No abstract

A microinjection technique was devised for inoculation of single Nicotiana clevelandii leaf trichome cells with virus particles. By removing inoculated trichomes at various times after microinjection, it was shown that at least 4 hr were required for tobacco rattle virus (TRV; tobravirus group) to move out of primarily inoculated cells. Effects of the early stages of TRV infection on plasmodesmatal permeability were examined by microinjection of fluorochrome-labeled molecules. Fluorescein-labeled insulin A chain (Mr, 2921) and fluorescein-labeled dextran (Mr, 4400) were observed to pass out of individual N. clevelandii trichome cells that had been inoculated with TRV by microinjection 5 hr previously. By contrast, Lucifer Yellow CH-labeled dextran (Mr, 10,000) was restricted to the inoculated cell. None of these fluorescent probes were able to move out of uninoculated cells or out of cells that had been inoculated with TRV only 2 hr previously. The movement of macromolecules through plasmodesmata, therefore, coincided with and probably resulted from cell-to-cell movement of TRV. The results are discussed with reference to the interaction of viruses and plasmodesmata and mechanisms of intercellular virus movement.

A microinjection technique was devised for inoculation of single Nicotiana clevelandii leaf trichome cells with virus particles. By removing inoculated trichomes at various times after microinjection, it was shown that at least 4 hr were required for tobacco rattle virus (TRV; tobravirus group) to move out of primarily inoculated cells. Effects of the early stages of TRV infection on plasmodesmatal permeability were examined by microinjection of fluorochrome-labeled molecules. Fluorescein-labeled insulin A chain (Mr, 2921) and fluorescein-labeled dextran (Mr, 4400) were observed to pass out of individual N. clevelandii trichome cells that had been inoculated with TRV by microinjection 5 hr previously. By contrast, Lucifer Yellow CH-labeled dextran (Mr, 10,000) was restricted to the inoculated cell. None of these fluorescent probes were able to move out of uninoculated cells or out of cells that had been inoculated with TRV only 2 hr previously. The movement of macromolecules through plasmodesmata, therefore, coincided with and probably resulted from cell-to-cell movement of TRV. The results are discussed with reference to the interaction of viruses and plasmodesmata and mechanisms of intercellular virus movement.

The masked and U1 strains of tobacco mosaic tobamovirus differ in symptom phenotype and in phloem-dependent accumulation in tobacco. The symptom phenotype is determined by eight amino acids in the 126- and 183-kDa proteins that differ between the two strains. In this study, slow phloem-dependent accumulation of the masked strain was shown to be determined by these same eight amino acids, but some symptomatically severe mutants altered at specific positions within the eight amino acids were inefficient in phloem-dependent accumulation. Therefore, the appearance of severe symptoms does not require rapid phloem-dependent accumulation. There was no consistent relationship between the accumulation of virus coat protein, movement protein, or 126- and 183-kDa protein in inoculated protoplasts and the efficiency of phloem-dependent accumulation in stem tissue. Therefore, the difference in phloem-dependent accumulation between the masked strain, its mutants, and the U1 strain in most instances resulted from the functional competence of the 126- and/or 183-kDa proteins or a host response to their change and not from their quantities or the quantities or functional competence of the movement proteins or coat proteins.

The molecular weight exclusion limit of plasmodesmata in subveinal epidermal cells of Nicotiana clevelandii (Gray) leaves was estimated by microinjection and fluorescence microscopy using fluorescein isothiocyanate-peptide conjugates, carboxyfluorescein and Lucifer Yellow CH. The largest fluorochrome which moved symplastically between cells had a molecular weight of 749, although movement did not appear to depend purely on molecular weight parameters. Systemic infection of plants by tobacco rattle tobravirus, tomato black ring nepovirus or potato Y potyvirus did not alter the limits of plasmodesmatal conductance of the fluorochromes. However, carrot mottle umbravirus and groundnut rosette umbravirus diminished the symplastic mobility of some fluorescent tracers. These results imply that intercellular movement of these viruses does not involve a long-lasting increase in the plasmodesmatal molecular size exclusion limit.