Global phosphoproteomic analysis of Ebola virions reveals a novel role for VP35 phosphorylation-dependent regulation of genome transcription

Andrey Ivanov, Palaniappan Ramanathan, Christian Parry, Philipp A. Ilinykh, Xionghao Lin, Michael Petukhov, Yuri Obukhov, Tatiana Ammosova, Gaya K. Amarasinghe, Alexander Bukreyev, Sergei Nekhai

    Research output: Contribution to journalArticle

    Abstract

    Ebola virus (EBOV) causes severe human disease with a high case fatality rate. The balance of evidence implies that the virus circulates in bats. The molecular basis for host–viral interactions, including the role for phosphorylation during infections, is largely undescribed. To address this, and to better understand the biology of EBOV, the phosphorylation of EBOV proteins was analyzed in virions purified from infected monkey Vero-E6 cells and bat EpoNi/22.1 cells using high-resolution mass spectrometry. All EBOV structural proteins were detected with high coverage, along with phosphopeptides. Phosphorylation sites were identified in all viral structural proteins. Comparison of EBOV protein phosphorylation in monkey and bat cells showed only partial overlap of phosphorylation sites, with shared sites found in NP, VP35, and VP24 proteins, and no common sites in the other proteins. Three-dimensional structural models were built for NP, VP35, VP40, GP, VP30 and VP24 proteins using available crystal structures or by de novo structure prediction to elucidate the potential role of the phosphorylation sites. Phosphorylation of one of the identified sites in VP35, Thr-210, was demonstrated to govern the transcriptional activity of the EBOV polymerase complex. Thr-210 phosphorylation was also shown to be important for VP35 interaction with NP. This is the first study to compare phosphorylation of all EBOV virion proteins produced in primate versus bat cells, and to demonstrate the role of VP35 phosphorylation in the viral life cycle. The results uncover a novel mechanism of EBOV transcription and identify novel targets for antiviral drug development.

    Original languageEnglish (US)
    JournalCellular and Molecular Life Sciences
    DOIs
    StateAccepted/In press - Jan 1 2019

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    Keywords

    • Ebola virus
    • Phosphorylation
    • Replication
    • Transcription

    ASJC Scopus subject areas

    • Molecular Medicine
    • Molecular Biology
    • Pharmacology
    • Cellular and Molecular Neuroscience
    • Cell Biology

    Cite this

    Ivanov, A., Ramanathan, P., Parry, C., Ilinykh, P. A., Lin, X., Petukhov, M., Obukhov, Y., Ammosova, T., Amarasinghe, G. K., Bukreyev, A., & Nekhai, S. (Accepted/In press). Global phosphoproteomic analysis of Ebola virions reveals a novel role for VP35 phosphorylation-dependent regulation of genome transcription. Cellular and Molecular Life Sciences. https://doi.org/10.1007/s00018-019-03303-1