The Plasmodesmal Localization Signal of TMV MP Is Recognized by Plant Synaptotagmin SYTA.

Plant viruses cross the barrier of the plant cell wall by moving through intercellular channels, termed plasmodesmata, to invade their hosts. They accomplish this by encoding movement proteins (MPs), which act to alter plasmodesmal gating. How MPs target to plasmodesmata is not well understood.

The Agrobacterium F-Box Protein Effector VirF Destabilizes the Arabidopsis GLABROUS1 Enhancer/Binding Protein-Like Transcription Factor VFP4, a Transcriptional Activator of Defense Response Genes.

Agrobacterium-mediated genetic transformation not only represents a technology of choice to genetically manipulate plants, but it also serves as a model system to study mechanisms employed by invading pathogens to counter the myriad defenses mounted against them by the host cell. Here, we uncover a new layer of plant defenses that is targeted by A. tumefaciens to facilitate infection.

Identification of Plasmodesmal Localization Sequences in Proteins In Planta.

Plasmodesmata (Pd) are cell-to-cell connections that function as gateways through which small and large molecules are transported between plant cells. Whereas Pd transport of small molecules, such as ions and water, is presumed to occur passively, cell-to-cell transport of biological macromolecules, such proteins, most likely occurs via an active mechanism that involves specific targeting signals on the transported molecule. The scarcity of identified plasmodesmata (Pd) localization signals (PLSs) has severely restricted the understanding of protein-sorting pathways involved in plant cell-to-cell macromolecular transport and communication.

Identification of a Functional Plasmodesmal Localization Signal in a Plant Viral Cell-To-Cell-Movement Protein.

Unlabelled: Our fundamental knowledge of the protein-sorting pathways required for plant cell-to-cell trafficking and communication via the intercellular connections termed plasmodesmata has been severely limited by the paucity of plasmodesmal targeting sequences that have been identified to date. To address this limitation, we have identified the plasmodesmal localization signal (PLS) in the Tobacco mosaic virus (TMV) cell-to-cell-movement protein (MP), which has emerged as the paradigm for dissecting the molecular details of cell-to-cell transport through plasmodesmata. We report here the identification of a bona fide functional TMV MP PLS, which encompasses amino acid residues between positions 1 and 50, with residues Val-4 and Phe-14 potentially representing critical sites for PLS function that most likely affect protein conformation or protein interactions.

Interaction of Arabidopsis Trihelix-Domain Transcription Factors VFP3 and VFP5 with Agrobacterium Virulence Protein VirF.

Agrobacterium is a natural genetic engineer of plants that exports several virulence proteins into host cells in order to take advantage of the cell machinery to facilitate transformation and support bacterial growth. One of these effectors is the F-box protein VirF, which presumably uses the host ubiquitin/proteasome system (UPS) to uncoat the packaging proteins from the invading bacterial T-DNA. By analogy to several other bacterial effectors, VirF most likely has several functions in the host cell and, therefore, several interacting partners among host proteins.

Synaptotagmin SYTA forms ER-plasma membrane junctions that are recruited to plasmodesmata for plant virus movement.

Metazoan synaptotagmins are Ca(2+) sensors that regulate exocytosis and endocytosis in various cell types, notably in nerve and neuroendocrine cells [1, 2]. Recently, the structurally related extended synaptotagmins were shown to tether the cortical ER to the plasma membrane in human and yeast cells to maintain ER morphology and stabilize ER-plasma membrane (ER-PM) contact sites for intracellular lipid and Ca(2+) signaling [3, 4]. The Arabidopsis synaptotagmin SYTA regulates endocytosis and the ability of plant virus movement proteins (MPs) to alter plasmodesmata to promote virus cell-to-cell transport [5, 6].

The Arabidopsis synaptotagmin SYTA regulates the cell-to-cell movement of diverse plant viruses.

Synaptotagmins are a large gene family in animals that have been extensively characterized due to their role as calcium sensors to regulate synaptic vesicle exocytosis and endocytosis in neurons, and dense core vesicle exocytosis for hormone secretion from neuroendocrine cells. Thought to be exclusive to animals, synaptotagmins have recently been characterized in Arabidopsis thaliana, in which they comprise a five gene family. Using infectivity and leaf-based functional assays, we have shown that Arabidopsis SYTA regulates endocytosis and marks an endosomal vesicle recycling pathway to regulate movement protein-mediated trafficking of the Begomovirus Cabbage leaf curl virus (CaLCuV) and the Tobamovirus Tobacco mosaic virus (TMV) through plasmodesmata (Lewis and Lazarowitz, 2010).

The tobamovirus Turnip Vein Clearing Virus 30-kilodalton movement protein localizes to novel nuclear filaments to enhance virus infection.

Plant viruses overcome the barrier of the plant cell wall by encoding cell-to-cell movement proteins (MPs), which direct newly replicated viral genomes to, and across, the wall. The paradigm for how a single MP regulates and coordinates these activities is the Tobacco mosaic virus (TMV) 30-kDa protein (MP(TMV)). Detailed studies demonstrate that TMV multiplies exclusively in the cytoplasm and have documented associations of MP(TMV) with endoplasmic reticulum (ER) membrane, microtubules, and plasmodesmata throughout the course of infection.

Arabidopsis synaptotagmin SYTA regulates endocytosis and virus movement protein cell-to-cell transport.

Synaptotagmins are calcium sensors that regulate synaptic vesicle exo/endocytosis. Thought to be exclusive to animals, they have recently been characterized in plants. We show that Arabidopsis synaptotagmin SYTA regulates endosome recycling and movement protein (MP)-mediated trafficking of plant virus genomes through plasmodesmata.

Functional transient genetic transformation of Arabidopsis leaves by biolistic bombardment.

Transient gene expression is an indispensable tool for studying functions of gene products. In the case of plants, transient introduction of genes by Agrobacterium infiltration is a method of choice for many species. However, this technique does not work efficiently in Arabidopsis leaf tissue, the most widely used model system for basic plant biology research.

The geminivirus nuclear shuttle protein NSP inhibits the activity of AtNSI, a vascular-expressed Arabidopsis acetyltransferase regulated with the sink-to-source transition.

DNA viruses can suppress or enhance the activity of cellular acetyltransferases to regulate virus gene expression and to affect cell cycle progression in support of virus replication. A role for protein acetylation in regulating the nuclear export of the bipartite geminivirus (Begomovirus) DNA genome was recently suggested by the findings that the viral movement protein NSP, a nuclear shuttle protein, interacts with the Arabidopsis (Arabidopsis thaliana) nuclear acetyltransferase AtNSI (nuclear shuttle protein interactor), and that this interaction and NSI expression are necessary for cabbage leaf curl virus infection and pathogenicity. To further investigate the consequences of NSI-NSP interactions, and the potential role of NSI in Arabidopsis growth and development, we used a reverse yeast two-hybrid selection and deletion analysis to identify NSI mutants that failed to interact with NSP, and promoter fusions to a uidA reporter gene to analyze the pattern of NSI expression during plant development.

Interaction of the movement protein NSP and the Arabidopsis acetyltransferase AtNSI is necessary for Cabbage leaf curl geminivirus infection and pathogenicity.

DNA viruses can modulate the activity of cellular acetyltransferases to regulate virus gene expression and to affect cell cycle progression in order to support virus replication. A role for protein acetylation in regulating the nuclear export of the bipartite geminivirus DNA genome was recently suggested by the findings that the viral movement protein NSP, which shuttles the viral genome between the nucleus and the cytoplasm, interacts with a novel Arabidopsis acetyltransferase, AtNSI, and the increased expression of AtNSI enhances susceptibility to Cabbage leaf curl virus infection. To further investigate the interaction of NSP and AtNSI and to establish the importance of this interaction in virus infections, we used a reverse yeast two-hybrid selection and deletion analysis to identify NSP mutants that were impaired in their ability to bind AtNSI.

The complete genome sequence of the Arabidopsis and tomato pathogen Pseudomonas syringae pv. tomato DC3000.

We report the complete genome sequence of the model bacterial pathogen Pseudomonas syringae pathovar tomato DC3000 (DC3000), which is pathogenic on tomato and Arabidopsis thaliana. The DC3000 genome (6.5 megabases) contains a circular chromosome and two plasmids, which collectively encode 5,763 ORFs.

A novel Arabidopsis acetyltransferase interacts with the geminivirus movement protein NSP.

Protein acetylation is important in regulating DNA-templated processes specifically and protein-protein interactions more generally in eukaryotes. The geminivirus movement protein NSP is essential for virus movement, shuttling the viral DNA genome between the nucleus and the cytoplasm. We have identified a novel Arabidopsis protein, AtNSI, that interacts with NSP.

Genomewide identification of Pseudomonas syringae pv. tomato DC3000 promoters controlled by the HrpL alternative sigma factor.

The ability of Pseudomonas syringae pv. tomato DC3000 to parasitize tomato and Arabidopsis thaliana depends on genes activated by the HrpL alternative sigma factor. To support various functional genomic analyses of DC3000, and specifically, to identify genes involved in pathogenesis, we developed a draft sequence of DC3000 and used an iterative process involving computational and gene expression techniques to identify virulence-implicated genes downstream of HrpL-responsive promoters.