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Influenza A/Hong Kong/156/1997(H5N1) virus NS1 gene mutations F103L and M106I both increase IFN antagonism, virulence and cytoplasmic localization but differ in binding to RIG-I and CPSF30.

Influenza A/Hong Kong/156/1997(H5N1) virus NS1 gene mutations F103L and M106I both increase IFN antagonism, virulence and cytoplasmic localization but differ in binding to RIG-I and CPSF30.
Influenza A/Hong Kong/156/1997(H5N1) virus NS1 gene mutations F103L and M106I both increase IFN antagonism, virulence and cytoplasmic localization but differ in binding to RIG-I and CPSF30.
BACKGROUND:
The genetic basis for avian to mammalian host switching in influenza A virus is largely unknown. The human A/HK/156/1997 (H5N1) virus that transmitted from poultry possesses NS1 gene mutations F103L + M106I that are virulence determinants in the mouse model of pneumonia; however their individual roles have not been determined. The emergent A/Shanghai/patient1/2013(H7N9)-like viruses also possess these mutations which may contribute to their virulence and ability to switch species.
METHODS:
NS1 mutant viruses were constructed by reverse genetics and site directed mutagenesis on human and mouse-adapted backbones. Mouse infections assessed virulence, virus yield, tissue infection, and IFN induction. NS1 protein properties were assessed for subcellular distribution, IFN antagonism (mouse and human), CPSF30 and RIG-I domain binding, host transcription (microarray); and the natural prevalence of 103L and 106I mutants was assessed.
RESULTS:
Each of the F103L and M106I mutations contributes additively to virulence to reduce the lethal dose by >800 and >3,200 fold respectively by mediating alveolar tissue infection with >100 fold increased infectious yields. The 106I NS1 mutant lost CPSF binding but the 103L mutant maintained binding that correlated with an increased general decrease in host gene expression in human but not mouse cells. Each mutation positively modulated the inhibition of IFN induction in mouse cells and activation of the IFN-? promoter in human cells but not in combination in human cells indicating negative epistasis. Each of the F103L and M106I mutations restored a defect in cytoplasmic localization of H5N1 NS1 in mouse cells. Human H1N1 and H3N2 NS1 proteins bound to the CARD, helicase and RD RIG-I domains, whereas the H5N1 NS1 with the same consensus 103F and 106M mutations did not bind these domains, which was totally or partially restored by the M106I or F103L mutations respectively.
CONCLUSIONS:
The F103L and M106I mutations in the H5N1 NS1 protein each increased IFN antagonism and mediated interstitial pneumonia in mice that was associated with increased cytoplasmic localization and altered host factor binding. These mutations may contribute to the ability of previous HPAI H5N1 and recent LPAI H7N9 and H6N1 (NS1-103L+106M) viruses to switch hosts and cause disease in humans.
ns1, nonstructural protein 1, influenza a virus, host switch, nuclear, cytoplasmic, localization, nuclear export signal, host gene expression, virulence, interferon, h7n9, h5n1, h6n1, alveoli, tropism
1743-422X
Dankar, S.K.
f76625e5-fbf2-4b8d-ba54-ad7f4ae1257e
Miranda, Elena
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Forbes, N.E.
88871c6d-5c5d-49bc-9368-b153ec657bbe
Pelchat, M.
dcf08606-4802-4ae9-8ec0-06c20164e9dc
Tavassoli, Ali
d561cf8f-2669-46b5-b6e1-2016c85d63b2
Selman, M.
3269cd8a-49d7-439f-aa11-7af00a3dc3e8
Ping, J.
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Jia, J.
2ff2714e-41c6-4b7a-9e75-e4126b17d213
Brown, E.G.
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Dankar, S.K.
f76625e5-fbf2-4b8d-ba54-ad7f4ae1257e
Miranda, Elena
ad727f7b-cc62-4904-b46d-043f74ef88ed
Forbes, N.E.
88871c6d-5c5d-49bc-9368-b153ec657bbe
Pelchat, M.
dcf08606-4802-4ae9-8ec0-06c20164e9dc
Tavassoli, Ali
d561cf8f-2669-46b5-b6e1-2016c85d63b2
Selman, M.
3269cd8a-49d7-439f-aa11-7af00a3dc3e8
Ping, J.
012bcab0-e897-4c2c-a8b1-4af7c8304b15
Jia, J.
2ff2714e-41c6-4b7a-9e75-e4126b17d213
Brown, E.G.
2672ccba-42b9-44d8-943b-ddf9fe5802fa

Dankar, S.K., Miranda, Elena, Forbes, N.E., Pelchat, M., Tavassoli, Ali, Selman, M., Ping, J., Jia, J. and Brown, E.G. (2013) Influenza A/Hong Kong/156/1997(H5N1) virus NS1 gene mutations F103L and M106I both increase IFN antagonism, virulence and cytoplasmic localization but differ in binding to RIG-I and CPSF30. Virology Journal, 10 (243). (doi:10.1186/1743-422X-10-243). (PMID:23886034)

Record type: Article

Abstract

BACKGROUND:
The genetic basis for avian to mammalian host switching in influenza A virus is largely unknown. The human A/HK/156/1997 (H5N1) virus that transmitted from poultry possesses NS1 gene mutations F103L + M106I that are virulence determinants in the mouse model of pneumonia; however their individual roles have not been determined. The emergent A/Shanghai/patient1/2013(H7N9)-like viruses also possess these mutations which may contribute to their virulence and ability to switch species.
METHODS:
NS1 mutant viruses were constructed by reverse genetics and site directed mutagenesis on human and mouse-adapted backbones. Mouse infections assessed virulence, virus yield, tissue infection, and IFN induction. NS1 protein properties were assessed for subcellular distribution, IFN antagonism (mouse and human), CPSF30 and RIG-I domain binding, host transcription (microarray); and the natural prevalence of 103L and 106I mutants was assessed.
RESULTS:
Each of the F103L and M106I mutations contributes additively to virulence to reduce the lethal dose by >800 and >3,200 fold respectively by mediating alveolar tissue infection with >100 fold increased infectious yields. The 106I NS1 mutant lost CPSF binding but the 103L mutant maintained binding that correlated with an increased general decrease in host gene expression in human but not mouse cells. Each mutation positively modulated the inhibition of IFN induction in mouse cells and activation of the IFN-? promoter in human cells but not in combination in human cells indicating negative epistasis. Each of the F103L and M106I mutations restored a defect in cytoplasmic localization of H5N1 NS1 in mouse cells. Human H1N1 and H3N2 NS1 proteins bound to the CARD, helicase and RD RIG-I domains, whereas the H5N1 NS1 with the same consensus 103F and 106M mutations did not bind these domains, which was totally or partially restored by the M106I or F103L mutations respectively.
CONCLUSIONS:
The F103L and M106I mutations in the H5N1 NS1 protein each increased IFN antagonism and mediated interstitial pneumonia in mice that was associated with increased cytoplasmic localization and altered host factor binding. These mutations may contribute to the ability of previous HPAI H5N1 and recent LPAI H7N9 and H6N1 (NS1-103L+106M) viruses to switch hosts and cause disease in humans.

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Published date: 25 July 2013
Keywords: ns1, nonstructural protein 1, influenza a virus, host switch, nuclear, cytoplasmic, localization, nuclear export signal, host gene expression, virulence, interferon, h7n9, h5n1, h6n1, alveoli, tropism

Identifiers

Local EPrints ID: 355323
URI: http://eprints.soton.ac.uk/id/eprint/355323
ISSN: 1743-422X
PURE UUID: 43a9c8c8-bab6-4b34-8cd3-7a299f5972f6
ORCID for Ali Tavassoli: ORCID iD orcid.org/0000-0002-7420-5063

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Date deposited: 20 Aug 2013 09:18
Last modified: 15 Mar 2024 03:26

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Contributors

Author: S.K. Dankar
Author: Elena Miranda
Author: N.E. Forbes
Author: M. Pelchat
Author: Ali Tavassoli ORCID iD
Author: M. Selman
Author: J. Ping
Author: J. Jia
Author: E.G. Brown

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