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Nucleoprotein phosphorylation site (Y385) mutation confers temperature sensitivity to influenza A virus due to impaired nucleoprotein oligomerization at a lower temperature

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Abstract

Mutations in viral proteins can lead to the cold adaption of influenza A virus and the cold-adapted virus is an important vaccination instrument. Here, we identify a novel strain of influenza A virus with cold sensitivity conferred by a mutation at a phosphorylation site within the nucleoprotein (NP). The highly conserved tyrosine 385 residue (Y385) of NP was identified as a phosphorylation site by mass spectrometry. The constructive NP phosphorylation mimicked by Y385E mutation was fatal for virus replication, while the continuous Y385 dephosphorylation mimicked by Y385F mutation had little impact on virus replication in vitro. Notably, the Y385F virus showed much lower replicative capacity in turbinates of mice compared with the wild type virus. Moreover, the replication of Y385F virus was significantly reduced in both A549 and MDCK cells grown at 33°C, when compared to that at 37°C. These results indicated that the Y385F mutation led to cold sensitivity of virus. We further found that the cold sensitivity of Y385F virus could be attributed to diminished NP oligomerization rather than any changes in intracellular localization. Taken together, these findings suggest that the phosphorylation of NP may be a critical factor that regulates the temperature sensitivity of influenza A virus.

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Acknowledgements

This work was supported by grants from the Strategic Priority Research Program of Chinese Academy of Sciences (XDB29010000), the National Natural Science Foundation of China (31630079, 31972657, and 31672531), the National Key Research and Development Program of China (2016YFD0500206), and the Mega-Project of Guangxi Natural Science Foundation (2015GXNSFEA139002). Jing Li is supported by Youth Innovation Promotion Association of CAS (2019091).

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Authors and Affiliations

  1. CAS Key Laboratory of Pathogenic Microbiology and Immunology, Institute of Microbiology, Chinese Academy of Sciences, Beijing, 100101, China

    Weinan Zheng, Liang Cui, Minghui Li, Yun Li, Wenhui Fan, Limin Yang, Jing Li, Lei Sun & Wenjun Liu

  2. University of Chinese Academy of Sciences, Beijing, 100101, China

    Liang Cui, Minghui Li, Lei Sun & Wenjun Liu

  3. State Key Laboratory for Conservation and Utilization of Subtropical Agro-Bioresourses & Laboratory of Animal Infectious Diseases, College of Animal Sciences and Veterinary Medicine, Guangxi University, Nanning, 530004, China

    Wenjun Liu

  4. Institute of Microbiology, Center for Biosafety Mega-Science, Chinese Academy of Sciences, Beijing, 100101, China

    Wenjun Liu

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  1. Weinan Zheng
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Correspondence to Lei Sun.

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Zheng, W., Cui, L., Li, M. et al. Nucleoprotein phosphorylation site (Y385) mutation confers temperature sensitivity to influenza A virus due to impaired nucleoprotein oligomerization at a lower temperature. Sci. China Life Sci. 64, 633–643 (2021). https://doi.org/10.1007/s11427-020-1727-y

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