Requirements for cell-to-cell movement of Barley stripe mosaic virus in monocot and dicot hosts

Summary The Barley stripe mosaic virus (BSMV) RNAss genome contains a series of overlapping open reading frames termed the triple gene block. The three most abundant proteins, betab, betac and betad, have been shown to have essential roles in infectivity, but their function in cell-to-cell movement has not previously been unambiguously defined, nor has the role of a minor translational read-through protein, betad' been characterized. We have now examined the direct involvement of each of these proteins in cell-to-cell movement in planta by engineering fusions of the green fluorescent protein (GFP) to a cysteine-rich regulatory protein designated gammab. Microscopic examination of inoculated and systemically infected barley and oat leaves revealed high levels of fluorescence that moved rapidly through the compact striate vascular tissue without infecting epidermal cells. In contrast, a radial pattern of fluorescence spread through a large number of epidermal and mesophyll cells before entry into the reticulate vascular tissue of the dicot hosts Nicotiania benthamiana and Chenopodium amaranticolor. Mutational analyses indicated that the betab, betac and betad proteins are each essential for cell-to-cell movement in local lesion and systemic hosts, whereas the betad' protein is dispensable. Collectively, these results demonstrate conclusively that the three major triple gene block-encoded proteins act in concert to mediate cell-to-cell movement of BSMV.