scholarly journals Identification of an Interferon-Stimulated Long Noncoding RNA (LncRNA ISR) Involved in Regulation of Influenza A Virus Replication

2019 ◽  
Vol 20 (20) ◽  
pp. 5118 ◽  
Author(s):  
Qidong Pan ◽  
Zhonghui Zhao ◽  
Yuan Liao ◽  
Shih-Hsin Chiu ◽  
Song Wang ◽  
...  
Keyword(s):  
Viral Replication ◽  
Influenza A Virus ◽  
Noncoding Rna ◽  
Influenza A ◽  
Ectopic Expression ◽  
A549 Cells ◽  
Mouse Cell ◽  
Type I ◽  
Human Beings ◽  
Inhibitor Treatment

Long noncoding RNAs (lncRNAs) are involved in a diversity of biological processes. It is known that differential expression of thousands of lncRNAs occurs in host during influenza A virus (IAV) infection. However, only few of them have been well characterized. Here, we identified a lncRNA, named as interferon (IFN)-stimulated lncRNA (ISR), which can be significantly upregulated in response to IAV infection in a mouse model. A sequence alignment revealed that lncRNA ISR is present in mice and human beings, and indeed, we found that it was expressed in several human and mouse cell lines and tissues. Silencing lncRNA ISR in A549 cells resulted in a significant increase in IAV replication, whereas ectopic expression of lncRNA ISR reduced the viral replication. Interestingly, interferon-β (IFN-β) treatment was able to induce lncRNA ISR expression, and induction of lncRNA ISR by viral infection was nearly abolished in host deficient of IFNAR1, a type I IFN receptor. Furthermore, the level of IAV-induced lncRNA ISR expression was decreased either in retinoic acid-inducible gene I (RIG-I) knockout A549 cells and mice or by nuclear factor κ-light-chain-enhancer of activated B cells (NF-κB) inhibitor treatment. Together, these data elucidate that lncRNA ISR is regulated by RIG-I-dependent signaling that governs IFN-β production during IAV infection, and has an inhibitory capacity in viral replication.

scholarly journals Cellular co-infection can modulate the efficiency of influenza A virus production and shape the interferon response

2019 ◽  
Author(s):  
Brigitte E. Martin ◽  
Jeremy D. Harris ◽  
Jiayi Sun ◽  
Katia Koelle ◽  
Christopher B. Brooke
Keyword(s):  
Influenza A Virus ◽  
Cell Lines ◽  
Influenza A ◽  
Mdck Cells ◽  
A549 Cells ◽  
Virus Production ◽  
Innate Immune ◽  
Type I ◽  
Functional Forms ◽  
A Cell

ABSTRACTDuring viral infection, the numbers of virions infecting individual cells can vary significantly over time and space. The functional consequences of this variation in cellular multiplicity of infection (MOI) remain poorly understood. Here, we rigorously quantify the phenotypic consequences of cellular MOI during influenza A virus (IAV) infection over a single round of replication in terms of cell death rates, viral output kinetics, interferon and antiviral effector gene transcription, and superinfection potential. By statistically fitting mathematical models to our data, we precisely define specific functional forms that quantitatively describe the modulation of these phenotypes by MOI at the single cell level. To determine the generality of these functional forms, we compare two distinct cell lines (MDCK cells and A549 cells), both infected with the H1N1 strain A/Puerto Rico/8/1934 (PR8). We find that a model assuming that infected cell death rates are independent of cellular MOI best fits the experimental data in both cell lines. We further observe that a model in which the rate and efficiency of virus production increase with cellular co-infection best fits our observations in MDCK cells, but not in A549 cells. In A549 cells, we also find that induction of type III interferon, but not type I interferon, is highly dependent on cellular MOI, especially at early timepoints. This finding identifies a role for cellular co-infection in shaping the innate immune response to IAV infection. Finally, we show that higher cellular MOI is associated with more potent superinfection exclusion, thus limiting the total number of virions capable of infecting a cell. Overall, this study suggests that the extent of cellular co-infection by influenza viruses may be a critical determinant of both viral production kinetics and cellular infection outcomes in a host cell type-dependent manner.AUTHOR SUMMARYDuring influenza A virus (IAV) infection, the number of virions to enter individual cells can be highly variable. Cellular co-infection appears to be common and plays an essential role in facilitating reassortment for IAV, yet little is known about how cellular co-infection influences infection outcomes at the cellular level. Here, we combine quantitative in vitro infection experiments with statistical model fitting to precisely define the phenotypic consequences of cellular co-infection in two cell lines. We reveal that cellular co-infection can increase and accelerate the efficiency of IAV production in a cell line-dependent fashion, identifying it as a potential determinant of viral replication kinetics. We also show that induction of type III, but not type I, interferon is highly dependent upon the number of virions that infect a given cell, implicating cellular co-infection as an important determinant of the host innate immune response to infection. Altogether, our findings show that cellular co-infection plays a crucial role in determining infection outcome. The integration of experimental and statistical modeling approaches detailed here represents a significant advance in the quantitative study of influenza virus infection and should aid ongoing efforts focused on the construction of mathematical models of IAV infection.

scholarly journals Properties of H7N7 influenza A virus strain SC35M lacking interferon antagonist NS1 in mice and chickens

2007 ◽  
Vol 88 (5) ◽  
pp. 1403-1409 ◽  
Author(s):  
Georg Kochs ◽  
Iris Koerner ◽  
Lena Thiel ◽  
Sonja Kothlow ◽  
Bernd Kaspers ◽  
...  
Keyword(s):  
Influenza A Virus ◽  
Virulence Factor ◽  
Influenza A ◽  
Severe Disease ◽  
A549 Cells ◽  
Type I ◽  
Upper Respiratory Tract ◽  
Structural Protein ◽  
Wild Type ◽  
Ns1 Gene

Non-structural protein NS1 of influenza A virus counteracts the host immune response by blocking the synthesis of type I interferon (IFN). As deletion of the complete NS1 gene has to date been reported only in the human H1N1 strain A/PR/8/34, it remained unclear whether NS1 is a non-essential virulence factor in other influenza A virus strains as well. In this report, the properties of NS1-deficient mutants derived from strain SC35M (H7N7) are described. A mutant of SC35M that completely lacks the NS1 gene was an excellent inducer of IFN in mammalian and avian cells in culture and, consequently, was able to multiply efficiently only in cell lines with defects in the type I IFN system. Virus mutants carrying C-terminally truncated versions of NS1 were less powerful inducers of IFN and were attenuated less strongly in human A549 cells. Although attenuated in wild-type mice, these mutants remained highly pathogenic for mice lacking the IFN-regulated antiviral factor Mx1. In contrast, the NS1-deficient SC35M mutant was completely non-pathogenic for wild-type mice, but remained pathogenic for mice lacking Mx1 and double-stranded RNA-activated protein kinase (PKR). Wild-type SC35M, but not the NS1-deficient mutant virus, was able to replicate in the upper respiratory tract of birds, but neither virus induced severe disease in adult chickens. Altogether, this study supports the view that NS1 represents a non-essential virulence factor of different influenza A viruses.

scholarly journals MicroRNA-21-3p modulates FGF2 to facilitate influenza A virus H5N1 replication by refraining type I interferon response

2020 ◽  
Vol 40 (5) ◽  
Author(s):  
Jianli Shi ◽  
Ping Feng ◽  
Tingting Gu
Keyword(s):  
Influenza A Virus ◽  
Influenza A ◽  
Type I Interferon ◽  
A549 Cells ◽  
Virus Titer ◽  
Interferon Response ◽  
Luciferase Reporter ◽  
Type I ◽  
Structural Factors ◽  
Oligoadenylate Synthetase

Abstract Background: Influenza A virus (IAV) has greatly affected public health in recent decades. Accumulating data indicated that host microRNAs (miRNAs) were related to IAV replication. The present study mainly focused on the effects of microRNA-21-3p (miR-21-3p) on H5N1 replication. Methods: The levels of miR-21-3p, virus structural factors (matrix 1 (M1), nucleoprotein (NP)), type I interferon (IFN) response markers (IFN-β, IFN-α), IFN-stimulated genes (protein kinase R (PKR), myxovirus resistance A (MxA), 2′-5′-oligoadenylate synthetase 2 (OAS)), and fibroblast growth factor 2 (FGF2) were measured by quantitative real-time polymerase chain reaction (qRT-PCR). The protein levels of M1, NP, and FGF2 were tested by Western blot assay. The virus titer was assessed by tissue culture infective dose 50% (TCID50) assay. The dual-luciferase reporter assay and ribonucleic acid (RNA) immunoprecipitation (RIP) assay were used to verify the interaction between miR-21-3p and FGF2. Results: MiR-21-3p was reduced in H5N1-infected patients and A549 cells. MiR-21-3p overexpression facilitated the levels of M1, NP, TCID50 value, and reduced the levels of IFN-β, IFN-α, PKR, MxA, and OAS in H5N1-infected A549 cells. FGF2 was verified as a direct target of miR-21-3p. The introduction of FGF2 counteracted miR-21-3p-mediated decrease in the levels of M1, NP, and TCID50 value, as well as reduction in the levels of IFN-β, IFN-α, PKR, MxA, and OAS in H5N1-infected A549 cells. Conclusion: MiR-21-3p down-regulated FGF2 expression to accelerate H5N1 replication and confine IFN response.

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 Upregulation of miR-101 during Influenza A Virus Infection Abrogates Viral Life Cycle by Targeting mTOR Pathway

Viruses ◽  
2020 ◽  
Vol 12 (4) ◽  
pp. 444 ◽  
Author(s):  
Shipra Sharma ◽  
Anirvan Chatterjee ◽  
Purnima Kumar ◽  
Sunil Lal ◽  
Kiran Kondabagil
Keyword(s):  
Influenza A Virus ◽  
Influenza A ◽  
Potent Inhibitor ◽  
Ectopic Expression ◽  
A549 Cells ◽  
Mtor Pathway ◽  
Viral Protein Synthesis ◽  
Viral Propagation ◽  
Qpcr Analysis ◽  
Micro Rnas

Micro RNAs (miRNAs) are a class of small non-coding single-stranded RNA, which play an important role in modulating host-Influenza A virus (IAV) crosstalk. The interplay between influenza and miRNA interaction is defined by a plethora of complex mechanisms, which are not fully understood yet. Here, we demonstrate that in IAV infected A549 cells, a synchronous increase was observed in the expression of mTOR up to 24 hpi and significant downregulation at 48 hpi. Additionally, NP of IAV interacts with mTOR and modulates the levels of mTOR mRNA and protein, thus regulating the translation of host cell. RNA sequencing and qPCR analysis of IAV-infected A549 cells and NP transfected cells revealed that miR-101 downregulates mTOR transcripts at later stages of infection. Ectopic expression of miR-101 mimic led to a decrease in expression of NP, a reduction in IAV titer and replication. Moreover, treatment of the cells with Everolimus, a potent inhibitor of mTOR, resulted in an increase of miR-101 transcript levels, which further suppressed the viral protein synthesis. Collectively, the data suggest a novel mechanism that IAV stimulates mTOR pathway at early stages of infection; however, at a later time-point, positive regulation of miR-101 restrains the mTOR expression, and hence, the viral propagation.

scholarly journals Influenza A Virus (H1N1) Infection Induces Glycolysis to Facilitate Viral Replication

2021 ◽  
Author(s):  
Lehao Ren ◽  
Wanju Zhang ◽  
Jing Zhang ◽  
Jiaxiang Zhang ◽  
Huiying Zhang ◽  
...  
Keyword(s):  
Viral Replication ◽  
Influenza A Virus ◽  
Influenza A ◽  
A549 Cells ◽  
Glycolytic Pathway ◽  
Pyruvate Dehydrogenase Kinase ◽  
Metabolic Inhibitors ◽  
Mouse Lung ◽  
Virus H1n1 ◽  
H1n1 Infection

AbstractViruses depend on host cellular metabolism to provide the energy and biosynthetic building blocks required for their replication. In this study, we observed that influenza A virus (H1N1), a single-stranded, negative-sense RNA virus with an eight-segmented genome, enhanced glycolysis both in mouse lung tissues and in human lung epithelial (A549) cells. In detail, the expression of hexokinase 2 (HK2), the first enzyme in glycolysis, was upregulated in H1N1-infected A549 cells, and the expression of pyruvate kinase M2 (PKM2) and pyruvate dehydrogenase kinase 3 (PDK3) was upregulated in H1N1-infected mouse lung tissues. Pharmacologically inhibiting the glycolytic pathway or targeting hypoxia-inducible factor 1 (HIF-1), the central transcriptional factor critical for glycolysis, significantly reduced H1N1 replication, revealing a requirement for glycolysis during H1N1 infection. In addition, pharmacologically enhancing the glycolytic pathway further promoted H1N1 replication. Furthermore, the change of H1N1 replication upon glycolysis inhibition or enhancement was independent of interferon signaling. Taken together, these findings suggest that influenza A virus induces the glycolytic pathway and thus facilitates efficient viral replication. This study raises the possibility that metabolic inhibitors, such as those that target glycolysis, could be used to treat influenza A virus infection in the future.

scholarly journals Human Properdin Released By Infiltrating Neutrophils Can Modulate Influenza A Virus Infection

2021 ◽  
Vol 12 ◽  
Author(s):  
Praveen M. Varghese ◽  
Shuvechha Mukherjee ◽  
Futwan A. Al-Mohanna ◽  
Souad M. Saleh ◽  
Fahad N. Almajhdi ◽  
...  
Keyword(s):  
Virus Infection ◽  
Inflammatory Response ◽  
Viral Replication ◽  
Influenza A Virus ◽  
Influenza A ◽  
Alternative Pathway ◽  
A549 Cells ◽  
H1n1 Subtype ◽  
H3n2 Subtype ◽  
Influenza A Virus Infection

The complement system is designed to recognise and eliminate invading pathogens via activation of classical, alternative and lectin pathways. Human properdin stabilises the alternative pathway C3 convertase, resulting in an amplification loop that leads to the formation of C5 convertase, thereby acting as a positive regulator of the alternative pathway. It has been noted that human properdin on its own can operate as a pattern recognition receptor and exert immune functions outside its involvement in complement activation. Properdin can bind directly to microbial targets via DNA, sulfatides and glycosaminoglycans, apoptotic cells, nanoparticles, and well-known viral virulence factors. This study was aimed at investigating the complement-independent role of properdin against Influenza A virus infection. As one of the first immune cells to arrive at the site of IAV infection, we show here that IAV challenged neutrophils released properdin in a time-dependent manner. Properdin was found to directly interact with haemagglutinin, neuraminidase and matrix 1 protein Influenza A virus proteins in ELISA and western blot. Furthermore, modelling studies revealed that properdin could bind HA and NA of the H1N1 subtype with higher affinity compared to that of H3N2 due to the presence of an HA cleavage site in H1N1. In an infection assay using A549 cells, properdin suppressed viral replication in pH1N1 subtype while promoting replication of H3N2 subtype, as revealed by qPCR analysis of M1 transcripts. Properdin treatment triggered an anti-inflammatory response in H1N1-challenged A549 cells and a pro-inflammatory response in H3N2-infected cells, as evident from differential mRNA expression of TNF-α, NF-κB, IFN-α, IFN-β, IL-6, IL-12 and RANTES. Properdin treatment also reduced luciferase reporter activity in MDCK cells transduced with H1N1 pseudotyped lentiviral particles; however, it was increased in the case of pseudotyped H3N2 particles. Collectively, we conclude that infiltrating neutrophils at the site of IAV infection can release properdin, which then acts as an entry inhibitor for pandemic H1N1 subtype while suppressing viral replication and inducing an anti-inflammatory response. H3N2 subtype can escape this immune restriction due to altered haemagglutinin and neuraminindase, leading to enhanced viral entry, replication and pro-inflammatory response. Thus, depending on the subtype, properdin can either limit or aggravate IAV infection in the host.

scholarly journals Influenza A Virus Inhibits Type I IFN Signaling via NF-κB-Dependent Induction of SOCS-3 Expression

2008 ◽  
Vol 4 (11) ◽  
pp. e1000196 ◽  
Author(s):  
Eva-K. Pauli ◽  
Mirco Schmolke ◽  
Thorsten Wolff ◽  
Dorothee Viemann ◽  
Johannes Roth ◽  
...  
Keyword(s):  
Influenza A Virus ◽  
Influenza A ◽  
Type I ◽  
Type I Ifn

scholarly journals Cyclophilin A interacts with influenza A virus M1 protein and impairs the early stage of the viral replication

2009 ◽  
Vol 11 (5) ◽  
pp. 730-741 ◽  
Author(s):  
Xiaoling Liu ◽  
Lei Sun ◽  
Maorong Yu ◽  
Zengfu Wang ◽  
Chongfeng Xu ◽  
...  
Keyword(s):  
Viral Replication ◽  
Influenza A Virus ◽  
Influenza A ◽  
Early Stage ◽  
Cyclophilin A ◽  
M1 Protein
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