Citation: Mengmeng Cao, Qiannan Jia, Jinghua Li, Lili Zhao, Li zhu, Yufan Zhang, Shan Li, Tao Deng. Naturally occurring PAE206K point mutation in 2009 H1N1 pandemic influenza viruses impairs viral replication at high temperatures .VIROLOGICA SINICA, 2024, 39(1) : 71-80.  http://dx.doi.org/10.1016/j.virs.2023.11.005

Naturally occurring PAE206K point mutation in 2009 H1N1 pandemic influenza viruses impairs viral replication at high temperatures

  • Corresponding author: Tao Deng, dengt@im.ac.cn
  • Received Date: 18 July 2023
    Accepted Date: 10 November 2023
    Available online: 16 November 2023
  • The emergence of influenza virus A pandemic H1N1 in April 2009 marked the first pandemic of the 21st century. In this study, we observed significant differences in the polymerase activities of two clinical 2009 H1N1 influenza A virus isolates from Chinese and Japanese patients. Sequence comparison of the three main protein subunits (PB2, PB1, and PA) of the viral RNA-dependent RNA polymerase complex and subsequent mutational analysis revealed that a single amino acid substitution (E206K) was responsible for the observed impaired replication phenotype. Further in vitro experiments showed that presence of PAE206K decreased the replication of influenza A/WSN/33 virus in mammalian cells and a reduction in the virus’s pathogenicity in vivo. Mechanistic studies revealed that PAE206K is a temperature-sensitive mutant associated with the inability to transport PB1–PA complex to the nucleus at high temperature (39.5 ℃). Hence, this naturally occurring variant in the PA protein represents an ideal candidate mutation for the development of live attenuated influenza vaccines.

  • 加载中
  • 10.1016j.virs.2023.11.005-ESM.docx
    1. Avit, D.F.S., Jain, S., Finelli, L., Shaw, M.W., Lindstrom, S., Garten, R.J., Gubareva, L.V., Xu, X., Bridges, C.B., Uyeki, T.M., 2009. Emergence of a novel swine-origin influenza A (H1N1) virus in humans. N. Engl. J. Med. 360, 2605-2615.

    2. Bussey, K.A., Desmet, E.A., Mattiacio, J.L., Hamilton, A., Bradel-Tretheway, B., Bussey, H.E., Kim, B., Dewhurst, S., Takimoto, T., 2011. PA residues in the 2009 H1N1 pandemic influenza virus enhance avian influenza virus polymerase activity in mammalian cells. J. Virol. 85, 7020-7028.

    3. Da Costa, B., Sausset, A., Munier, S., Ghounaris, A., Naffakh, N., Le Goffic, R., Delmas, B., 2015. Temperature-sensitive mutants in the influenza A virus RNA polymerase: Alterations in the PA linker reduce nuclear targeting of the PB1-PA dimer and result in viral attenuation. J. Virol. 89, 6376-6390.

    4. Fan, H., Walker, A.P., Carrique, L., Keown, J.R., Martin, I.S., Karia, D., Sharps, J., Hengrung,N., Pardon, E., Steyaert, J., Grimes, J.M., Fodor, E., 2019. Influenza A virus RNA polymerase structures provide insights into viral genome replication. Nature. 573, 287-290.

    5. Fan, S., Hatta, M., Kim, J.H., Le, M.Q., Neumann, G., Kawaoka, Y., 2014. Amino acid changes in the influenza A virus PA protein that attenuate avian H5N1 viruses in mammals. J. Virol. 88, 13737-13746.

    6. Fodor E., 2013. The RNA polymerase of influenza A virus: Mechanisms of viral transcription and replication. Acta virol. 57, 113-122.

    7. Fodor, E., Crow, M., Mingay, L.J., Deng, T., Sharps, J., Fechter, P., Brownlee, G.G., 2002. A single amino acid mutation in the PA subunit of the influenza virus RNA polymerase inhibits endonucleolytic cleavage of capped RNAs. J. Virol. 76, 8989 -9001.

    8. Hengrung, N., Omari, K.E., Martin, I.S., Vreede, F.T., Cusack, S., Rambo, R.P., Vonrhein, C., Bricogne, G., Stuart, D.I., Grimes, J.M., Fodor, E., 2015. Crystal structure of the RNA-dependent RNA polymerase from influenza C virus. Nature. 527, 114-117.

    9. Hoffmann, E., Neumann, G., Kawaoka, Y., Hobom, G., Webster, R.G., 2000. A DNA transfection system for generation of influenza A virus from eight plasmids. Proc. Natl. Acad. Sci. U. S. A. 97, 6108-6113.

    10. Hsieh, E.F., Lin, S.J., Mok, C.K., Chen, G.W., Huang, C.H., Wang, Y.C., Chen, T.C., Chen, C.J., Ojcius, D.M., Shih, S.R., 2011. Altered pathogenicity for seasonal influenza virus by single reassortment of the RNP genes derived from the 2009 pandemic influenza virus. J. Infect. Dis. 204, 864-872.

    11. Hu, J., Liu, X., 2015. Crucial role of PA in virus life cycle and host adaptation of influenza A virus. Med. Microbiol. Immunol. 204, 137-149.

    12. Huarte, M., Sanz-Ezquerro, J.J., Roncal, F., Ortin, J., Nieto, A., 2001. PA subunit from influenza virus polymerase complex interacts with a cellular protein with homology to a family of transcriptional activators. J. Virol. 75, 8597-8604.

    13. Huarte, M., Falcon, A., Nakaya, Y., Ortin, J., Garcia-Sastre, A., Nieto, A., 2003. Threonine 157 of influenza virus PA polymerase subunit modulates RNA replication in infectious viruses. J. Virol. 77, 6007-6013.

    14. Hutchinson, E.C., Fodor, E., 2012. Nuclear import of the influenza A virus transcriptional machinery. Vaccine. 30,7353-7358.

    15. Hsu, W.B., Shih, J.L., Shih, J.R., Du, J.L., Teng, S.C., Huang, L.M., Wang, W.B., 2013. Cellular protein HAX1 interacts with the influenza A virus PA polymerase subunit and impedes its nuclear translocation. J. Virol. 87, 110-123.

    16. Krumbholz, A., Philipps, A., Oehring, H., Schwarzer, K., Eitner, A., Wutzler, P., Zell, R., 2011. Current knowledge on PB1-F2 of influenza A viruses. Med. Microbiol. Immunol. 200, 69-75.

    17. Lee, I., Il Kim, J., Park, S., Bae, J.Y., Yoo, K., Yun, S.H., Lee, J.Y., Kim, K., Kang, C., Park, M.S., 2017. Single PA mutation as a high yield determinant of avian influenza vaccines. Sci. Rep. 7, 40675.

    18. Lopez, C.E., Legge, K.L., 2020. Influenza A virus vaccination: Immunity, protection, and recent advances toward A universal vaccine. Vaccines. 8, 434.

    19. Lukarska, M., Fournier, G., Pflug, A., Resa-Infante, P., Reich, S., Naffakh, N., Cusack, S., 2017. Structural basis of an essential interaction between influenza polymerase and Pol II CTD. Nature. 541, 117-121.

    20. Mehle, A., Dugan, V.G., Taubenberger, J.K., Doudna, J.A., 2012. Reassortment and mutation of the avian influenza virus polymerase PA subunit overcome species barriers. J. Virol. 86, 1750-1757.

    21. Meyer, L., Sausset, A., Sedano, L., Da Costa, B., Le Goffic, R., Delmas, B., 2016. Codon deletions in the influenza A virus PA gene generate temperature-sensitive viruses. J. Virol. 90, 3684-3693.

    22. Mishra, A.C., Chadha, M.S., Choudhary, M.L., Potdar, V.A., 2010. Pandemic influenza (H1N1) 2009 is associated with severe disease in India. PLOS ONE. 5, e10540.

    23. Muramoto, Y., Noda, T., Kawakami, E., Akkina, R., Kawaoka, Y., 2013. Identification of novel influenza A virus proteins translated from PA mRNA. J. Virol. 87, 2455-2462.

    24. Nogales, A., Martinez-Sobrido, L., Chiem, K., Topham, D.J., DeDiego, M.L., 2018. Functional evolution of the 2009 pandemic H1N1 influenza virus NS1 and PA in humans. J. Virol. 92, e01206-e01218.

    25. Palese, P., Shaw. M.L., 2013. Orthomyxoviridae, In Knipe, D.M., Howley, P.M., Cohen, J.I., Griffifin, D.E., Lamb, R.A., Martin, M.A., Racaniello, V.R., Roizman, B. (Eds.), Fields virology, 6th ed. Lippincott Williams and Wilkins, Philadelphia, pp. 1151-1185.

    26. Perales, B., Sanz-Ezquerro, J.J., Gastaminza, P., Ortega, J., Santaren, J.F., Ortin, J., Nieto, A., 2000. The replication activity of influenza virus polymerase is linked to the capacity of the PA subunit to induce proteolysis. J. Virol. 74, 1307-1312.

    27. Rodriguez-Frandsen, A., Alfonso, R., Nieto, A., 2015. Influenza virus polymerase: Functions on host range, inhibition of cellular response to infection and pathogenicity. Virus Res. 209, 23-38.

    28. Selman, M., Dankar, S.K., Forbes, N.E., Jia, J.J., Brown, E.G., 2012. Adaptive mutation in influenza A virus non-structural gene is linked to host switching and induces a novel protein by alternative splicing. Emerg. Microbes Infect. 1, e42.

    29. Sherbany, H., McCauley, J., Meningher, T., Hindiyeh, M., Dichtiar, R., Markovich, M.P., Mendelson, E., Mandelboim, M., 2014. Return of pandemic H1N1 influenza virus. BMC Infect. Dis. 14, 710.

    30. Smith, G.J., Vijaykrishna, D., Bahl, J., Lycett, S.J., Worobey, M., Pybus, O.G., Ma, S.K., Cheung, C.L., Raghwani, J., Bhatt, S., Peiris, J.S., Guan, Y., Rambaut, A., 2009. Origins and evolutionary genomics of the 2009 swine-origin H1N1 influenza A epidemic. Nature. 459, 1122-1125.

    31. Song, M.S., Pascua, P.N., Lee, J.H., Baek, Y.H., Lee, O.J., Kim, C.J., Kim, H., Webby, R.J., Webster, R.G., Choi, Y.K., 2009. The polymerase acidic protein gene of influenza A virus contributes to pathogenicity in a mouse model. J. Virol. 83, 12325-12335.

    32. Stevaert, A., Naesens, L., 2016. The influenza virus polymerase complex: An update on its structure, functions, and significance for antiviral drug design. Med. Res. Rev. 36, 1127-1173.

    33. Stubbs, T.M., Te Velthuis. A.J., 2014. The RNA-dependent RNA polymerase of the influenza A virus. Future Virol. 9, 863-876.

    34. Subbarao, K., 2020. Live attenuated influenza vaccines for pandemic preparedness. J. Pediatr. Infect. Dis. Soc. 9, S15-S18.

    35. Treanor, J., 2020. History of live, attenuated influenza vaccine. J. Pediatr. Infect. Dis. Soc. 9, S3-S9.

    36. Trifonov, V., Khiabanian, H., Rabadan, R., 2009. Geographic dependence, surveillance, and origins of the 2009 influenza A (H1N1) virus. N. Engl. J. Med. 361, 115-119.

    37. Wang, L., Wu, A., Wang, Y.E., Quanquin, N., Li, C., Wang, J., Chen, H.W., Liu, S., Liu, P., Zhang, H., Qin, F.X.F., Jiang, T.J., Cheng, G.H., 2015. Functional genomics reveals linkers critical for influenza virus polymerase. J. Virol. 90, 2938-2947.

    38. Watanabe, T., Watanabe, S., Kawaoka, Y., 2010. Cellular networks involved in the influenza virus life cycle. Cell Host Microbe. 7, 427-439.

    39. Weinberg, G.A., 2020. Live, attenuated influenza vaccine: Present and future roles. J. Pediatr. Infect. Dis. Soc. 9, S1-S2.

    40. Wise, H.M., Hutchinson, E.C., Jagger, B.W., Stuart, A.D., Kang, Z.H., Robb, N., Schwartzman, L.M., Kash, J.C., Fodor, E., Firth, A.E., Gog, J.R., Taubenberger, J.K., Digard, P., 2012. Identification of a novel splice variant form of the influenza A virus M2 ion channel with an antigenically distinct ectodomain. PLOS Pathog. 8, e1002998.

    41. Xu, G., Zhang, X., Gao, W., Wang, C., Wang, J., Sun, H., Sun, Y., Guo, L., Zhang, R., Chang, K.C., Liu, J.H., Pu, J., 2016. Prevailing PA mutation K356R in avian influenza H9N2 virus increases mammalian replication and pathogenicity. J. Virol. 90, 8105-8114.

    42. Yamaji, R., Yamada, S., Le, M.Q., Ito, M., Sakai-Tagawa, Y., Kawaoka, Y., 2015. Mammalian adaptive mutations of the PA protein of highly pathogenic avian H5N1 influenza virus. J. Virol. 89, 4117-4125.

    43. Yang, H.,Dong, Y. R., Bian, Y., Huo, C.Z., Zhu, C.C., Qin, T., Chen, S.J, Peng, D.X., Liu, X.F., 2023. The synergistic effect of residues 32T and 550L in the PA protein of H5 subtype avian influenza virus contributes to viral pathogenicity in mice. PLoS Pathog. 19, e1011489.

    44. Yi, C.Y., Zhao, Z.Z., Wang, S.Y., Sun, X., Zhang, D., Sun,X.M., Zhang,A.D., Jin, M.D., 2017. Influenza A Virus PA Antagonizes Interferon-βby Interacting with Interferon Regulatory Factor 3. Front. Immunol. 8, 1051-1059.

    45. Zhang, H., Chen, L., 2009. Possible origin of current influenza A H1N1 viruses. Lancet Infect. Dis. 9, 456-457.

    46. Zhang, W., Tu, J., Zhao, Z., Chen, H., Jin, M., 2012. The new temperature-sensitive mutation PA-F35S for developing recombinant avian live attenuated H5N1 influenza vaccine. Virol. J. 9, 97.

  • 加载中

Article Metrics

Article views(2944) PDF downloads(17) Cited by()

Related
Proportional views

    Naturally occurring PAE206K point mutation in 2009 H1N1 pandemic influenza viruses impairs viral replication at high temperatures

      Corresponding author: Tao Deng, dengt@im.ac.cn
    • a. National Institute of Pathogen Biology, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, 100730, China;
    • b. CAS Key Laboratory of Pathogen Microbiology and Immunology, Institute of Microbiology, Chinese Academy of Sciences, Beijing, 100101, China

    Abstract: The emergence of influenza virus A pandemic H1N1 in April 2009 marked the first pandemic of the 21st century. In this study, we observed significant differences in the polymerase activities of two clinical 2009 H1N1 influenza A virus isolates from Chinese and Japanese patients. Sequence comparison of the three main protein subunits (PB2, PB1, and PA) of the viral RNA-dependent RNA polymerase complex and subsequent mutational analysis revealed that a single amino acid substitution (E206K) was responsible for the observed impaired replication phenotype. Further in vitro experiments showed that presence of PAE206K decreased the replication of influenza A/WSN/33 virus in mammalian cells and a reduction in the virus’s pathogenicity in vivo. Mechanistic studies revealed that PAE206K is a temperature-sensitive mutant associated with the inability to transport PB1–PA complex to the nucleus at high temperature (39.5 ℃). Hence, this naturally occurring variant in the PA protein represents an ideal candidate mutation for the development of live attenuated influenza vaccines.

    Reference (46) Relative (20)

    目录

    /

    DownLoad:  Full-Size Img  PowerPoint
    Return
    Return