scholarly journals The dynamic proteome of influenza A virus infection identifies M segment splicing as a host range determinant

2018 ◽  
Author(s):  
Boris Bogdanow ◽  
Katrin Eichelbaum ◽  
Anne Sadewasser ◽  
Xi Wang ◽  
Immanuel Husic ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Host Range ◽  
Influenza A Virus ◽  
Rna Splicing ◽  
Influenza A ◽  
Matrix Protein ◽  
Shotgun Proteomics ◽  
Influenza Viruses ◽  
Species Barrier ◽  
Viral Protein Synthesis

SUMMARYA century ago, influenza A virus (IAV) infection caused the 1918 flu pandemic and killed an estimated 20-40 million people. Pandemic IAV outbreaks occur when strains from animal reservoirs acquire the ability to infect and spread among humans. The molecular details of this species barrier are incompletely understood. We combined metabolic pulse labeling and quantitative shotgun proteomics to globally monitor protein synthesis upon infection of human cells with a human-and a bird-adapted IAV strain. While production of host proteins was remarkably similar, we observed striking differences in the kinetics of viral protein synthesis over the course of infection. Most importantly, the matrix protein M1 was inefficiently produced by the bird-adapted strain at later stages. We show that impaired production of M1 from bird-adapted strains is caused by increased splicing of the M segment RNA to alternative isoforms. Experiments with reporter constructs and recombinant influenza viruses revealed that strain-specific M segment splicing is controlled by the 3’ splice site and functionally important for permissive infection. Independentin silicoevidence shows that avian-adapted M segments have evolved different conserved RNA structure features than human-adapted sequences. Thus, our data identifies M segment RNA splicing as a viral determinant of host range.

scholarly journals The dynamic proteome of influenza A virus infection identifies M segment splicing as a host range determinant

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Boris Bogdanow ◽  
Xi Wang ◽  
Katrin Eichelbaum ◽  
Anne Sadewasser ◽  
Immanuel Husic ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Host Range ◽  
Influenza A Virus ◽  
Rna Splicing ◽  
Rna Structure ◽  
Influenza A ◽  
Matrix Protein ◽  
Species Barrier ◽  
Viral Protein Synthesis ◽  
The Matrix

AbstractPandemic influenza A virus (IAV) outbreaks occur when strains from animal reservoirs acquire the ability to infect and spread among humans. The molecular basis of this species barrier is incompletely understood. Here we combine metabolic pulse labeling and quantitative proteomics to monitor protein synthesis upon infection of human cells with a human- and a bird-adapted IAV strain and observe striking differences in viral protein synthesis. Most importantly, the matrix protein M1 is inefficiently produced by the bird-adapted strain. We show that impaired production of M1 from bird-adapted strains is caused by increased splicing of the M segment RNA to alternative isoforms. Strain-specific M segment splicing is controlled by the 3′ splice site and functionally important for permissive infection. In silico and biochemical evidence shows that avian-adapted M segments have evolved different conserved RNA structure features than human-adapted sequences. Thus, we identify M segment RNA splicing as a viral host range determinant.

scholarly journals Antiviral Activity of Benzoic Acid Derivative NC-5 Against Influenza A Virus and Its Neuraminidase Inhibition

2019 ◽  
Vol 20 (24) ◽  
pp. 6261
Author(s):  
Min Guo ◽  
Jiawei Ni ◽  
Jie Yu ◽  
Jing Jin ◽  
Lingman Ma ◽  
...  
Keyword(s):  
Antiviral Activity ◽  
Benzoic Acid ◽  
Influenza A Virus ◽  
Influenza A ◽  
Matrix Protein ◽  
Influenza Viruses ◽  
Drug Candidate ◽  
Dependent Manner ◽  
Drug Resistant

The currently available drugs against influenza A virus primarily target neuraminidase (NA) or the matrix protein 2 (M2) ion channel. The emergence of drug-resistant viruses requires the development of new antiviral chemicals. Our study applied a cell-based approach to evaluate the antiviral activity of a series of newly synthesized benzoic acid derivatives, and 4-(2,2-Bis(hydroxymethyl)-5-oxopyrrolidin-l-yl)-3-(5-cyclohexyl-4H-1,2,4-triazol-3-yl)amino). benzoic acid, termed NC-5, was found to possess antiviral activity. NC-5 inhibited influenza A viruses A/FM/1/47 (H1N1), A/Beijing/32/92 (H3N2) and oseltamivir-resistant mutant A/FM/1/47-H275Y (H1N1-H275Y) in a dose-dependent manner. The 50% effective concentrations (EC50) for H1N1 and H1N1-H275Y were 33.6 μM and 32.8 μM, respectively, which showed that NC-5 had a great advantage over oseltamivir in drug-resistant virus infections. The 50% cytotoxic concentration (CC50) of NC-5 was greater than 640 μM. Orally administered NC-5 protected mice infected with H1N1 and H1N1-H275Y, conferring 80% and 60% survival at 100 mg/kg/d, reducing body weight loss, and alleviating virus-induced lung injury. NC-5 could suppress NP and M1 protein expression levels during the late stages of viral biosynthesis and inhibit NA activity, which may influence virus release. Our study proved that NC-5 has potent anti-influenza activity in vivo and in vitro, meaning that it could be regarded as a promising drug candidate to treat infection with influenza viruses, including oseltamivir-resistant viruses.

scholarly journals Influenza A Virus Strains Differ in Sensitivity to the Antiviral Action of Mx-GTPase

2008 ◽  
Vol 82 (7) ◽  
pp. 3624-3631 ◽  
Author(s):  
Jan Dittmann ◽  
Silke Stertz ◽  
Daniel Grimm ◽  
John Steel ◽  
Adolfo García-Sastre ◽  
...  
Keyword(s):  
Influenza A Virus ◽  
Influenza A ◽  
Influenza Viruses ◽  
Host Responses ◽  
Influenza A Viruses ◽  
Species Barrier ◽  
Spanish Flu ◽  
Avian Origin ◽  
Flu Virus ◽  
Virus Strains

ABSTRACT Interferon-mediated host responses are of great importance for controlling influenza A virus infections. It is well established that the interferon-induced Mx proteins possess powerful antiviral activities toward most influenza viruses. Here we analyzed a range of influenza A virus strains for their sensitivities to murine Mx1 and human MxA proteins and found remarkable differences. Virus strains of avian origin were highly sensitive to Mx1, whereas strains of human origin showed much weaker responses. Artificial reassortments of the viral components in a minireplicon system identified the viral nucleoprotein as the main target structure of Mx1. Interestingly, the recently reconstructed 1918 H1N1 “Spanish flu” virus was much less sensitive than the highly pathogenic avian H5N1 strain A/Vietnam/1203/04 when tested in a minireplicon system. Importantly, the human 1918 virus-based minireplicon system was almost insensitive to inhibition by human MxA, whereas the avian influenza A virus H5N1-derived system was well controlled by MxA. These findings suggest that Mx proteins provide a formidable hurdle that hinders influenza A viruses of avian origin from crossing the species barrier to humans. They further imply that the observed insensitivity of the 1918 virus-based replicon to the antiviral activity of human MxA is a hitherto unrecognized characteristic of the “Spanish flu” virus that may contribute to the high virulence of this unusual pandemic strain.

scholarly journals Stress granule-inducing eukaryotic translation initiation factor 4A inhibitors block influenza A virus replication

2017 ◽  
Author(s):  
Patrick D. Slaine ◽  
Mariel Kleer ◽  
Nathan Smith ◽  
Denys A. Khaperskyy ◽  
Craig McCormick
Keyword(s):  
Protein Synthesis ◽  
Influenza A Virus ◽  
Viral Protein ◽  
Influenza A ◽  
Translation Initiation Factor ◽  
Host Protein ◽  
Initiation Factor ◽  
Viral Protein Synthesis ◽  
Infected Cells ◽  
Pateamine A

ABSTRACTEukaryotic translation initiation factor 4A (eIF4A) is a helicase that facilitates assembly of the translation preinitiation complex by unwinding structured mRNA 5’ untranslated regions. Pateamine A (PatA) and silvestrol are natural products that disrupt eIF4A function and arrest translation, thereby triggering the formation of cytoplasmic aggregates of stalled preinitiation complexes known as stress granules (SGs). Here we examined the effects of eIF4A inhibition by PatA and silvestrol on influenza A virus (IAV) protein synthesis and replication in cell culture. Treatment of infected cells with either PatA or silvestrol at early times post-infection results in SG formation, arrest of viral protein synthesis and failure to replicate the viral genome. PatA, which irreversibly binds to eIF4A, sustained long-term blockade of IAV replication following drug withdrawal, and inhibited IAV replication at concentrations that had minimal cytotoxicity. By contrast, the antiviral effects of silvestrol were fully reversible; drug withdrawal caused rapid SG dissolution and resumption of viral protein synthesis. IAV inhibition by silvestrol was invariably associated with cytotoxicity. PatA blocked replication of genetically divergent IAV strains, suggesting common dependence on host eIF4A activity. This study demonstrates the feasibility of targeting core host protein synthesis machinery to prevent viral replication.IMPORTANCEInfluenza A virus (IAV) relies on cellular protein synthesis to decode viral messenger RNAs. Pateamine A and silvestrol are natural products that inactivate an essential protein synthesis protein known as eIF4A. Here we show that IAV is sensitive to these eIF4A inhibitor drugs. Treatment of infected cells with pateamine A or silvestrol prevented synthesis of viral proteins, viral genome replication and release of infectious virions. The irreversible eIF4A inhibitor pateamine A sustained long-term blockade of viral replication, whereas viral protein synthesis quickly resumed after silvestrol was removed from infected cells. Prolonged incubation of either infected or uninfected cells with these drugs induced the programmed cell death cascade called apoptosis. Our findings suggest that core components of the host protein synthesis machinery are viable targets for antiviral drug discovery. The most promising drug candidates should selectively block protein synthesis in infected cells without perturbing bystander uninfected cells.

scholarly journals Isolation and Characterization of a Distinct Influenza A Virus from Egyptian Bats

2018 ◽  
Vol 93 (2) ◽  
Author(s):  
Ahmed Kandeil ◽  
Mokhtar R. Gomaa ◽  
Mahmoud M. Shehata ◽  
Ahmed N. El Taweel ◽  
Sara H. Mahmoud ◽  
...  
Keyword(s):  
Influenza A Virus ◽  
Influenza A ◽  
Mdck Cells ◽  
Genomic Analysis ◽  
Cross Reactivity ◽  
Influenza Viruses ◽  
Avian Host ◽  
Influenza A Viruses ◽  
Species Barrier ◽  
Isolation And Characterization

ABSTRACT Recently, two genetically distinct influenza viruses were detected in bats in Guatemala and Peru. We conducted influenza A virus surveillance among four bat species in Egypt. Out of 1,202 swab specimens, 105 were positive by real-time PCR. A virus was successfully isolated in eggs and propagated in MDCK cells in the presence of N-tosyl-l-phenylalanine chloromethyl ketone-treated trypsin. Genomic analysis revealed that the virus was phylogenetically distinct from all other influenza A viruses. Analysis of the hemagglutinin gene suggested a common ancestry with other H9 viruses, and the virus showed a low level of cross-reactivity with serum raised against H9N2 viruses. Bats were seropositive for the isolated viruses. The virus replicated in the lungs of experimentally infected mice. While it is genetically distinct, this virus shares several avian influenza virus characteristics suggesting a more recent avian host origin. IMPORTANCE Through surveillance, we isolated and characterized an influenza A virus from Egyptian fruit bats. This virus had an affinity to avian-like receptors but was also able to infect mice. Our findings indicate that bats may harbor a diversity of influenza A viruses. Such viruses may have the potential to cross the species barrier to infect other species, including domestic birds, mammals, and, possibly, humans.

scholarly journals RNase L limits host and viral protein synthesis via inhibition of mRNA export

2021 ◽  
Author(s):  
James M. Burke ◽  
Alison R. Gilchrist ◽  
Sara L. Sawyer ◽  
Roy Parker
Keyword(s):  
Protein Synthesis ◽  
Influenza A Virus ◽  
Viral Protein ◽  
Influenza A ◽  
Cytokine Production ◽  
Mrna Export ◽  
Virus Protein ◽  
Rnase L ◽  
Viral Mrna ◽  
Viral Protein Synthesis

AbstractRNase L is widely thought to limit viral protein synthesis by cleaving host rRNA and viral mRNA, resulting in translation arrest and viral mRNA degradation. Herein, we show that the mRNAs of dengue virus and influenza A virus largely escape RNase L-mediated mRNA decay, and this permits viral protein production. However, activation of RNase L arrests nuclear mRNA export, which strongly inhibits influenza A virus protein synthesis and reduces cytokine production. Importantly, the heterogeneous and temporal nature of the mRNA export block in individual cells permits sufficient production of antiviral cytokines from transcriptionally induced host mRNAs. This defines RNase L-mediated arrest of mRNA export as a key antiviral shutoff and cytokine regulatory pathway.One Sentence SummaryRNase L-mediated shutoff of nuclear mRNA export limits viral protein synthesis and regulates antiviral cytokine production.

scholarly journals Inhibition of Ongoing Influenza A Virus Replication Reveals Different Mechanisms of RIG-I Activation

2019 ◽  
Vol 93 (6) ◽  
Author(s):  
GuanQun Liu ◽  
Yao Lu ◽  
Qiang Liu ◽  
Yan Zhou
Keyword(s):  
Protein Synthesis ◽  
Viral Replication ◽  
Influenza A Virus ◽  
Viral Protein ◽  
Influenza A ◽  
Viral Rna ◽  
Chain Elongation ◽  
Type I ◽  
Viral Polymerase ◽  
Viral Protein Synthesis

ABSTRACTPattern recognition receptors provide essential nonself immune surveillance within distinct cellular compartments. Retinoic acid-inducible gene I (RIG-I) is one of the primary cytosolic RNA sensors, with an emerging role in the nucleus. It is involved in the spatiotemporal sensing of influenza A virus (IAV) replication, leading to the induction of type I interferons (IFNs). Nonetheless, the physiological viral ligands activating RIG-I during IAV infection remain underexplored. Other than full-length viral genomes, cellular constraints that impede ongoing viral replication likely potentiate an erroneous viral polymerase generating aberrant viral RNA species with RIG-I-activating potential. Here, we investigate the origins of RIG-I-activating viral RNA under two such constraints. Using chemical inhibitors that inhibit continuous viral protein synthesis, we identify the incoming, but notde novo-synthesized, viral defective interfering (DI) genomes contributing to RIG-I activation. In comparison, deprivation of viral nucleoprotein (NP), the key RNA chain elongation factor for the viral polymerase, leads to the production of aberrant viral RNA species activating RIG-I; however, their nature is likely to be distinct from that of DI RNA. Moreover, RIG-I activation in response to NP deprivation is not adversely affected by expression of the nuclear export protein (NEP), which diminishes the generation of a major subset of aberrant viral RNA but facilitates the accumulation of small viral RNA (svRNA). Overall, our results indicate the existence of fundamentally different mechanisms of RIG-I activation under cellular constraints that impede ongoing IAV replication.IMPORTANCEThe induction of an IFN response by IAV is mainly mediated by the RNA sensor RIG-I. The physiological RIG-I ligands produced during IAV infection are not fully elucidated. Cellular constraints leading to the inhibition of ongoing viral replication likely potentiate an erroneous viral polymerase producing aberrant viral RNA species activating RIG-I. Here, we demonstrate that RIG-I activation during chemical inhibition of continuous viral protein synthesis is attributable to the incoming DI genomes. Erroneous viral replication driven by NP deprivation promotes the generation of RIG-I-activating aberrant viral RNA, but their nature is likely to be distinct from that of DI RNA. Our results thus reveal distinct mechanisms of RIG-I activation by IAV under cellular constraints impeding ongoing viral replication. A better understanding of RIG-I sensing of IAV infection provides insight into the development of novel interventions to combat influenza virus infection.

scholarly journals RNase L limits host and viral protein synthesis via inhibition of mRNA export

2021 ◽  
Vol 7 (23) ◽  
pp. eabh2479
Author(s):  
James M. Burke ◽  
Alison R. Gilchrist ◽  
Sara L. Sawyer ◽  
Roy Parker
Keyword(s):  
Protein Synthesis ◽  
Influenza A Virus ◽  
Viral Protein ◽  
Influenza A ◽  
Mrna Export ◽  
Virus Protein ◽  
Rnase L ◽  
Viral Mrna ◽  
Viral Protein Synthesis ◽  
Translation Arrest

RNase L is widely thought to limit viral protein synthesis by cleaving host rRNA and viral mRNA, resulting in translation arrest and viral mRNA degradation. Here, we show that the mRNAs of dengue virus and influenza A virus largely escape RNase L–mediated mRNA decay, and this permits viral protein production. However, activation of RNase L arrests nuclear mRNA export, which strongly inhibits influenza A virus protein synthesis and reduces cytokine production. The heterogeneous and temporal nature of the mRNA export block in individual cells permits sufficient production of antiviral cytokines from transcriptionally induced host mRNAs. This defines RNase L–mediated arrest of mRNA export as a key antiviral shutoff and cytokine regulatory pathway.

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