scholarly journals Host Shutoff in Influenza A Virus: Many Means to an End

Viruses ◽  
2018 ◽  
Vol 10 (9) ◽  
pp. 475 ◽  
Author(s):  
Rachel Levene ◽  
Marta Gaglia
Keyword(s):  
Immune Responses ◽  
Influenza A Virus ◽  
Influenza A ◽  
Rna Synthesis ◽  
Viral Proteins ◽  
Host Gene Expression ◽  
Structural Protein ◽  
Infected Cells ◽  
Host Shutoff ◽  
Virus Replication Cycle

Influenza A virus carries few of its own proteins, but uses them effectively to take control of the infected cells and avoid immune responses. Over the years, host shutoff, the widespread down-regulation of host gene expression, has emerged as a key process that contributes to cellular takeover in infected cells. Interestingly, multiple mechanisms of host shutoff have been described in influenza A virus, involving changes in translation, RNA synthesis and stability. Several viral proteins, notably the non-structural protein NS1, the RNA-dependent RNA polymerase and the endoribonuclease PA-X have been implicated in host shutoff. This multitude of host shutoff mechanisms indicates that host shutoff is an important component of the influenza A virus replication cycle. Here we review the various mechanisms of host shutoff in influenza A virus and the evidence that they contribute to immune evasion and/or viral replication. We also discuss what the purpose of having multiple mechanisms may be.

scholarly journals The influenza A virus host shutoff factor PA-X is rapidly turned over in a strain-specific manner

2021 ◽  
Author(s):  
Rachel Emily Levene ◽  
Shailab D. Shrestha ◽  
Marta Maria Gaglia
Keyword(s):  
Immune Responses ◽  
Influenza A Virus ◽  
Half Life ◽  
Influenza A ◽  
Pandemic H1n1 ◽  
Host Gene Expression ◽  
X Protein ◽  
Protein Levels ◽  
H1n1 Strain ◽  
Host Shutoff

The influenza A endoribonuclease PA-X regulates virulence and transmission of the virus by reducing host gene expression and thus regulating immune responses to influenza A virus. Despite this key function in viral biology, the levels of PA-X protein remain markedly low during infection, and previous results suggest that these low levels are not solely the result of regulation of the level of translation and RNA stability. How PA-X is regulated post-translationally remains unknown. We now report that the PA-X protein is rapidly turned over. PA-X from multiple viral strains are short-lived, although the half-life of PA-X ranges from ∼30 minutes to ∼3.5 hours depending on the strain. Moreover, sequences in the variable PA-X C-terminal domain are primarily responsible for regulating PA-X half-life, although the N-terminal domain also accounts for some differences among strains. Interestingly, we find that the PA-X from the 2009 pandemic H1N1 strain has a longer half-life compared to the other variants we tested. This PA-X isoform has been reported to have a higher host shutoff activity, suggesting a role for protein turnover in regulating PA-X activity. Collectively, this study reveals a novel regulatory mechanism of PA-X protein levels that may impact host shutoff activity during influenza A virus infection. IMPORTANCE The PA-X protein from influenza A virus reduces host immune responses to infection through suppressing host gene expression, including genes encoding the antiviral response. Thus, it plays a central role in influenza A virus biology. Despite its key function, PA-X was only discovered in 2012 and much remains to be learned including how PA-X activity is regulated to promote optimal levels of viral infection. In this study, we reveal that PA-X protein levels are very low likely because of rapid turnover. We show that instability is a conserved property among PA-X variants from different strains of influenza A virus, but that the half-lives of PA-X variants differ. Moreover, the longer half-life of PA-X from the 2009 pandemic H1N1 strain correlates with its reported higher activity. Therefore, PA-X stability may be a way to regulate its activity and may contribute to the differential virulence of influenza A virus strains.

scholarly journals The influenza A virus host shutoff factor PA-X is rapidly turned over in a strain-specific manner

2020 ◽  
Author(s):  
Rachel Emily Levene ◽  
Shailab D. Shrestha ◽  
Marta Maria Gaglia
Keyword(s):  
Immune Responses ◽  
Influenza A Virus ◽  
Half Life ◽  
Influenza A ◽  
Pandemic H1n1 ◽  
Host Gene Expression ◽  
X Protein ◽  
Protein Levels ◽  
H1n1 Strain ◽  
Host Shutoff

ABSTRACTThe influenza A endoribonuclease PA-X regulates virulence and transmission of the virus by reducing host gene expression and thus regulating immune responses to influenza A virus. Despite this key function in viral biology, the levels of PA-X protein remain markedly low during infection, and previous results suggest that these low levels are not solely the result of regulation of the level of translation and RNA stability. How PA-X is regulated post-translationally remains unknown. We now report that the PA-X protein is rapidly turned over. PA-X from multiple viral strains are short-lived, although the half-life of PA-X ranges from ∼30 minutes to ∼3.5 hours depending on the strain. Moreover, sequences in the variable PA-X C-terminal domain are primarily responsible for regulating PA-X half-life, although the N-terminal domain also accounts for some differences among strains. Interestingly, we find that the PA-X from the 2009 pandemic H1N1 strain has a longer half-life compared to the other variants we tested. This PA-X isoform has been reported to have a higher host shutoff activity, suggesting a role for protein turnover in regulating PA-X activity. Collectively, this study reveals a novel regulatory mechanism of PA-X protein levels that may impact host shutoff activity during influenza A virus infection.IMPORTANCEThe PA-X protein from influenza A virus reduces host immune responses to infection through suppressing host gene expression, including genes encoding the antiviral response. Thus, it plays a central role in influenza A virus biology. Despite its key function, PA-X was only discovered in 2012 and much remains to be learned including how PA-X activity is regulated to promote optimal levels of viral infection. In this study, we reveal that PA-X protein levels are very low likely because of rapid turnover. We show that instability is a conserved property among PA-X variants from different strains of influenza A virus, but that the half-lives of PA-X variants differ. Moreover, the longer half-life of PA-X from the 2009 pandemic H1N1 strain correlates with its reported higher activity. Therefore, PA-X stability may be a way to regulate its activity and may contribute to the differential virulence of influenza A virus strains.

scholarly journals The influenza A virus endoribonuclease PA-X usurps host mRNA processing machinery to limit host gene expression

2018 ◽  
Author(s):  
Lea Gaucherand ◽  
Brittany K. Porter ◽  
Summer K. Schmaling ◽  
Christopher Harley Rycroft ◽  
Yuzo Kevorkian ◽  
...  
Keyword(s):  
Gene Expression ◽  
Influenza A Virus ◽  
Rna Splicing ◽  
Influenza A ◽  
Host Gene ◽  
Host Gene Expression ◽  
Viral Control ◽  
New Host ◽  
Antiviral Responses ◽  
Host Shutoff

SUMMARYMany viruses globally shut off host gene expression to inhibit activation of cell-intrinsic antiviral responses. However, host shutoff is not indiscriminate, since viral proteins and host proteins required for viral replication are still synthesized during shutoff. The molecular determinants of target selectivity in host shutoff remain incompletely understood. Here, we report that the influenza A virus shutoff factor PA-X usurps RNA splicing to selectively target host RNAs for destruction. PA-X preferentially degrades spliced mRNAs, both transcriptome-wide and in reporter assays. Moreover, proximity-labeling proteomics revealed that PA-X interacts with cellular proteins involved in RNA splicing. The interaction with splicing contributes to target discrimination and is unique among viral host shutoff nucleases. This novel mechanism sheds light on the specificity of viral control of host gene expression and may provide opportunities for development of new host-targeted antivirals.

scholarly journals Influenza A Virus Protein PA-X Contributes to Viral Growth and Suppression of the Host Antiviral and Immune Responses

2015 ◽  
Vol 89 (12) ◽  
pp. 6442-6452 ◽  
Author(s):  
Tsuyoshi Hayashi ◽  
Leslie A. MacDonald ◽  
Toru Takimoto
Keyword(s):  
Protein Synthesis ◽  
Influenza Virus ◽  
Immune Responses ◽  
Influenza A Virus ◽  
Influenza A ◽  
Host Protein ◽  
Viral Growth ◽  
Host Shutoff ◽  
Host Protein Synthesis ◽  
Host Antiviral Response

ABSTRACTInfluenza virus infection causes global inhibition of host protein synthesis in infected cells. This host shutoff is thought to allow viruses to escape from the host antiviral response, which restricts virus replication and spread. Although the mechanism of host shutoff is unclear, a novel viral protein expressed by ribosomal frameshifting, PA-X, was found to play a major role in influenza virus-induced host shutoff. However, little is known about the impact of PA-X expression on currently circulating influenza A virus pathogenicity and the host antiviral response. In this study, we rescued a recombinant influenza A virus, A/California/04/09 (H1N1, Cal), containing mutations at the frameshift motif in the polymerase PA gene (Cal PA-XFS). Cal PA-XFS expressed significantly less PA-X than Cal wild type (WT). Cal WT, but not Cal PA-XFS, induced degradation of host β-actin mRNA and suppressed host protein synthesis, supporting the idea that PA-X induces host shutoff via mRNA decay. Moreover, Cal WT inhibited beta interferon (IFN-β) expression and replicated more rapidly than Cal PA-XFS in human respiratory cells. Mice infected with Cal PA-XFS had significantly lower levels of viral growth and greater expression of IFN-β mRNA in their lungs than mice infected with Cal WT. Importantly, more antihemagglutinin and neutralizing antibodies were produced in Cal PA-XFS-infected mice than in Cal WT-infected mice, despite the lower level of virus replication in the lungs. Our data indicate that PA-X of the pandemic H1N1 virus has a strong impact on viral growth and the host innate and acquired immune responses to influenza virus.IMPORTANCEVirus-induced host protein shutoff is considered to be a major factor allowing viruses to evade innate and acquired immune recognition. We provide evidence that the 2009 H1N1 influenza A virus protein PA-X plays a role in virus replication and inhibition of host antiviral response by means of its host protein synthesis shutoff activity bothin vitroandin vivo. We also demonstrated that, while the growth of Cal PA-XFS was attenuated in the lungs of infected animals, this mutant induced a stronger humoral response than Cal WT. Our findings clearly highlight the importance of PA-X in counteracting the host innate and acquired immune responses to influenza virus, an important global pathogen. This work demonstrates that inhibition of PA-X expression in influenza virus vaccine strains may provide a novel way of safely attenuating viral growth while inducing a more robust immune response.

scholarly journals Biochemical and Structural Evidence in Support of a Coherent Model for the Formation of the Double-Helical Influenza A Virus Ribonucleoprotein

mBio ◽  
2012 ◽  
Vol 4 (1) ◽  
Author(s):  
Qiaozhen Ye ◽  
Tom S. Y. Guu ◽  
Douglas A. Mata ◽  
Rei-Lin Kuo ◽  
Bartram Smith ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Influenza A Virus ◽  
Influenza A ◽  
Rna Synthesis ◽  
Viral Rna ◽  
Helical Structures ◽  
Structural Element ◽  
X Ray Crystallography ◽  
Multiple Copies ◽  
Infected Cells

ABSTRACTInfluenza A virions contain eight ribonucleoproteins (RNPs), each comprised of a negative-strand viral RNA, the viral polymerase, and multiple nucleoproteins (NPs) that coat the viral RNA. NP oligomerization along the viral RNA is mediated largely by a 28-amino-acid tail loop. Influenza viral RNPs, which serve as the templates for viral RNA synthesis in the nuclei of infected cells, are not linear but rather are organized in hairpin-like double-helical structures. Here we present results that strongly support a coherent model for the assembly of the double-helical influenza virus RNP structure. First, we show that NP self-associates much more weakly in the absence of RNA than in its presence, indicating that oligomerization is very limited in the cytoplasm. We also show that once NP has oligomerized, it can dissociate in the absence of bound RNA, but only at a very slow rate, indicating that the NP scaffold remains intact when viral RNA dissociates from NPs to interact with the polymerase during viral RNA synthesis. In addition, we identify a previously unknown NP-NP interface that is likely responsible for organizing the double-helical viral RNP structure. This identification stemmed from our observation that NP lacking the oligomerization tail loop forms monomers and dimers. We determined the crystal structure of this NP dimer, which reveals this new NP-NP interface. Mutation of residues that disrupt this dimer interface does not affect oligomerization of NPs containing the tail loop but does inactivate the ability of NPs containing the tail loop to support viral RNA synthesis in minigenome assays.IMPORTANCEInfluenza A virus, the causative agent of human pandemics and annual epidemics, contains eight RNA gene segments. Each RNA segment assumes the form of a rod-shaped, double-helical ribonucleoprotein (RNP) that contains multiple copies of a viral protein, the nucleoprotein (NP), which coats the RNA segment along its entire length. Previous studies showed that NP molecules can polymerize via a structural element called the tail loop, but the RNP assembly process is poorly understood. Here we show that influenza virus RNPs are likely assembled from NP monomers, which polymerize through the tail loop only in the presence of viral RNA. Using X-ray crystallography, we identified an additional way that NP molecules interact with each other. We hypothesize that this new interaction is responsible for organizing linear, single-stranded influenza virus RNPs into double-helical structures. Our results thus provide a coherent model for the assembly of the double-helical influenza virus RNP structure.

scholarly journals Identification of a Novel Viral Protein Expressed from the PB2 Segment of Influenza A Virus

2015 ◽  
Vol 90 (1) ◽  
pp. 444-456 ◽  
Author(s):  
Seiya Yamayoshi ◽  
Mariko Watanabe ◽  
Hideo Goto ◽  
Yoshihiro Kawaoka
Keyword(s):  
Influenza Virus ◽  
Influenza A Virus ◽  
Viral Protein ◽  
Influenza A ◽  
Viral Infections ◽  
Viral Proteins ◽  
Wild Type Virus ◽  
Infected Cells

ABSTRACTOver the past 2 decades, several novel influenza virus proteins have been identified that modulate viral infectionsin vitroand/orin vivo. The PB2 segment, which is one of the longest influenza A virus segments, is known to encode only one viral protein, PB2. In the present study, we used reverse transcription-PCR (RT-PCR) targeting viral mRNAs transcribed from the PB2 segment to look for novel viral proteins encoded by spliced mRNAs. We identified a new viral protein, PB2-S1, encoded by a novel spliced mRNA in which the region corresponding to nucleotides 1513 to 1894 of the PB2 mRNA is deleted. PB2-S1 was detected in virus-infected cells and in cells transfected with a protein expression plasmid encoding PB2. PB2-S1 localized to mitochondria, inhibited the RIG-I-dependent interferon signaling pathway, and interfered with viral polymerase activity (dependent on its PB1-binding capability). The nucleotide sequences around the splicing donor and acceptor sites for PB2-S1 were highly conserved among pre-2009 human H1N1 viruses but not among human H1N1pdm and H3N2 viruses. PB2-S1-deficient viruses, however, showed growth kinetics in MDCK cells and virulence in mice similar to those of wild-type virus. The biological significance of PB2-S1 to the replication and pathogenicity of seasonal H1N1 influenza A viruses warrants further investigation.IMPORTANCETranscriptome analysis of cells infected with influenza A virus has improved our understanding of the host response to viral infection, because such analysis yields considerable information about bothin vitroandin vivoviral infections. However, little attention has been paid to transcriptomes derived from the viral genome. Here we focused on the splicing of mRNA expressed from the PB2 segment and identified a spliced viral mRNA encoding a novel viral protein. This result suggests that other, as yet unidentified viral proteins encoded by spliced mRNAs could be expressed in virus-infected cells. A viral transcriptome including the viral spliceosome should be evaluated to gain new insights into influenza virus infection.

scholarly journals Closing in on the causes of host shutoff

eLife ◽  
2016 ◽  
Vol 5 ◽  
Author(s):  
Ian Mohr
Keyword(s):  
Influenza A Virus ◽  
Influenza A ◽  
Infected Cells ◽  
Host Shutoff

Influenza A virus suppresses the translation of host mRNA by selectively remodeling and dominating the pool of mRNA in infected cells.

scholarly journals Necroptosis restricts influenza A virus as a stand-alone cell death mechanism

2020 ◽  
Vol 217 (11) ◽  
Author(s):  
Maria Shubina ◽  
Bart Tummers ◽  
David F. Boyd ◽  
Ting Zhang ◽  
Chaoran Yin ◽  
...  
Keyword(s):  
Cell Death ◽  
Immune Responses ◽  
Influenza A Virus ◽  
Respiratory Infection ◽  
Host Defense ◽  
Influenza A ◽  
Cell Death Mechanism ◽  
Virus Clearance ◽  
Infected Cells ◽  
Fail Safe

Influenza A virus (IAV) activates ZBP1-initiated RIPK3-dependent parallel pathways of necroptosis and apoptosis in infected cells. Although mice deficient in both pathways fail to control IAV and succumb to lethal respiratory infection, RIPK3-mediated apoptosis by itself can limit IAV, without need for necroptosis. However, whether necroptosis, conventionally considered a fail-safe cell death mechanism to apoptosis, can restrict IAV—or indeed any virus—in the absence of apoptosis is not known. Here, we use mice selectively deficient in IAV-activated apoptosis to show that necroptosis drives robust antiviral immune responses and promotes effective virus clearance from infected lungs when apoptosis is absent. We also demonstrate that apoptosis and necroptosis are mutually exclusive fates in IAV-infected cells. Thus, necroptosis is an independent, “stand-alone” cell death mechanism that fully compensates for the absence of apoptosis in antiviral host defense.

scholarly journals Dicer is involved in protection against influenza A virus infection

2007 ◽  
Vol 88 (10) ◽  
pp. 2627-2635 ◽  
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.

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

2010 ◽  
Vol 84 (24) ◽  
pp. 12832-12840 ◽  
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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