Dynamics of Bean Dwarf Mosaic Geminivirus Cell-to-Cell and Long-Distance Movement in Phaseolus vulgaris Revealed, Using the Green Fluorescent Protein

Abstract

The cell-to-cell and long-distance movement of the bipartite geminivirus, bean dwarf mosaic (BDMV), in Phaseolus vulgaris plants was examined with the noninvasive reporter, the green fluorescent protein (GFP). A modified GFP gene (mGFP4) was inserted into the BDMV DNA-A component in place of the coat protein gene (BDMVA-mGFP4), and particle bombardment was used to introduce viral DNA into bean seedlings (radicle and hypocotyl tissues). Fluorescence analysis of GFP expressed from BDMVA-mGFP4 established that particle bombardment introduced viral DNA only into epidermal cells, and the requirement for the DNA-B-encoded proteins (BV1 and BC1) in the cell-to-cell movement of BDMVA-mGFP4. This GFP reporter system was used to follow the viral infection process from the seedling stage throughout the entire plant life cycle. In inoculated hypocotyls, BDMV moved from cell to cell through the cortex and showed a striking phloem tropism. Upon entry into phloem tissues, BDMV moved rapidly toward the root via the long-distance transport system, and toward the shoot apex by a combination of cell-to-cell and long-distance movement. Analysis of GFP distribution in systemically infected tissues revealed that BDMV was restricted to phloem cells in both roots and stems. In systemically infected primary and trifoliolate leaves, BDMV infected phloem cells associated with all vein orders (first through fifth), and the capacity of BDMV to exit from phloem tissue into nonphloem cells was correlated with the stage of plant development. Finally, fluorescence analysis of GFP in reproductive tissues established that BDMV infected flower, pod, and seed-coat tissues, but was excluded from the embryo.