Copperacion Viral

The Plant Cell, Vol. 7, 1185-1194, August 1995 O 1995 American Society of Plant Physiologists

Cooperation in Vira1 Movement: The Geminivirus BL1 Movement Protein Interacts with BR1 and Redirects It from the Nucleus to the'Cell Periphery
Anton A. Sanderfoot and Sondra G. Lazarowitz'
Department of Microbiology, University of lllinois at Urbana-Champaign, Urbana, lllinois 61801

For plantviruses to systemically infect a host requires the active participation of viral-encoded movement proteins. It has been suggested that BL1 and BR1, the two movement proteins encoded by the bipartite geminivirus squash leaf curl virus (SqLCV), act cooperatively to facilitate movement of the viral single-stranded DNA genome from its site of replication in the nucleus to the cell periphery and acrossthe cell wall to adjacent uninfected cells. To better understand the mechanism of SqLCV movement, we investigated the ability of BL1 and BR1 to interact specifically with each other using transient expression assays in insect cells and Nicotiana tabacum cv Xanthi protoplasts. In this study, we showed that when individually expressed, BL1 is localized to the periphery and BR1 to nuclei in both cellsystems. However, when coexpressed in either cell type, BL1 relocalized BR1 from the nucleus to the cell periphery. This interaction was found to be specific for BL1 and BR1, because BL1 did not relocalize the SqLCV nuclear-localized AL2 or coat protein. In addition, mutations in BL1 known to affect viral infectivity and pathogenicity were found to be defective in either their subcellularlocalization or their ability to relocalize BR1, and, thus, identified regions of BL1 required for correct subcellular targeting or interaction with BR1. These findings extend our model for SqLCV movement, demonstrating that BL1 and BR1 appear to interact directly with each other to facilitate movement cooperatively and that BL1 is responsible for providing directionality to movement of the viral genome.INTRODUCTION To successfully infect a host plant and cause disease, a plant virus must cross the barrier of the cell wall to move cell to cell and reach the phloem sieve elements. From the sieve elements, it systemically infects the host. Plant viruses accomplish this by encoding movement proteins (MPs), which are nonstructural proteins that are not essential for viral replication orencapsidation but are required for systemic infection of the host (Atabekov and Dorokhov, 1984; Hull, 1991). Our current understanding of MP function is based primarily on molecu'lar studies of the single MP encoded by tobacco mosaic virus (TMV) and red clover necrotic mosaic virus. In vitro studies have shown each to be a sequence-nonspecific nucleic acid binding protein that appears to bind RNA in acooperative manner (Citovsky et al., 1990, 1992; Fujiwara et al., 1993; Giesman-Cookmeyerand Lommel, 1993). In transgenic plants, the TMV 30-kD MP localizes to secondary plasmodesmata in primarily nonvascular cells (Ding et al., 1992) and increases the size exclusion limit (SEL) of plasmodesmata 40-fold between mesophyll and bundle sheath cells. When microinjected into mesophyll cells, bacteriallyexpressed fusions of the TMV or red clover necrotic mosaic virus MP increase measured SELs of plasmodesmata, and each MP rapidly moves from cell to cell and functions to move single-strandedRNA (Fujiwara et al., 1993; Waigmann et al., 1994). Based on these studies, it has been proposed that these MPs are molecular chaperones that bind the viral RNA genome and target it to plasmodesmata, where the MPfunctions to increase the SEL and thus facilitates movement of the viral genome to adjacent cells. A second model has been proposed that is based primarily on electron microscopic studies of cauliflower mosaic virus (CaMV), cowpea mosaic virus, tomato ringspot virus, and tomato spotted wilt virus infections. In this model, the single viral-encoded MP is associatedwith tubular structures that...