and bovine embryonic trachea (EBTr) cells were assessed for molecules capable of binding BPV. Competition studies were carried out on both erythrocyte and nucleated cell targets using a variety of sialated compounds and sialic acid-negative compounds. Glycophorin A was found to inhibit BPV binding, while mucin exhibited low-level inhibition. These two sialated compounds also blocked attachment of BPV-modified microsphere carriers to the Bu cell membrane. Influenza A virus was used as a sialic acid competitor and interfered with BPV attachment to erythrocytes and replication in Bu cells. Significantly, the enzyme sialidase removed BPV-binding sites from Bu and EBTr cells. The binding sites could be reconstituted on sialidase-treated cells by the enzymes α-2,3-O-sialyltransferase and α-2,3-N-sialyltransferase. These results indicated that BPV can attach to sialic acid on cell membranes and that the sialylglycoproteins available for virus attachment appear to contain both N- and O-linked carbohydrate moieties, but that not all members of the sialic acid family can bind BPV. Moreover, there may be other moieties that can bind BPV, which may act as either primary or secondary receptors.

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The Functional Role of Loops and Flanking Sequences of G-Quadruplex Aptamer to the Hemagglutinin of Influenza a Virus

International Journal of Molecular Sciences ◽

10.3390/ijms22052409 ◽

2021 ◽

Vol 22 (5) ◽

pp. 2409

Author(s):

Anastasia A. Bizyaeva ◽

Dmitry A. Bunin ◽

Valeria L. Moiseenko ◽

Alexandra S. Gambaryan ◽

Sonja Balk ◽

...

Keyword(s):

Influenza A Virus ◽

Influenza A ◽

Core Structure ◽

Dna Aptamer ◽

Tertiary Structures ◽

Quadruplex Dna ◽

G Quadruplex ◽

Flanking Sequences ◽

Related Proteins

Nucleic acid aptamers are generally accepted as promising elements for the specific and high-affinity binding of various biomolecules. It has been shown for a number of aptamers that the complexes with several related proteins may possess a similar affinity. An outstanding example is the G-quadruplex DNA aptamer RHA0385, which binds to the hemagglutinins of various influenza A virus strains. These hemagglutinins have homologous tertiary structures but moderate-to-low amino acid sequence identities. Here, the experiment was inverted, targeting the same protein using a set of related, parallel G-quadruplexes. The 5′- and 3′-flanking sequences of RHA0385 were truncated to yield parallel G-quadruplex with three propeller loops that were 7, 1, and 1 nucleotides in length. Next, a set of minimal, parallel G-quadruplexes with three single-nucleotide loops was tested. These G-quadruplexes were characterized both structurally and functionally. All parallel G-quadruplexes had affinities for both recombinant hemagglutinin and influenza virions. In summary, the parallel G-quadruplex represents a minimal core structure with functional activity that binds influenza A hemagglutinin. The flanking sequences and loops represent additional features that can be used to modulate the affinity. Thus, the RHA0385–hemagglutinin complex serves as an excellent example of the hypothesis of a core structure that is decorated with additional recognizing elements capable of improving the binding properties of the aptamer.

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Effects of Basic Amino Acids and Their Derivatives on SARS-CoV-2 and Influenza-A Virus Infection

Viruses ◽

10.3390/v13071301 ◽

2021 ◽

Vol 13 (7) ◽

pp. 1301

Author(s):

Ivonne Melano ◽

Li-Lan Kuo ◽

Yan-Chung Lo ◽

Po-Wei Sung ◽

Ni Tien ◽

...

Keyword(s):

Amino Acids ◽

Virus Infection ◽

Influenza A Virus ◽

Influenza A ◽

Enveloped Viruses ◽

Virus Attachment ◽

Basic Amino Acids ◽

Ester Derivative ◽

Endosomal Acidification

Amino acids have been implicated with virus infection and replication. Here, we demonstrate the effects of two basic amino acids, arginine and lysine, and their ester derivatives on infection of two enveloped viruses, SARS-CoV-2, and influenza A virus. We found that lysine and its ester derivative can efficiently block infection of both viruses in vitro. Furthermore, the arginine ester derivative caused a significant boost in virus infection. Studies on their mechanism of action revealed that the compounds potentially disturb virus uncoating rather than virus attachment and endosomal acidification. Our findings suggest that lysine supplementation and the reduction of arginine-rich food intake can be considered as prophylactic and therapeutic regimens against these viruses while also providing a paradigm for the development of broad-spectrum antivirals.

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Potent sialic acid inhibitors that target influenza A virus hemagglutinin

Scientific Reports ◽

10.1038/s41598-021-87845-0 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Yu-Jen Chang ◽

Cheng-Yun Yeh ◽

Ju-Chien Cheng ◽

Yu-Qi Huang ◽

Kai-Cheng Hsu ◽

...

Keyword(s):

Sialic Acid ◽

Antiviral Activity ◽

Binding Site ◽

Influenza A Virus ◽

Receptor Binding ◽

Influenza A ◽

The United States ◽

Ligand Complex ◽

Receptor Binding Site ◽

Computational Strategy

AbstractEradicating influenza A virus (IAV) is difficult, due to its genetic drift and reassortment ability. As the infectious cycle is initiated by the influenza glycoprotein, hemagglutinin (HA), which mediates the binding of virions to terminal sialic acids moieties, HA is a tempting target of anti-influenza inhibitors. However, the complexity of the HA structure has prevented delineation of the structural characterization of the HA protein–ligand complex. Our computational strategy efficiently analyzed > 200,000 records of compounds held in the United States National Cancer Institute (NCI) database and identified potential HA inhibitors, by modeling the sialic acid (SA) receptor binding site (RBS) for the HA structure. Our modeling revealed that compound NSC85561 showed significant antiviral activity against the IAV H1N1 strain with EC50 values ranging from 2.31 to 2.53 µM and negligible cytotoxicity (CC50 > 700 µM). Using the NSC85561 compound as the template to generate 12 derivatives, robust bioassay results revealed the strongest antiviral efficacies with NSC47715 and NSC7223. Virtual screening clearly identified three SA receptor binding site inhibitors that were successfully validated in experimental data. Thus, our computational strategy has identified SA receptor binding site inhibitors against HA that show IAV-associated antiviral activity.

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Dicer is involved in protection against influenza A virus infection

Journal of General Virology ◽

10.1099/vir.0.83103-0 ◽

2007 ◽

Vol 88 (10) ◽

pp. 2627-2635 ◽

Cited By ~ 80

Author(s):

Alexey A. Matskevich ◽

Karin Moelling

Keyword(s):

Virus Infection ◽

Influenza A Virus ◽

Mammalian Cells ◽

Influenza A ◽

Virus Production ◽

Vero Cells ◽

Antiviral Response ◽

Protein Levels ◽

Infected Cells ◽

Influenza A Virus Infection

In mammals the interferon (IFN) system is a central innate antiviral defence mechanism, while the involvement of RNA interference (RNAi) in antiviral response against RNA viruses is uncertain. Here, we tested whether RNAi is involved in the antiviral response in mammalian cells. To investigate the role of RNAi in influenza A virus-infected cells in the absence of IFN, we used Vero cells that lack IFN-α and IFN-β genes. Our results demonstrate that knockdown of a key RNAi component, Dicer, led to a modest increase of virus production and accelerated apoptosis of influenza A virus-infected cells. These effects were much weaker in the presence of IFN. The results also show that in both Vero cells and the IFN-producing alveolar epithelial A549 cell line influenza A virus targets Dicer at mRNA and protein levels. Thus, RNAi is involved in antiviral response, and Dicer is important for protection against influenza A virus infection.

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Influenza A Virus Replication Induces Cell Cycle Arrest in G0/G1 Phase

Journal of Virology ◽

10.1128/jvi.01216-10 ◽

2010 ◽

Vol 84 (24) ◽

pp. 12832-12840 ◽

Cited By ~ 85

Author(s):

Yuan He ◽

Ke Xu ◽

Bjoern Keiner ◽

Jianfang Zhou ◽

Volker Czudai ◽

...

Keyword(s):

Cell Cycle ◽

Cell Cycle Arrest ◽

Influenza A Virus ◽

Influenza A ◽

Cell Cycle Progression ◽

Virus Production ◽

Phase Cell ◽

Cycle Progression ◽

Cycle Arrest ◽

Infected Cells

ABSTRACT Many viruses interact with the host cell division cycle to favor their own growth. In this study, we examined the ability of influenza A virus to manipulate cell cycle progression. Our results show that influenza A virus A/WSN/33 (H1N1) replication results in G0/G1-phase accumulation of infected cells and that this accumulation is caused by the prevention of cell cycle entry from G0/G1 phase into S phase. Consistent with the G0/G1-phase accumulation, the amount of hyperphosphorylated retinoblastoma protein, a necessary active form for cell cycle progression through late G1 into S phase, decreased after infection with A/WSN/33 (H1N1) virus. In addition, other key molecules in the regulation of the cell cycle, such as p21, cyclin E, and cyclin D1, were also changed and showed a pattern of G0/G1-phase cell cycle arrest. It is interesting that increased viral protein expression and progeny virus production in cells synchronized in the G0/G1 phase were observed compared to those in either unsynchronized cells or cells synchronized in the G2/M phase. G0/G1-phase cell cycle arrest is likely a common strategy, since the effect was also observed in other strains, such as H3N2, H9N2, PR8 H1N1, and pandemic swine H1N1 viruses. These findings, in all, suggest that influenza A virus may provide favorable conditions for viral protein accumulation and virus production by inducing a G0/G1-phase cell cycle arrest in infected cells.

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