Intracellular trafficking of Potato leafroll virus movement protein in transgenic Arabidopsis

Intracellular trafficking of viral movement proteins (MPs) in plants has mainly been studied using Tobacco mosaic virus MP30 (TMV MP30) as a model system. Because of the limitations of TMV MP30 expression in Arabidopsis thaliana, these studies have mostly been restricted to tobacco plants. Here we present data on the analysis of transgenic Arabidopsis plants expressing Potato leafroll virus 17-kDa movement protein (MP17) fused to green fluorescent protein. MP17 localizes to secondary branched plasmodesmata (PD) in source but not to simple PD in sink tissues, where MP17 is believed to be degraded by proteolysis. To unravel the intracellular transport path of MP17, we analyzed the relevance of the cytoskeleton and of the secretory pathway on MP17 targeting. To this end, a new incubation system for in vivo analysis of immediate and long-term responses of whole Arabidopsis plants to inhibitor treatments was established. Microscopic and histochemical analysis showed that MP17 is targeted to PD in an actin- and endoplasmic reticulum-Golgi-dependent manner. In contrast, degradation of MP17 in sink tissues required intact microtubules and occurred at 26S proteasomes. Interestingly, inhibition of the 26S proteasome led to aggregation of MP17 in aggresome-like structures. Formation of these structures could be inhibited by colchicine, as was shown for aggresomes in mammalian cells.     

Nonspecific intercellular protein trafficking probed by green-fluorescent protein in plants

Summary Plasmodesmata mediate intercellular transport of proteins, nucleic acids, and small molecules in plants. We show that transiently produced green-fluorescent protein (GFP) trafficked intercellularly in the epidermis of sink leaves, but not of source leaves, in tobacco and cucumber. In contrast, the protein did not traffic in either sink or source leaves of tomato. On the other hand, the protein spread extensively from cell to cell in the epidermis of all leaves and stems ofArabidopsis thaliana as well as in young hypocotyls and cotyledons of tomato and cucumber. GFP could traffic from epidermis to ground tissues in hypocotyls but not in cotyledons of cucumber. GFP fused to a number of mutant forms of the cucumber mosaic virus 3a movement protein (CMV 3a MP) failed to traffic from cell to cell, suggesting that GFP does not have a specific motif for plasmodesmal trafficking. Our data, together with previous findings, indicate that plasmodesmata can mediate both specific and nonspecific intercellular trafficking of proteins. Furthermore, our data suggest that nonspecific protein trafficking is controlled by species-, development-, organ-, and tissue-specific factors. Since GFP can readily traffic from cell to cell, it raises the questions of how metabolites are compartmentalized intercellularly in a plant and of whether some endogenous plant proteins traffic nonspecifically from cell to cell to perform physiological functions yet to be elucidated.Abbreviations CMV cucumber mosaic virus - GFP green-fluorescent protein - MP movement protein - SEL size exclusion limit     

Plasmodesma-mediated selective protein traffic between "symplasmically isolated" cells probed by a viral movement protein

Intercellular communication is essential for differentiation and development. In plants, plasmodesmata (PD) form cytoplasmic channels for direct communication. During plant development, programmed reduction in PD number and transport capacity creates the so-called symplasmic domains. Small fluorescent dyes and ions can diffuse among cells within a domain but not across domain boundaries. Such symplasmic isolation is thought to allow groups of cells to differentiate and develop into tissues with distinct structures and functions. Whether or how "symplasmically isolated" cells communicate with one another is poorly understood. One well-documented symplasmic domain is the sieve element-companion cell (SE-CC) complex in the phloem tissue. We report here that, when produced in the CC of transgenic tobacco, the 3a movement protein (3a MP) of Cucumber mosaic virus fused to green fluorescent protein (GFP) can traffic out of the SE-CC complex via PD. The extent of 3a MP:GFP traffic across the boundary between vascular and nonvascular tissues depends on organ type and developmental stage. Our findings provide experimental evidence that endogenous machinery exists for protein traffic between the symplasmically isolated SE-CC complex and neighboring cells. We suggest that PD-mediated traffic of selected macromolecules can be a mechanism for symplasmically isolated cells to communicate with one another.     

The silver lining of a viral agent: increasing seed yield and harvest index in Arabidopsis by ectopic expression of the potato leaf roll virus movement protein

Ectopic expression of viral movement proteins (MPs) has previously been shown to alter plasmodesmata (PD) function and carbon partitioning in transgenic plants, giving rise to the view of PD being dynamic and highly regulated structures that allow resource allocation to be adapted to environmental and developmental needs. However, most work has been restricted to solanaceous species and the potential use of MP expression to improve biomass and yield parameters has not been addressed in detail. Here we demonstrate that MP-mediated modification of PD function can substantially alter assimilate allocation, biomass production, and reproductive growth in Arabidopsis (Arabidopsis thaliana). These effects were achieved by constitutive expression of the potato leaf roll virus 17-kD MP (MP17) fused to green fluorescent protein (GFP) in different Arabidopsis ecotypes. The resulting transgenic plants were analyzed for PD localization of the MP17:GFP fusion protein and different lines with low to high expression levels were selected for further analysis. Low-level accumulation of MP17 resulted in enhanced sucrose efflux from source leaves and a considerably increased vegetative biomass production. In contrast, high MP17 levels impaired sucrose export, resulting in source leaf-specific carbohydrate accumulation and a strongly reduced vegetative growth. Surprisingly, later during development the MP17-mediated inhibition of resource allocation was reversed, and final seed yield increased in average up to 30% in different transgenic lines as compared to wild-type plants. This resulted in a strongly improved harvest index. The release of the assimilate export block was paralleled by a reduced PD binding of MP17 in senescing leaves, indicating major structural changes of PD during leaf senescence.     

Fructose 2,6-bisphosphate levels in mature leaves of tobacco (Nicotiana tabacum) and potato (Solanum tuberosum)

Here we show that fructose 2,6-bisphosphate cannot be reliably measured in mature leaves of tobacco (Nicotiana tabacum L.), potato (Solanum tuberosum L.), or stinging nettle (Urtica dioica L.) using conventional extraction techniques, since the recoveries of fructose 2,6-bisphosphate added during extraction are poor. However, fructose 2,6-bisphosphate could be extracted by boiling leaves in ethanol and aqueous buffer. Evidence for the reliability of this technique is provided by high recovery measurements of fructose 2,6-bisphosphate added to the leaves before extraction. This extraction method was used to measure changes in the level of fructose 2,6-bisphosphate throughout the photoperiod in tobacco and potato leaves. These changes are compared with the rate of accumulation of sucrose and starch in the leaf samples. Variations in the levels of fructose 2,6-bisphosphate, and the relationship between this metabolite and sucrose and starch accumulation in these leaves during the photoperiod are similar to the pattern observed in leaves of other plant species.Abbreviations BSA bovine serum albumin - Fru-2,6-P₂ fructose 2,6-bisphosphate This research was supported by the Agricultural and Food Research Council (Grant no. PG43/531), and the Royal Society.     

Development of a quantitative tool for measuring changes in the coefficient of conductivity of plasmodesmata induced by developmental, biotic, and abiotic signals

Summary. The regulation of intercellular and interorgan communication is pivotal for cell fate decisions in plant development and probably plays a significant role in the systemic regulation of gene expression and in defense reactions against pathogens or other biotic and abiotic environmental factors. In plants, symplasmic cell-to-cell communication is provided by plasmodesmata (Pd), coaxial membranous tunnels that span cell walls interconnecting adjacent cytoplasms. Macromolecules, proteins, and RNA may be transported through Pd by passive diffusion or by a facilitated mechanism. A quantitative tool was developed to measure the coefficient of conductivity, C(Pd), for diffusion-driven transport via Pd and to assess changes in the coefficient induced by developmental, biotic and abiotic signals. ^GFPC(Pd), the coefficient of conductivity for cell-to-cell spread of green-fluorescent protein (GFP), a protein with a Stokes radius of 2.82 nm, was determined in epidermal cells of sink and source leaves of wild-type and transgenic Nicotiana benthamiana plants expressing the movement protein of tobacco mosaic virus (MP^TMV) incubated both in dark and light and at 16 and 25°C. Under all conditions, Pd in source leaves conducted macromolecules, with ^GFPC(Pd)_sink > ^GFPC(Pd)_source. Light down-regulated ^GFPC(Pd) (all conditions); down-regulation was stronger for sink cells. The effect of MP^TMV on ^GFPC(Pd) between epidermal cells was dependent on temperature and leaf development; at 16°C, MP^TMV down-regulated ^GFPC(Pd) only in source leaves, while at 25°C, MP^TMV had no significant effect. This quantitative tool should be useful for investigating differences in Pd conductivity that are induced by mutations or silencing. Correspondence and reprints: Department of Plant Sciences, George S. Wise Faculty of Life Sciences, Tel Aviv University, Tel Aviv 69978, Israel. Present address: Department of Biological Chemistry, Weizmann Institute of Science, Rehovot, Israel.     

The actin cytoskeleton is involved in the regulation of the plasmodesmal size exclusion limit

Plasmodesmata (PD) are the communication channels which allow the trafficking of macromolecules between neighboring cells. Such cell-to-cell movement of macromolecules is regulated during plant growth and development; however, little is known about the regulation mechanism of PD size exclusion limit (SEL). Plant viral movement proteins (MPs) enhance the invasion of viruses from cell to cell by increasing the SEL of the PD and are therefore a powerful means for the study of the plasmodesmal regulation mechanisms. In a recent study, we reported that the actin cytoskeleton is involved in the increase of the PD SEL induced by MPs. Microinjection experiments demonstrated that actin depolymerization was required for the Cucumber mosaic virus (CMV) MP-induced increase in the PD SEL. In vitro experiments showed that CMV MP severs actin filaments (F-actin). Furthermore, through the analyses of two CMV MP mutants, we demonstrated that the F-actin severing ability of CMV MP was required to increase the PD SEL. These results are similar to what has been found in Tobacco mosaic virus MP. Thus, our data suggest that actin dynamics may participate in the regulations of the PD SEL.Key words: plasmodesmata, size exclusion limit, movement protein, actin filaments, F-actin severing     

Sieve elements caught in the act

Phloem is a puzzling plant tissue owing to the unique natural defence responses of the sieve elements to any kind of mechanical manipulation. Recent non-invasive studies have enabled real-time observation of events in intact sieve tubes, including mass transport, sieve-pore sealing and conformational changes of structural proteins. These studies further highlighted the importance of the symplasmic setting for development and functioning of the sieve elements. Exchange of macromolecules between companion cells and sieve elements is indispensable for the survival of the sieve element, but also seems to be involved in long-distance communication. How the branched plasmodesmata between sieve element and companion cell function as corridors for the passage of macromolecules is an intriguing but unresolved story.     

Developmental Regulation of Intercellular Protein Trafficking through Plasmodesmata in Tobacco Leaf Epidermis

Plasmodesmata mediate direct cell-to-cell communication in plants. One of their significant features is that primary plasmodesmata formed at the time of cytokinesis often undergo structural modifications, by the de novo addition of cytoplasmic strands across cell walls, to become complex secondary plasmodesmata during plant development. Whether such modifications allow plasmodesmata to gain special transport functions has been an outstanding issue in plant biology. Here we present data showing that the cucumber mosaic virus 3a movement protein (MP):green fluorescent protein (GFP) fusion was not targeted to primary plasmodesmata in the epidermis of young or mature leaves in transgenic tobacco (Nicotiana tabacum) plants constitutively expressing the 3a:GFP fusion gene. Furthermore, the cucumber mosaic virus 3a MP:GFP fusion protein produced in planta by biolistic bombardment of the 3a:GFP fusion gene did not traffic between cells interconnected by primary plasmodesmata in the epidermis of a young leaf. In contrast, the 3a MP:GFP was targeted to complex secondary plasmodesmata and trafficked from cell to cell when a leaf reached a certain developmental stage. These data provide the first experimental evidence, to our knowledge, that primary and complex secondary plasmodesmata have different protein-trafficking functions and suggest that complex secondary plasmodesmata may be formed to traffic specific macromolecules that are important for certain stages of leaf development.     

ANK, a host cytoplasmic receptor for the Tobacco mosaic virus cell-to-cell movement protein, facilitates intercellular transport through plasmodesmata

Plasmodesma (PD) is a channel structure that spans the cell wall and provides symplastic connection between adjacent cells. Various macromolecules are known to be transported through PD in a highly regulated manner, and plant viruses utilize their movement proteins (MPs) to gate the PD to spread cell-to-cell. The mechanism by which MP modifies PD to enable intercelluar traffic remains obscure, due to the lack of knowledge about the host factors that mediate the process. Here, we describe the functional interaction between Tobacco mosaic virus (TMV) MP and a plant factor, an ankyrin repeat containing protein (ANK), during the viral cell-to-cell movement. We utilized a reverse genetics approach to gain insight into the possible involvement of ANK in viral movement. To this end, ANK overexpressor and suppressor lines were generated, and the movement of MP was tested. MP movement was facilitated in the ANK-overexpressing plants, and reduced in the ANK-suppressing plants, demonstrating that ANK is a host factor that facilitates MP cell-to-cell movement. Also, the TMV local infection was largely delayed in the ANK-suppressing lines, while enhanced in the ANK-overexpressing lines, showing that ANK is crucially involved in the infection process. Importantly, MP interacted with ANK at PD. Finally, simultaneous expression of MP and ANK markedly decreased the PD levels of callose, β-1,3-glucan, which is known to act as a molecular sphincter for PD. Thus, the MP-ANK interaction results in the downregulation of callose and increased cell-to-cell movement of the viral protein. These findings suggest that ANK represents a host cellular receptor exploited by MP to aid viral movement by gating PD through relaxation of their callose sphincters.     

Close association of centrosomes to the distal ends of the microbody during its growth, division and partitioning in the green alga Klebsormidium flaccidum

Summary. Division and partitioning of microbodies (peroxisomes) of the green alga Klebsormidium flaccidum, whose cells contain a single microbody, were investigated by electron microscopy. In interphase, the rod-shaped microbody is present between the nucleus and the single chloroplast, oriented perpendicular to the pole-to-pole direction of the future spindle. A centriole pair associates with one distal end of the microbody. In prophase, the microbody changes not only in shape, from a rodlike to a branched form, but also in orientation, from perpendicular to parallel to the future pole-to-pole direction. Duplicated centriole pairs are localized in close proximity to both distal ends of the microbody. In metaphase, the elongated microbody flanks the open spindle, with both distal ends close to the centriole pair at either spindle pole. The microbody further elongates in telophase and divides after septum formation (cytokinesis) has started. The association between the centrioles and both distal ends of the microbody is maintained throughout mitosis, resulting in the distal ends of the elongated microbody being fixed at the cellular poles. This configuration of the microbody may be favorable for faithful transmission of the organelle during cell division. After cytokinesis is completed, the microbody reverts to the perpendicular orientation by changing its shape. Microtubules radiating from the centrosomes flank the side of the microbody throughout mitosis. The close association of centrosomes and microtubules with the microbody is discussed in respect to the partitioning of the microbody in this alga. Correspondence: H. Hashimoto, Department of Life Sciences, Graduate School of Arts and Sciences, University of Tokyo, Komaba, Meguro-ku, Tokyo 153-8902, Japan. Present address: M. Honda, Department of Computational Biology, Graduate School of Frontier Sciences, University of Tokyo, Kashiwa, Chiba, Japan.     

Detection of Tobacco Mosaic Virus Movement Protein in Association with Tobacco Nuclei Isolated from Intact and Detached Leaves

The movement protein (MP) of tobacco mosaic virus was observed to cofractionate with nuclei from intact and detached tobacco (Nicotiana tahacum cv. Xanthi-nn) leaves. When purified nuclei were treated with proteinase K, MP disappeared indicating that MP is localized close to but not inside nuclei. Moreover, the amount of MP was showti to increase greatly in nuclei isolated from detached leaves, thus facilitating the detection of MP. Accumulation close to nuclei was strongest early in infection, results indicating that it may be an early step in the pathway of MP from cell cytoplasm to plasmodesmata. Other TMV-specific proteins were not detected in the nuclei fraction.     

Choline transporter‐like1 (CHER1) is crucial for plasmodesmata maturation in Arabidopsis thaliana

Plasmodesmata (PD) are microscopic pores connecting plant cells and enable cell‐to‐cell transport. Currently, little information is known about the molecular mechanisms regulating PD formation and development. To uncover components of PD development we made use of the 17 kDa movement protein (MP17) encoded by the Potato leafroll virus (PLRV). The protein is required for cell‐to‐cell movement of the virus and localises to complex PD. Forward genetic screening for Arabidopsis mutants with altered PD binding of MP17 revealed several mutant lines, while molecular genetics, biochemical and microscopic studies allowed further characterisation. Map‐based cloning of one mutant revealed a point mutation in the choline transporter‐like 1 (CHER1) protein, changing glycine²⁴⁷ into glutamate. Mutation in CHER1 resulted in a starch excess phenotype and stunted growth. Ultrastructure analysis of shoot apical meristems, developing and fully developed leaves showed reduced PD numbers and the absence of complex PD in fully developed leaves. This indicates that cher1 mutants are impaired in PD formation and development. Global lipid profiling revealed only slight modifications in the overall lipid composition, however, altered composition of PD‐associated lipids cannot be ruled out. Thus, cher1 is devoid of complex PD in developed leaves and provides insights into the formation of complex PD at the molecular level.     

Changes in the protein composition of cytoplasmic ribsomes during the greening of etiolated barley leaves

We examined changes in the protein composition of cytoplasmic ribosomes in etiolated barley leaves following illumination. Cytoplasmic ribosomes were isolated from greening barley leaves by sucrose density gradient centrifugation, and were analyzed by radical-free highly reducing polyacrylamide gel electrophoresis (RFHR-PAGE). Eighty-nine proteins were resolved from the ribosomal fraction; among them, 8 proteins changed their copy numbers depending on the stage of greening. We designated these as phase dependent ribosomal proteins (PD1–PD8). Two of the proteins (PD1 and 5) present in the ribosomes of etiolated leaves showed a decrease in level during greening. In contrast, the levels of 6 ribosomal proteins (PD2, 3, 4, 6, 7 and 8) increased as greening proceeded. N-terminal amino acid sequence of PD8 showed high homology to rat ribosomal protein L34. The ribosomal proteins that appeared after illumination were not found in any fraction of the etiolated leaves, suggesting that they were synthesized after the onset of illumination. Copy numbers of other ribosomal proteins did not change during greening.     

烟草GA合成调控转录因子RSG应答甲醇和乙醇刺激的分子机理初探

为了考察甲醇或乙醇促进植物生长与赤霉素(GA)的合成关系,该研究在MS固体培养基中培养并添加外源GA和GA合成抑制剂多效唑(PAC),分析其对2mmol/L甲醇或乙醇促进烟草生长的影响及GA合成调控转录因子RSG(for repression of shoot growth)应答甲醇或乙醇刺激的分子机理。结果显示:(1)外源添加GA可增强甲醇或乙醇对烟草生长的刺激作用,而添加PAC却抑制甲醇和乙醇对烟草生长的刺激作用。(2)14-3-3蛋白与RSG结合抑制RSG进入细胞核及其转录调控活性;甲醇和乙醇诱导烟草14-3-3基因的转录和表达,对RSG蛋白表达也有诱导作用。(3)甲醇和乙醇可降低14-3-3蛋白与RSG的相互作用,同时增强RSG与GA20ox1启动子的结合。研究表明,甲醇和乙醇刺激烟草的生长可能通过增加RSG表达,且减弱RSG与14-3-3蛋白的结合来增加RSG细胞核定位作用,从而增强RSG与GA20ox1启动子的结合,最终增加GA的合成,从而促进烟草的生长,这可能是甲醇和乙醇促进烟草生长的一种重要的分子机制。     

Expression of a luteoviral movement protein in transgenic plants leads to carbohydrate accumulation and reduced photosynthetic capacity in source leaves

Elucidating the role of viral genes in transgenic plants revealed that the movement protein (MP) from tobacco mosaic virus is responsible for altered carbohydrate allocation in tobacco and potato plants. To study whether this is a general feature of viral MPs, the movement protein MP17 of potato leafroll virus (PLRV), a phloem-restricted luteovirus, was constitutively expressed in tobacco plants. Transgenic lines were strongly reduced in height and developed bleached and sometimes even necrotic areas on their source leaves. Levels of soluble sugars and starch were significantly increased in source leaves. Yet, in leaf laminae the hexose—phosphate content was unaltered and ATP reduced to only a small extent, indicating that these leaves were able to maintain homeostatic conditions by compartmentalization of soluble sugars, probably in the vacuole. On the contrary, midribs contained lower levels of soluble sugars, ATP, hexose—phosphates and UDP-glucose supporting the concept of limited uptake and catabolism of sucrose in the phloem. The accumulation of carbohydrates led to a decreased photosynthetic capacity and carboxylation efficiency of ribulose-1,5-bisphosphate carboxylase/oxygenase (rubisco) probably owing to decreased expression of photosynthetic proteins. In parallel, levels of pathogenesis-related proteins were elevated which may be the reason for the obtained limited resistance against the unrelated potato virus Y (PVY)^N in the transgenic tobacco plants. Ultrathin sections of affected leaves harvested from 2-week-old plants revealed plasmodesmal alterations in the phloem tissue while plasmodesmata between mesophyll cells were indistinguishable from wild-type. These data favour the phloem tissue to be the primary site of PLRV MP17 action in altering carbohydrate metabolism.     

Cucumber mosaic virus movement protein affects sugar metabolism and transport in tobacco and melon plants

In addition to its influence on plasmodesmal function, tobacco mosaic virus movement protein (TMV‐MP) causes an alteration in carbon metabolism in source leaves and in resource partitioning among the various plant organs. The present study was aimed at characterizing the influence of cucumber mosaic virus (CMV)‐MP on carbohydrate metabolism and transport in both tobacco and melon plants. Transgenic tobacco plants expressing the CMV‐MP had reduced levels of soluble sugars and starch in their source leaves and a significantly reduced root‐to‐shoot ratio in comparison with control plants. A novel virus‐vector system was employed to express the CMV‐coat protein (CP), the CMV‐MP or the TMV‐MP in melon plants. This set of experiments indicated that the viral MPs cause a significant elevation in the proportion of sucrose in the phloem sap collected from petioles of source leaves, whereas this sugar was at very low levels or even absent from the sap of control melon plants. The mode by which the CMV‐MP exerts its effect on phloem‐sap sugar composition is discussed in terms of possible alterations in the mechanism of phloem loading.     

Non-targeted and targeted protein movement through plasmodesmata in leaves in different developmental and physiological states

Plant cells rely on plasmodesmata for intercellular transport of small signaling molecules as well as larger informational macromolecules such as proteins. A green fluorescent protein (GFP) reporter and low-pressure microprojectile bombardment were used to quantify the degree of symplastic continuity between cells of the leaf at different developmental stages and under different growth conditions. Plasmodesmata were observed to be closed to the transport of GFP or dilated to allow the traffic of GFP. In sink leaves, between 34% and 67% of the cells transport GFP (27 kD), and between 30% and 46% of the cells transport double GFP (54 kD). In leaves in transition transport was reduced; between 21% and 46% and between 2% and 9% of cells transport single and double GFP, respectively. Thus, leaf age dramatically affects the ability of cells to exchange proteins nonselectively. Further, the number of cells allowing GFP or double GFP movement was sensitive to growth conditions because greenhouse-grown plants exhibited higher diffusion rates than culture-grown plants. These studies reveal that leaf cell plasmodesmata are dynamic and do not have a set size exclusion limit. We also examined targeted movement of the movement protein of tobacco mosaic virus fused to GFP, P30::GFP. This 58-kD fusion protein localizes to plasmodesmata, consistently transits from up to 78% of transfected cells, and was not sensitive to developmental age or growth conditions. The relative number of cells containing dilated plasmodesmata varies between different species of tobacco, with Nicotiana clevelandii exhibiting greater diffusion of proteins than Nicotiana tabacum.     

Plasmodesmal-associated protein kinase in tobacco and Arabidopsis recognizes a subset of non-cell-autonomous proteins

Cell-to-cell communication in plants involves the trafficking of macromolecules through specialized intercellular organelles, termed plasmodesmata. This exchange of proteins and RNA is likely regulated, and a role for protein phosphorylation has been implicated, but specific components remain to be identified. Here, we describe the molecular characterization of a plasmodesmal-associated protein kinase (PAPK). A 34-kD protein, isolated from a plasmodesmal preparation, exhibits calcium-independent kinase activity and displays substrate specificity in that it recognizes a subset of viral and endogenous non-cell-autonomous proteins. This PAPK specifically phosphorylates the C-terminal residues of tobacco mosaic virus movement protein (TMV MP); this posttranslational modification has been shown to affect MP function. Molecular analysis of purified protein established that tobacco (Nicotiana tabacum) PAPK is a member of the casein kinase I family. Subcellular localization studies identified a possible Arabidopsis thaliana PAPK homolog, PAPK1. TMV MP and PAPK1 are colocalized within cross-walls in a pattern consistent with targeting to plasmodesmata. Moreover, Arabidopsis PAPK1 also phosphorylates TMV MP in vitro at its C terminus. These results strongly suggest that Arabidopsis PAPK1 is a close homolog of tobacco PAPK. Thus, PAPK1 represents a novel plant protein kinase that is targeted to plasmodesmata and may play a regulatory role in macromolecular trafficking between plant cells.