scholarly journals Adaptive Mutations in Influenza A/California/07/2009 Enhance Polymerase Activity and Infectious Virion Production

Viruses ◽  
2018 ◽  
Vol 10 (5) ◽  
pp. 272 ◽  
Author(s):  
Patrick Slaine ◽  
Cara MacRae ◽  
Mariel Kleer ◽  
Emily Lamoureux ◽  
Sarah McAlpine ◽  
...  
Keyword(s):  
Influenza A ◽  
Polymerase Activity ◽  
Adaptive Mutations ◽  
Infectious Virion ◽  
Virion Production

scholarly journals Equine Mx1 Restricts Influenza A Virus Replication by Targeting at Distinct Site of its Nucleoprotein

Viruses ◽  
2019 ◽  
Vol 11 (12) ◽  
pp. 1114 ◽  
Author(s):  
Urooj Fatima ◽  
Zhenyu Zhang ◽  
Haili Zhang ◽  
Xue-Feng Wang ◽  
Ling Xu ◽  
...  
Keyword(s):  
Influenza A Virus ◽  
Influenza A ◽  
Substantial Reduction ◽  
Polymerase Activity ◽  
Influenza Viruses ◽  
Dynamic Force ◽  
Equine Influenza Virus ◽  
Equine Influenza ◽  
Adaptive Mutations ◽  
Antiviral Activities

Interferon-mediated host factors myxovirus (Mx) proteins are key features in regulating influenza A virus (IAV) infections. Viral polymerases are essential for viral replication. The Mx1 protein has been known to interact with viral nucleoprotein (NP) and PB2, resulting in the influence of polymerase activity and providing interspecies restriction. The equine influenza virus has evolved as an independent lineage to influenza viruses from other species. We estimated the differences in antiviral activities between human MxA (huMxA) and equine Mx1 (eqMx1) against a broad range of IAV strains. We found that huMxA has antiviral potential against IAV strains from non-human species, whereas eqMx1 could only inhibit the polymerase activity of non-equine species. Here, we demonstrated that NP is the main target of eqMx1. Subsequently, we found adaptive mutations in the NP of strains A/equine/Jilin/1/1989 (H3N8JL89) and A/chicken/Zhejiang/DTID-ZJU01/2013 (H7N9ZJ13) that confer eqMx1 resistance and sensitivity respectively. A substantial reduction in Mx1 resistance was observed for the two mutations G34S and H52N in H3N8JL89 NP. Thus, eqMx1 is an important dynamic force in IAV nucleoprotein evolution. We, therefore, suggest that the amino acids responsible for Mx1 resistance should be regarded as a robust indicator for the pandemic potential of lately evolving IAVs.

scholarly journals Computational and molecular analysis of conserved influenza A virus RNA secondary structures involved in infectious virion production

RNA Biology ◽  
2016 ◽  
Vol 13 (9) ◽  
pp. 883-894 ◽  
Author(s):  
Yuki Kobayashi ◽  
Bernadeta Dadonaite ◽  
Neeltje van Doremalen ◽  
Yoshiyuki Suzuki ◽  
Wendy S. Barclay ◽  
...  
Keyword(s):  
Influenza A Virus ◽  
Molecular Analysis ◽  
Influenza A ◽  
Secondary Structures ◽  
Rna Secondary Structures ◽  
Infectious Virion ◽  
Virion Production ◽  
Virus Rna

scholarly journals Key Role of the Influenza A Virus PA Gene Segment in the Emergence of Pandemic Viruses

Viruses ◽  
2020 ◽  
Vol 12 (4) ◽  
pp. 365 ◽  
Author(s):  
Michael M. Lutz ◽  
Megan M. Dunagan ◽  
Yuki Kurebayashi ◽  
Toru Takimoto
Keyword(s):  
Mammalian Cells ◽  
Influenza A ◽  
Gene Segment ◽  
Polymerase Activity ◽  
Influenza A Viruses ◽  
Adaptive Mutations ◽  
Natural Reservoir ◽  
Wide Range ◽  
Seasonal Epidemics

Influenza A viruses (IAVs) are a significant human pathogen that cause seasonal epidemics and occasional pandemics. Avian waterfowl are the natural reservoir of IAVs, but a wide range of species can serve as hosts. Most IAV strains are adapted to one host species and avian strains of IAV replicate poorly in most mammalian hosts. Importantly, IAV polymerases from avian strains function poorly in mammalian cells but host adaptive mutations can restore activity. The 2009 pandemic H1N1 (H1N1pdm09) virus acquired multiple mutations in the PA gene that activated polymerase activity in mammalian cells, even in the absence of previously identified host adaptive mutations in other polymerase genes. These mutations in PA localize within different regions of the protein suggesting multiple mechanisms exist to activate polymerase activity. Additionally, an immunomodulatory protein, PA-X, is expressed from the PA gene segment. PA-X expression is conserved amongst many IAV strains but activity varies between viruses specific for different hosts, suggesting that PA-X also plays a role in host adaptation. Here, we review the role of PA in the emergence of currently circulating H1N1pdm09 viruses and the most recent studies of host adaptive mutations in the PA gene that modulate polymerase activity and PA-X function.

scholarly journals Adaptive Mutations in Influenza A/California/07/2009 Enhance Polymerase Activity and Infectious Virion Production

2018 ◽  
Author(s):  
Patrick D. Slaine ◽  
Cara MacRae ◽  
Mariel Kleer ◽  
Emily Lamoureux ◽  
Sarah McAlpine ◽  
...  
Keyword(s):  
Influenza A ◽  
Serial Passage ◽  
Polymerase Activity ◽  
Influenza A Viruses ◽  
Adaptation Studies ◽  
Adaptive Mutations ◽  
Susceptibility To Infection ◽  
Natural Hosts ◽  
2009 H1n1 ◽  
Polymerase Basic 1

Mice are not natural hosts for influenza A viruses (IAVs), but they are useful models for studying antiviral immune responses and pathogenesis. Serial passage of IAV in mice invariably causes the emergence of adaptive mutations and increased virulence. Typically, mouse-adaptation studies are conducted in inbred laboratory strains BALB/c and C57BL/6, which have defects in the antiviral Mx1 gene that results in increased susceptibility to infection and disease severity. Here, we report the adaptation of IAV reference strain A/California/07/2009(H1N1) (a.k.a. CA/07) in outbred Swiss Webster mice. Serial passage led to increased virulence and lung titers, and dissemination of the virus to brains. We adapted a deep-sequencing protocol to identify and enumerate adaptive mutations across all genome segments. Among mutations that emerged during mouse-adaptation, we focused on amino acid substitutions in polymerase subunits: polymerase basic-1 (PB1) T156A and F740L, and polymerase acidic (PA) E349G. These mutations were evaluated singly and in combination in minigenome replicon assays, which revealed that PA E349G increased polymerase activity. By selectively engineering these three adaptive PB1 and PA mutations into the parental CA/07 strain, we demonstrated that adaptive mutations in polymerase subunits decreased the production of defective viral genome segments with internal deletions, and dramatically increased the release of infectious virions from mouse cells. Together, these findings increase our understanding of the contribution of polymerase subunits to successful host adaptation.

scholarly journals PB2 and Hemagglutinin Mutations Are Major Determinants of Host Range and Virulence in Mouse-Adapted Influenza A Virus

2010 ◽  
Vol 84 (20) ◽  
pp. 10606-10618 ◽  
Author(s):  
Jihui Ping ◽  
Samar K. Dankar ◽  
Nicole E. Forbes ◽  
Liya Keleta ◽  
Yan Zhou ◽  
...  
Keyword(s):  
Host Range ◽  
Influenza A Virus ◽  
Influenza A ◽  
Lethal Dose ◽  
Polymerase Activity ◽  
Mouse Lung ◽  
Viral Gene ◽  
Adaptive Mutations

ABSTRACT Serial mouse lung passage of a human influenza A virus, A/Hong Kong/1/68 (H3N2) (HK-wt), produced a mouse-adapted variant, MA, with nine mutations that was >103.8-fold more virulent. In this study, we demonstrate that MA mutations of the PB2 (D701N) and hemagglutinin (HA) (G218W in HA1 and T156N in HA2) genes were the most adaptive genetic determinants for increased growth and virulence in the mouse model. Recombinant viruses expressing each of the mutated MA genome segments on the HK-wt backbone showed significantly increased disease severity, whereas only the mouse-adapted PB2 gene increased virulence, as determined by the 50% lethal dose ([LD50] >101.4-fold). The converse comparisons of recombinant MA viruses expressing each of the HK-wt genome segments showed the greatest decrease in virulence due to the HA gene (102-fold), with lesser decreases due to the M1, NS1, NA, and PB1 genes (100.3- to 100.8-fold), and undetectable effects on the LD50 for the PB2 and NP genes. The HK PB2 gene did, however, attenuate MA infection, as measured by weight loss and time to death. Replication of adaptive mutations in vivo and in vitro showed both viral gene backbone and host range effects. Minigenome transcription assays showed that PB1 and PB2 mutations increased polymerase activity and that the PB2 D701N mutation was comparable in effect to the mammalian adaptive PB2 E627K mutation. Our results demonstrate that host range and virulence are controlled by multiple genes, with major roles for mutations in PB2 and HA.

scholarly journals Adaptive Mutations Resulting in Enhanced Polymerase Activity Contribute to High Virulence of Influenza A Virus in Mice

2009 ◽  
Vol 83 (13) ◽  
pp. 6673-6680 ◽  
Author(s):  
Thierry Rolling ◽  
Iris Koerner ◽  
Petra Zimmermann ◽  
Kristian Holz ◽  
Otto Haller ◽  
...  
Keyword(s):  
Influenza A Virus ◽  
Influenza A ◽  
Polymerase Activity ◽  
Host Cells ◽  
Mammalian Host ◽  
Avian Host ◽  
Viral Polymerase ◽  
Adaptive Mutations ◽  
High Virulence ◽  
Adaptation Experiment

ABSTRACT High virulence of influenza virus A/Puerto Rico/8/34 in mice carrying the Mx1 resistance gene was recently shown to be determined by the viral surface proteins and the viral polymerase. Here, we demonstrated high-level polymerase activity in mammalian host cells but not avian host cells and investigated which mutations in the polymerase subunits PB1, PB2, and PA are critical for increased polymerase activity and high virus virulence. Mutational analyses demonstrated that an isoleucine-to-valine change at position 504 in PB2 was the most critical and strongly enhanced the activity of the reconstituted polymerase complex. An isoleucine-to-leucine change at position 550 in PA further contributed to increased polymerase activity and high virulence, whereas all other mutations in PB1, PB2, and PA were irrelevant. To determine whether this pattern of acquired mutations represents a preferred viral strategy to gain virulence, two independent new virus adaptation experiments were performed. Surprisingly, the conservative I504V change in PB2 evolved again and was the only mutation present in an aggressive virus variant selected during the first adaptation experiment. In contrast, the virulent virus selected in the second adaptation experiment had a lysine-to-arginine change at position 208 in PB1 and a glutamate-to-glycine change at position 349 in PA. These results demonstrate that a variety of minor amino acid changes in the viral polymerase can contribute to enhanced virulence of influenza A virus. Interestingly, all virulence-enhancing mutations that we identified in this study resulted in substantially increased viral polymerase activity.

scholarly journals Structural and Thermodynamic Analysis of the Resistance Development to Pimodivir (VX-787), the Clinical Inhibitor of Cap Binding to PB2 Subunit of Influenza A Polymerase

Molecules ◽  
2021 ◽  
Vol 26 (4) ◽  
pp. 1007
Author(s):  
Jiří Gregor ◽  
Kateřina Radilová ◽  
Jiří Brynda ◽  
Jindřich Fanfrlík ◽  
Jan Konvalinka ◽  
...  
Keyword(s):  
Thermodynamic Analysis ◽  
Influenza A ◽  
Polymerase Activity ◽  
Second Phase ◽  
Missense Mutations ◽  
Wild Type ◽  
Resistance Development ◽  
Third Phase ◽  
Major Mutation ◽  
Control And Prevention

Influenza A virus (IAV) encodes a polymerase composed of three subunits: PA, with endonuclease activity, PB1 with polymerase activity and PB2 with host RNA five-prime cap binding site. Their cooperation and stepwise activation include a process called cap-snatching, which is a crucial step in the IAV life cycle. Reproduction of IAV can be blocked by disrupting the interaction between the PB2 domain and the five-prime cap. An inhibitor of this interaction called pimodivir (VX-787) recently entered the third phase of clinical trial; however, several mutations in PB2 that cause resistance to pimodivir were observed. First major mutation, F404Y, causing resistance was identified during preclinical testing, next the mutation M431I was identified in patients during the second phase of clinical trials. The mutation H357N was identified during testing of IAV strains at Centers for Disease Control and Prevention. We set out to provide a structural and thermodynamic analysis of the interactions between cap-binding domain of PB2 wild-type and PB2 variants bearing these mutations and pimodivir. Here we present four crystal structures of PB2-WT, PB2-F404Y, PB2-M431I and PB2-H357N in complex with pimodivir. We have thermodynamically analysed all PB2 variants and proposed the effect of these mutations on thermodynamic parameters of these interactions and pimodivir resistance development. These data will contribute to understanding the effect of these missense mutations to the resistance development and help to design next generation inhibitors.

scholarly journals Human Cytomegalovirus Inhibitor AL18 Also Possesses Activity against Influenza A and B Viruses

2012 ◽  
Vol 56 (11) ◽  
pp. 6009-6013 ◽  
Author(s):  
Giulia Muratore ◽  
Beatrice Mercorelli ◽  
Laura Goracci ◽  
Gabriele Cruciani ◽  
Paul Digard ◽  
...  
Keyword(s):  
Influenza A Virus ◽  
Dna Polymerase ◽  
Human Cytomegalovirus ◽  
Influenza A ◽  
Molecular Model ◽  
Polymerase Activity

ABSTRACTAL18, an inhibitor of human cytomegalovirus DNA polymerase, was serendipitously found to also block the interaction between the PB1 and PA polymerase subunits of influenza A virus. Furthermore, AL18 effectively inhibited influenza A virus polymerase activity and the overall replication of influenza A and B viruses. A molecular model to explain the binding of AL18 to both cytomegalovirus and influenza targets is proposed. Thus, AL18 represents an interesting lead for the development of new antivirals.

scholarly journals Multiple polymerase acidic (PA) I38X substitutions in influenza A(H1N1)pdm09 virus permit polymerase activity and cause reduced baloxavir inhibition

2020 ◽  
Author(s):  
Jeremy C Jones ◽  
Philippe N Q Pascua ◽  
Walter N Harrington ◽  
Richard J Webby ◽  
Elena A Govorkova
Keyword(s):  
Inhibitory Activity ◽  
Influenza A ◽  
Antiviral Drug ◽  
Polymerase Activity ◽  
Resistance Marker ◽  
Complex Activity ◽  
Replication Complex ◽  
Virus Rescue ◽  
Influenza A H1n1 ◽  
Targeted Approach

Abstract Background Baloxavir marboxil is an antiviral drug that targets the endonuclease activity of the influenza virus polymerase acidic (PA) protein. PA I38T/M/F substitutions reduce its antiviral efficacy. Objectives To understand the effects of the 19 possible amino acid (AA) substitutions at PA 38 on influenza A(H1N1)pdm09 polymerase activity and inhibition by baloxavir acid, the active metabolite of baloxavir marboxil. Methods Influenza A(H1N1)pdm09 viral polymerase complexes containing all 19 I38X AA substitutions were reconstituted in HEK293T cells in a mini-replicon assay. Polymerase complex activity and baloxavir inhibitory activity were measured in the presence or absence of 50 nM baloxavir acid. Results Only three substitutions (R, K, P) reduced polymerase activity to <79% of I38-WT. When compared with the prototypical baloxavir marboxil resistance marker T38, 5 substitutions conferred 10%–35% reductions in baloxavir acid inhibitory activity (M, L, F, Y, C) and 11 substitutions conferred >50% reductions (R, K, S, N, G, W, A, Q, E, D, H), while two substitutions (V, P) maintained baloxavir acid inhibitory activity. Conclusions Most PA 38 substitutions permit a functional replication complex retaining some drug resistance in the mini-replicon assay. This study provides a targeted approach for virus rescue and analysis of novel baloxavir marboxil reduced-susceptibility markers, supports the consideration of a broader range of these markers during antiviral surveillance and adds to the growing knowledge of baloxavir marboxil resistance profiles.

scholarly journals HDAC6 Restricts Influenza A Virus by Deacetylation of the RNA Polymerase PA Subunit

2018 ◽  
Vol 93 (4) ◽  
Author(s):  
Huan Chen ◽  
Yingjuan Qian ◽  
Xin Chen ◽  
Zhiyang Ruan ◽  
Yuetian Ye ◽  
...  
Keyword(s):  
Life Cycle ◽  
Rna Polymerase ◽  
Influenza A Virus ◽  
Influenza A ◽  
Molecular Mechanisms ◽  
Polymerase Activity ◽  
Negative Regulator ◽  
Host Protein ◽  
Spectrometric Analysis ◽  
Rna Polymerase Activity

ABSTRACT The life cycle of influenza A virus (IAV) is modulated by various cellular host factors. Although previous studies indicated that IAV infection is controlled by HDAC6, the deacetylase involved in the regulation of PA remained unknown. Here, we demonstrate that HDAC6 acts as a negative regulator of IAV infection by destabilizing PA. HDAC6 binds to and deacetylates PA, thereby promoting the proteasomal degradation of PA. Based on mass spectrometric analysis, Lys(664) of PA can be deacetylated by HDAC6, and the residue is crucial for PA protein stability. The deacetylase activity of HDAC6 is required for anti-IAV activity, because IAV infection was enhanced due to elevated IAV RNA polymerase activity upon HDAC6 depletion and an HDAC6 deacetylase dead mutant (HDAC6-DM; H216A, H611A). Finally, we also demonstrate that overexpression of HDAC6 suppresses IAV RNA polymerase activity, but HDAC6-DM does not. Taken together, our findings provide initial evidence that HDAC6 plays a negative role in IAV RNA polymerase activity by deacetylating PA and thus restricts IAV RNA transcription and replication. IMPORTANCE Influenza A virus (IAV) continues to threaten global public health due to drug resistance and the emergence of frequently mutated strains. Thus, it is critical to find new strategies to control IAV infection. Here, we discover one host protein, HDAC6, that can inhibit viral RNA polymerase activity by deacetylating PA and thus suppresses virus RNA replication and transcription. Previously, it was reported that IAV can utilize the HDAC6-dependent aggresome formation mechanism to promote virus uncoating, but HDAC6-mediated deacetylation of α-tubulin inhibits viral protein trafficking at late stages of the virus life cycle. These findings together will contribute to a better understanding of the role of HDAC6 in regulating IAV infection. Understanding the molecular mechanisms of HDAC6 at various periods of viral infection may illuminate novel strategies for developing antiviral drugs.

Sign in / Sign up

Export Citation Format

Share Document