The Coat and Cylindrical Inclusion Proteins of a Potyvirus Are Associated with Connections between Plant Cells

Abstract

The subcellular locations of two potyviral proteins, the coat (CP) and nonstructural cylindrical inclusion (CI) proteins of tobacco vein mottling virus (TVMV), during early stages in the development of systemic infections in plants, have been investigated. Ultrathin sections of newly emerged leaves in infected plants were treated with antibodies specific to these proteins and then with gold-labeled secondary antibodies and examined by electron microscopy. CI was detected near plasmodesmatal connections between mesophyll cells prior to the appearance of CP or any virus-induced features or effects. Further accumulation of CI was evident in the form of conical structures, many of which appeared to penetrate the cell wall and to be connected to cones in neighboring cells. Prior to its appearance in other parts of the cells, the viral CP was detected, often in linear arrays, near the vertices or inside the cones and in plasmodesmata. In situ hybridization analysis of similar tissue sections with a TVMV RNA-specific oligoribonucleotide probe revealed the presence of the viral RNA in plasmodesmata. These results lend support to the notion that the formation of specific structures by potyviral CI proteins is required for and plays a direct role in the intercellular passage of viral genetic material, in the form of virus particles or complexes containing viral CP and RNA, in infected plants.

In Situ Localization and Tissue Distribution of the Replication-Associated Proteins of Cucumber Mosaic Virus in Tobacco and Cucumber

“…For ISH, the protocol described by Rodriguez-Cerezo et al (26), with some modifications, was used as standard. Briefly, grids were incubated in prehybridization buffer (50% deionized formamide, 12.5% dextran sulfate, 0.2 M NaCl, 6 mM PIPES, 1 mM EDTA, 6 mg of tRNA/ml) for 2 h at 42°C and hybridized overnight at the same temperature in the same buffer containing 1 to 5 ng of the probe/l.…”

Cell-to-cell movement of three genera (+) ss RNA plant viruses

“…In combination with ultrastructural analyses and genetic data Carrington et al (1998) results support a model in which Potyvirus CI protein interacts directly with PD and capsid protein within ribonucleoprotein complexes to facilitate Potyvirus cell-tocell movement. Ultrastructural research of tissues at an early stage of infection with immunogold labeling of specific Potyvirus proteins, have supported the role CI in local transport with cooperation in CP (Rodriguez-Cerezo et al 1997;Roberts et al 1998). Observation of young tobacco leaves infected with TVMV (Tobacco vein mottling virus; Rodriguez-Cerezo et al 1997) or cells during advanced PSbMV (Pea seed-borne mosaic virus) infection in pea cotyledons showed that cytoplasmic inclusions immunolabeled for CI protein and CP were attached to the plasma membrane, close to or over the plasmodesmal openings.…”

“…Ultrastructural research of tissues at an early stage of infection with immunogold labeling of specific Potyvirus proteins, have supported the role CI in local transport with cooperation in CP (Rodriguez-Cerezo et al 1997;Roberts et al 1998). Observation of young tobacco leaves infected with TVMV (Tobacco vein mottling virus; Rodriguez-Cerezo et al 1997) or cells during advanced PSbMV (Pea seed-borne mosaic virus) infection in pea cotyledons showed that cytoplasmic inclusions immunolabeled for CI protein and CP were attached to the plasma membrane, close to or over the plasmodesmal openings. The plasmodesmal aperture contained CP and also viral RNA during TVMV tobacco infection.…”

Detection, Understanding and Controlof Soybean Mosaic Virus

“…The potyvirus CI has been suggested to play a role in cell to cell movement (Laín et al 1990;Eagles et al 1994;Klein et al 1994). High-resolution ultrastructural analyses indicate that CI forms the cone-shaped structures at the cell periphery adjacent to PD (Rodríguez-Cerezo et al 1997;Roberts et al 1998Roberts et al , 2003. In the case of the newly found P3N-PIPO protein, it has been shown that mutation of the putative SMV PIPO domain impeded cell-to-cell movement (Wen et al,2010) and that P3N-PIPO mediated the formation of CI conical structures at PD (Wei et al, 2010b).…”