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Orchid Fleck Virus Structural Proteins N and P Form Intranuclear Viroplasm-Like Structures in the Absence of Viral Infection

文献类型： 外文期刊

作者： Kondo, Hideki 1 ; Chiba, Sotaro 1 ; Andika, Ida Bagus 1 ; Maruyama, Kazuyuki 1 ; Tamada, Tetsuo 1 ; Suzuki, Nobuhir 1 ;

作者机构： 1.Okayama Univ, Inst Plant Sci & Resources IPSR, Kurashiki, Okayama, Japan

2.Zhejiang Acad Agr Sci, Inst Virol & Biotechnol, Hangzhou, Zhejiang, Peoples R China

期刊名称：JOURNAL OF VIROLOGY （ 影响因子：5.103； 五年影响因子：5.078 ）

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收录情况： SCI

摘要： Orchid fleck virus (OFV) has a unique two-segmented negative-sense RNA genome that resembles that of plant nucleorhabdovi-ruses. In infected plant cells, OFV and nucleorhabdoviruses induce an intranuclear electron-lucent viroplasm that is believed tobe the site for virus replication. In this study, we investigated the molecular mechanism by which OFV viroplasms are produced in vivo. Among OFV-encoded proteins, the nucleocapsid protein (N) and the putative phosphoprotein (P) were present in nuclear fractions of OFV-infected Nicotiana benthamiana plants. Transient coexpression of N and P, in the absence of virus infection, was shown to be sufficient for formation of an intranuclear viroplasm-like structure in plant cells. When expressed independentlyas a fluorescent protein fusion product in uninfected plant cells, N protein accumulated throughout the cell, while P protein accumulated in the nucleus. However, the N protein, when coexpressed with P, was recruited to a subnuclear region to induce a large viroplasm-like focus. Deletion and substitution mutagenesis demonstrated that the P protein contains a nuclear localization signal (NLS). Artificial nuclear targeting of the N-protein mutant was insufficient for formation of viroplasm-like structures in the absence of P. A bimolecular fluorescence complementation assay confirmed interactions between the N and P proteins within subnuclear viroplasm-like foci and interactions of two of the N. benthamiana importin-a homologues with the P protein but not with the N protein. Taken together, our results suggest that viroplasm formation by OFV requires nuclear accumulation of both the N and P proteins, which is mediated by P-NLS, unlike nucleorhabdovirus viroplasm utilizing the NLS on protein N.

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