Identification and characterization of the interaction between viroplasm-associated proteins from two different plant-infecting reoviruses and eEF-1A of rice.

A rice protein homologous to eukaryotic translation elongation factor 1A (eEF-1A) was found to interact with the Pns6 of rice ragged stunt virus (RRSV), the type member of the genus Oryzavirus, family Reoviridae, in yeast two-hybrid screening. The interaction between the rice protein, designated OseEF-1A, and RRSV Pns6 was confirmed by bimolecular fluorescence complementation. Besides Pns6, OseEF-1A also interacted with the viroplasm matrix protein, Pns10, of RRSV. When expressed together, OseEF-1A co-localized with RRSV Pns10 in epidermal cells of Nicotiana benthamiana. Pns6 of southern rice black-streaked dwarf virus (SRBSDV), a newly reported member of the genus Fijivirus, family Reoviridae, was the only non-structural SRBSDV protein studied here that also interacted with OseEF-1A. Like Pns6 of rice black-streaked dwarf virus (RBSDV), SRBSDV Pns6 interacted with itself and co-localized with Pns9-1 in N. benthamiana. In the presence of Pns6, OseEF-1A co-localized with Pns9-1, the putative viroplasm matrix protein of SRBSDV.

Expression of Rice Gall Dwarf Virus Outer Coat Protein Gene (S8) in Insect Cells / 中国病毒学·英文版

To obtain the P8 protein of Rice gall dwarf virus (RGDV) with biological activity, its outer coat protein gene S8 was expressed in Spodoptera frugiperda (Sf9) insect cells using the baculovirus expression system. The S8 gene was subcloned into the pFastBacTM1 vector, to produce the recombinant baculovirus transfer vector pFB-S8. After transformation, pFB-S8 was introduced into the competent cells (E. coli DH10Bac) containing a shuttle vector, Bacmid, generating the recombinant bacmid rbpFB-S8. After being infected by recombinant baculovirus rvpFB-S8 at different multiplicities of infection, Sf9 cells were collected at different times and analyzed by SDS-PAGE, Western blotting and immunofluorescence microscopy. The expression level of the P8 protein was highest between 48-72 h after transfection of Sf9 cells. Immunofluorescence microscopy showed that P8 protein of RGDV formed punctate structures in the cytoplasm of Sf9 cells.

Characterization and Amino Acid Sequence of y3,an Antiviral Protein from Mushroom Coprinus comatus / 中国生物化学与分子生物学报

Many proteins from mushrooms can be used as potential antiviral agents and in plant protection.An antiviral protein,designated as y3,Was isolated from fruiting bodies of the fungus Coprinus comatus by fastflow ion-exchange column chromatography coupled with high-resolution molecular sieve chromatography.This glycoprotein was detected in both fruiting body and mycelium by Western blot analysis.According to its Nterminal sequence.an amino acid sequence and a partial cDNA sequence of the y3 gene were obtained.Protein y3 at 2.0μg/ml achieved 50.0% inhibition of tobacco mosaic virus(TMV,20μg/ml)lesions in Nicotiana glutinosa leaves.It also inhibited multiplication of TMV in Nicotiana tabacum Var.K326.

Characterization of Proteins and Nucleic Acid of Rice Stripe Virus / 中国病毒学

An isolate of rice stripe virus (designated as RSV-YL) was purified. The particles showed to be pleomorphisms under electron microscope, mainly branched filaments of about 80-250 nm in length and about 8 nm in width. There are also some open circular filaments of 3 nm and 8 nm in width, and some filaments of 13 nm in width and 130-190 nm in length. The basic morphism of RSV particles should be filaments of 3 nm in width and various length. By SDS-PAGE analysis, the molecular weight of disease-specific protein (SP) encoded by vRNA4 was 19.9 kDa and that of coat protein (CP) encoded by vcRNA3 was 33.6 kDa. When nucleic acid extracted from the purified RSV was electrophoresed under nondenaturing condition, the size of four dsRNAs (designated as dsRNA1-4 in order of decreasing size) was 4.9×106,2.7×106,2.0×106 and 1.7×106 Da, respectively, and that of four ssRNAs (designated as ssRNA1-4 in order of decreasing size) was 3.0×106,1.2×106,0.9×106 and 0.8×106 Da, respectively. A fifth segment with a size of 0.58×106 Da identified as ssRNA5 associated with the purified virus sometimes. The antiserum against the coat protein further purified by preparative electrophoresis was raised and used to investigate the serological relationships between RSV-CP and RSV-SP, CP and SP of rice grassy stunt virus (RGSV) which is also a member of Tenuivirus. The results showed that RSV-CP had no serological reaction with SP of RSV and PGSV, but could weakly react with antiserum of RGSV-CP, which confirmed that there is distantly evolutionary relationship between RGSV and RSV.