Formation And Biological Function Of The Viroplasm Derived From Rice Black Streaked Dwarf Virus

Rice black-streaked dwarf virus(RBSDV), a member of the genus Fijivirus, infects maize and rice plants and causes tremendous yield loss in Asia. RBSDV is transmitted in nature by the small brown planthopper(laodelphax striatellus)and can be replicated in cells of both host plants and insect vector. It has genomic segments of10dsRNAs(S1-S10) encased in icosahedral double-shelled particles and encodes at least12primary transcription products, which are named from their respective genomic segments (P1to P10). RBSDV, like other member of the family Reovirus, is thought to replicate and assemble within cytoplasmic viral inclusion bodies, commonly called viroplasms. RBSDV P9-1encoded by the first open reading frame in the ninth segment(S9)of the genomic RNAs is an essential protein for the formation of viroplasms, but so far little is known about the other proteins of the viroplasm and the molecular interactions amongst its components. Fused with green fluorescent protein(GFP), P9-1is able to form inclusion bodies when expressed alone in Nicotiana benthamiana epidermal cell. Bimolecular fluorescence complementation (BiFC) assay in Nicotiana benthamiana epidermal cell suggested P9-1had the ability of self-interaction. Non-structural proteins were screened for their association with P9-1using a co-immunopreciptation assay, only P6was found to interact with P9-1. Actually the interaction of P5-1and P6on a par with the interaction of P6and P9-1was also confirmed by bimolecular fluorescence complementation assay in Spodoptera frugiperdd (sf9) cells through the baculovirus expression. Immunoelectron microscopy showed that P5-1, P6and P9-1co-localized in electron-dense inclusion bodies, indicating that P5-1and P6were ingredients of the viroplasms. In sf9cells, P6was diffusely distributed throughout the cytoplasm when expressed alone, but localized to inclusion when co-expressed with P9-1, suggesting that P6is recruited to viral inclusions bodies by binding to P9-1.