Plasmodesmata signaling: many roles, sophisticated statutes.

Recent advances have increased our understanding of plasmodesmata function, their architecture as it relates to signaling capacity, the temporal and spatial regulation of their permeability, and their roles in systemic transport of macromolecules, non-cell autonomous development, and, potentially, plant defense.     

Plasmodesmata and pit development in secondary xylem elements

Developing pit membranes of secondary xylem elements in Drimys winteri, Fagus sylvatica, Quercus robur, Sorbus aucuparia, Tilia vulgaris and Trochodendron aralioides have been examined by transmission electron microscopy. Absence of plasmodesmata from the membranes of vessel elements and tracheids indicates that their pits develop independently of these structures. On the other hand, plasmodesmata are abundant in pit membranes between fibres, parenchyma cells, and combinations of these cell types in Fagus, Quercus and Tilia. In each case the plasmodesmata pass right through the developing pit membrane. In the case of Sorbus fibres, however, plasmodesmata were absent from the majority of pit membrane profiles seen in sections. Occasionally they were observed in large numbers associated with a swollen region on one side of the pit membrane between fibres and between fibres and parenchyma, radiating from a small area of the middle lamella. In the case of fibre to parenchyma pitting, this swelling was always found on the fibre side of the membrane, while on the other side a small number of plasmodesmata were present completing communication with the parenchyma cytoplasm. These observations are discussed with regard to the role of plasmodesmata in pit formation, and in the differentiation of the various cell types in secondary xylem. The significance their distribution may have for our understanding of xylem evolution is also discussed.     

Developmental changes in plasmodesmata in transgenic tobacco expressing the movement protein of tobacco mosaic virus

Summary Cell-to-cell communication in plants occurs through plasmodesmata, cytoplasmic channels that traverse the cell wall between neighboring cells. Plasmodesmata are also exploited by many viruses as an avenue for spread of viral progeny. In the case of tobacco mosaic virus (TMV), a virally-encoded movement protein (MP) enables the virus to move through plasmodesmata during infection. We have used thin section electron microscopy and immunocytochemistry to examine the structure of plasmodesmata in transgenic tobacco plants expressing the TMV MP. We observed a change in structure of the plasmodesmata as the leaves age, both in control and MP expressing [MP(+)] plants. In addition, the plasmodesmata of older cells of MP(+) plants accumulate a fibrous material in the central cavity. The presence of the fibers is correlated with the ability to label plasmodesmata with anti-MP antibodies. The developmental stage of leaf tissue at which this material is observed is the stage at which an increase in the size exclusion limit of the plasmodesmata can be measured in MP(+) plants. Using cell fractionation and aqueous phase partitioning studies, we identified the plasma membrane and cell wall as the compartments with which the MP stably associates. The nature of the interaction between the MP and the plasma membrane was studied using sodium carbonate and Triton X-100 washes. The MP behaves as an integral membrane protein. Identifying the mechanism by which the MP associates with plasma membrane and plasmodesmata will lead to a better understanding of how the MP alters the function of the plasmodesmata.Abbreviations MP movement protein - TMV tobacco mosaic virus     

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.     

Plasmodesmata 2010: plasmodesmata down under

More than 60 attendees from more than a dozen countries attended the International Plasmodesmata Meeting (Plasmodesmata 2010) held in Sydney, Australia. The structure of plasmodesmata continued to attract interest, with particular focus on how technological progress is advancing our ability to identify and characterise proteins associated with plasmodesmata. Also of major research interest was the movement of proteins and RNAs through plasmodesmata and how this is controlled by host chaperones, cytoskeletal elements and callose. There was also much new information on viral movement through plasmodesmata, with a focus on the ways that viral movement proteins interact with host cell components to modify plasmodesmata. The conference, as a whole, provided a stimulating forum for the discussion of future directions in this expanding field.     

Plasmodesmata form and function

Intercellular transport via plasmodesmata controls cell fate decisions in plants, and is of fundamental importance in viral movement, disease resistance, and the spread of RNAi signals. Although plasmodesmata appear to be unique to plant cells, they may have structural and functional similarities to the newly discovered tunneling nanotubes that connect animal cells. Recently, proteins that localize to plasmodesmata have been identified, and a microtubule-associated protein was found to negatively regulate the trafficking of viral movement proteins. Other advances have delivered new insights into the function and molecular nature of plasmodesmata and have shown that protein trafficking through plasmodesmata is developmentally regulated, opening up the possibility that the genetic control of plasmodesmal function will soon be understood.     

Plasmodesmata as a Modulator of Osmotic Water Fluxes in Plants

Solutions to some key problems in the relationships between the structure and functions of plasmodesmata, a component of the plant intercellular communication system, are proposed on the basis of the theory of osmotic flows through porous membranes. The theory accounts for structural characteristics of plasmodesmata, such as their dimension, shape, and length. It considers the steric and adsorption potentials of the solution–cell wall interaction and estimates water and solute (e.g., sucrose) flows under the sustained difference of osmotic pressures at the ends of plasmodesmata. The theory predicts that the water flow through plasmodesmata increases with the widening of the neck constriction and reaches its peak when its size is equal to the diameter of the solute molecule. The water-flow direction was found to depend on the opening of the annulus in neck constrictions at negative adsorption potentials of the plasmodesmata channel walls. Taking into account the presence of sphincters in the neck constrictions, our data suggest the role of plasmodesmata as a modulator of osmotic water fluxes in plants.     

Comparative ultrastructure of plasmodesmata of Chara and selected bryophytes: toward an elucidation of the evolutionary origin of plant plasmodesmata

We have used transmission electron microscopy to examine plasmodesmata of the charophycean green alga Chara zeylanica, and of the putatively early divergent bryophytes Monoclea gottschei (liverwort), Notothylas orbicularis (hornwort), and Sphagnum fimbriatum (moss), in an attempt to learn when seed plant plasmodesmata may have originated. The three bryophytes examined have desmotubules. In addition, Monoclea was found to have branched plasmodesmata, and plasmodesmata of Sphagnum displayed densely staining regions around the neck region, as well as ring-like wall specializations. In Chara, longitudinal sections revealed endoplasmic reticulum (ER) that sometimes appeared to be associated with plasmodesmata, but this was rare, despite abundant ER at the cell periphery. Across all three fixation methods, cross-sectional views showed an internal central structure, which in some cases appeared to be connected to the plasma membrane via spoke-like structures. Plasmodesmata were present even in the incompletely formed reticulum of forming cell plates, from which we conclude that primary plasmodesmata are formed at cytokinesis in Chara zeylanica. Based on these results it appears that plasmodesmata of Chara may be less specialized than those of seed plants, and that complex plasmodesmata probably evolved in the ancestor of land plants before extant lineages of bryophytes diverged.     

The distribution of plasmodesmata and its relationship to morphogenesis in fern gametophytes

Fern (Onoclea sensibilis) gametophytes when grown in the dark form a linear file of cells (one-dimensional) called a protonema. In the light two-dimensional growth occurs which results in a heart-shaped prothallus one cell thick. The objective of this paper is to relate the most common pattern of cell division observed in developing gametophytes to the formation of the plasmodesmatal network. Since the prothalli are only two dimensional, we can easily determine from thin sections the total number and the density (number per unit surface area) of plasmodesmata at each developmental stage. As the prothallus grows the number of plasmodesmata increases 50-fold in the apical or meristematic cell. This number eventually reaches a plateau even though the density continues to increase with each new cell division. What is particularly striking is that both the number and density of plasmodesmata between adjacent cells is precisely determined. Furthermore, the pattern of plasmodesmata distribution is predictable so that (1) we can identify the apical meristematic cells by their plasmodesmata number, or density, as well as by their size, shape and location, (2) we can predict, again from plasmodesmata number, the location of a future wall of the apical cell prior to its actual formation, (3) we can show that the density of plasmodesmata in the triangular apical cell of the prothallus (14 plasmodesmata microns-2) is comparable to those reported for secretory glands which are known to have high rates of plasmodesmatal transport and (4) we can show that once the plasmodesmata have been formed during division, no subsequent change in the number of plasmodesmata occurs following cell plate formation.     

Specific targeting of a plasmodesmal protein affecting cell-to-cell communication

Plasmodesmata provide the cytoplasmic conduits for cell-to-cell communication throughout plant tissues and participate in a diverse set of non–cell-autonomous functions. Despite their central role in growth and development and defence, resolving their modus operandi remains a major challenge in plant biology. Features of protein sequences and/or structure that determine protein targeting to plasmodesmata were previously unknown. We identify here a novel family of plasmodesmata-located proteins (called PDLP1) whose members have the features of type I membrane receptor-like proteins. We focus our studies on the first identified type member (namely At5g43980, or PDLP1a) and show that, following its altered expression, it is effective in modulating cell-to-cell trafficking. PDLP1a is targeted to plasmodesmata via the secretory pathway in a Brefeldin A–sensitive and COPII-dependent manner, and resides at plasmodesmata with its C-terminus in the cytoplasmic domain and its N-terminus in the apoplast. Using a deletion analysis, we show that the single transmembrane domain (TMD) of PDLP1a contains all the information necessary for intracellular targeting of this type I membrane protein to plasmodesmata, such that the TMD can be used to target heterologous proteins to this location. These studies identify a new family of plasmodesmal proteins that affect cell-to-cell communication. They exhibit a mode of intracellular trafficking and targeting novel for plant biology and provide technological opportunities for targeting different proteins to plasmodesmata to aid in plasmodesmal characterisation.     

Maintaining apical dominance in the fern gametophyte

A kinetic model is developed for cell differentiation in the fern gametophyte to test hypotheses on the role of spatially patterned plasmodesmata networks in development. Of particular interest is the establishment and maintenance of apical cell type in a single cell, with concurrent suppression of this character in all other cells (apical dominance). Steps towards understanding apical cell localization in geometrically simple gametophytes may shed light on the establishment and maintenance of apical meristems in higher plants. The model, based on the plasmodesmata maps of Tilney and colleagues and involving kinetics for a requisite minimum of two morphogens. successfully produces the apical/non-apical cell differentiation patterns of normal development, and redifferentiation due to cell isolation, in six stages from 0-30 d of development. Our results indicate that increasing apical cell plasmodesmata number, as development progresses, is not required for effective transport across apical cell walls in maintaining apical dominance.     

The distribution of plasmodesmata in the root tip of maize

Summary The distribution of plasmodesmata in different regions of the root apex of Zea mays has been analysed from electron micrographs. There are many more plasmodesmata traversing transverse walls than across longitudinal walls in all the regions studied. When the number of plasmodesmata per unit cell volume is calculated, cells in non-dividing tissue have a considerably lower value than cells in dividing tissue. Evidence for the transport of materials between cells via plasmodesmata is summarised. If it is accepted that plasmodesmata do act as channels for intercellular communication then we believe that their pattern of distribution may be a contributory factor to the process of cell differentiation.     

Are unusual plasmodesmata in the embryo-suspensor restricted to species from the genus Sedum among Crassulaceae?

The Crassulaceae family comprises mainly herbaceous leaf succulents, some of which have an ornamental value. During embryogenesis, they produce a suspensor with a giant polyploid basal cell. It has recently been shown that in Sedum acre and S. hispanicum this cell has compound plasmodesmata with an unusual dome of electron-dense material associated on the cell's side. These compound plasmodesmata differ from the typical ones occurring in other angiosperms. In this study, the hypothesis was tested that the unusual plasmodesmata in the embryo-suspensor are a feature not only restricted to species from the genus Sedum, but are also found in other Crassulaceae genera. Suspensors of example species from the genera Sempervivum and Jovibarba, which have vegetative morphologies quite different from Sedum and which are placed in the traditional classification into another subfamily, were first examined using an electron microscope. It was found that the unusual compound plasmodesmata in the suspensor are not only restricted to species from the genus Sedum but are also found in species from other Crassulaceae genera (Sempervivum arachnoideum and Jovibarba sobolifera). It should be noted that some ultrastructural features of compound plasmodesmata in the analyzed genera (e.g. the character of the wall with plasmodesmata, plasmodesmata diameter or occurrence of the electron-dense material) are different from the suspensor plasmodesmata recorded in species from the Sedum genus. We found that in Sempervivum arachnoideum the size of the plasmodesmata diameter varies according to the micropylar-chalazal axis of the embryo. This is the first report of variation in the diameter of the plasmodesmata within the embryo of angiosperms. Further study will be needed to show whether compound plasmodesmata occur in other Crassulaceae clades, whether they are a stable feature at the genus level in this family, and also whether they have evolved several times or only once in Crassulaceae.     

Phloem loading. A reevaluation of the relationship between plasmodesmatal frequencies and loading strategies

The incidence of plasmodesmata in the minor vein phloem of leaves varies widely between species. On this basis, two pathways of phloem loading have been proposed: symplastic where frequencies are high, and apoplastic where they are low. However, putative symplastic-loading species fall into at least two categories. In one, the plants translocate raffinose-family oligosaccharides (RFOs). In the other, the primary sugar in the phloem sap is sucrose (Suc). While a thermodynamically feasible mechanism of symplastic loading has been postulated for species that transport RFOs, no such mechanism is known for Suc transporters. We used p-chloromercuribenzenesulfonic acid inhibition of apoplastic loading to distinguish between the two pathways in three species that have abundant minor vein plasmodesmata and are therefore putative symplastic loaders. Clethra barbinervis and Liquidambar styraciflua transport Suc, while Catalpa speciosa transports RFOs. The results indicate that, contrary to the hypothesis that all species with abundant minor vein plasmodesmata load symplastically, C. barbinervis and L. styraciflua load from the apoplast. C. speciosa, being an RFO transporter, loads from the symplast, as expected. Data from these three species, and from the literature, also indicate that plants with abundant plasmodesmata in the minor vein phloem have abundant plasmodesmata between mesophyll cells. Thus, plasmodesmatal frequencies in the minor veins may be a reflection of overall frequencies in the lamina and may have limited relevance to phloem loading. We suggest that symplastic loading is restricted to plants that translocate oligosaccharides larger than Suc, such as RFOs, and that other plants, no matter how many plasmodesmata they have in the minor vein phloem, load via the apoplast.     

Some speculations on the possible roles of the plasmodesmata in the control of differentiation

Most plasmodesmata are formed across the cell plate at cytokinesis. Most of them persist until the cell is mature. Depending upon the pattern of elongation of the cell in differentiation, the frequency of plasmodesmata per unit area will suffer dilution to a greater or lesser extent. This dilution effect is now well understood and results commonly in high concentrations of plasmodesmata across transverse walls which have undergone little elongation and low concentrations on the longitudinal walls.Apart from their obvious role in cell to cell communication it is now believed that some plasmodesmata may offer preferential sites from which endogenous wall lytic enzymes may attack some or all of the constituent polymers of the surrounding wall. The effects of the asymmetrical distribution of large numbers of plasmodesmata, leading to the asymmetrical penetration of the wall by lytic enzymes are described and a hypothesis concerning the later stages of cell differentiation is constructed. In addition the late stage differentiation of individual plasmodesmata based on the same proposed lytic action, is described and re-interpreted.     

Current perspectives on plasmodesmata: structure and function

Recent studies on plasmodesmata have shown that these important intercellular passages for communication and transport are much more sophisticated in both structure and regulatory abilities than previously imagined. A complex, but not well understood, substructure has been revealed by a variety of increasingly reliable ultrastructural techniques. Proteinaceous particles are seen within the cytoplasmic sleeve surrounding the desmotubule. Dye-coupling studies have provided experimental evidence for the physical pathway of solute movement, supporting conclusions about substructural dimensions within plasmodesmata drawn from the ultrastructural studies. Calcium has been identified as a major factor in the regulation of intercellular communication via plasmodesmata. Evidence from studies on virus movement through plasmodesmata suggests a direct interaction between virallycoded movement proteins and plasmodesmata in the systemic spread of many viruses. There is increasing evidence, albeit indirect, that in some plant species phloem loading may involve transport of photoassimilate entirely within the symplast from mesophyll cells to the sieve element-companion cell complexes of minor veins.     

Integument cell gelatinisation—the fate of the integumentary cells in <Emphasis Type="Italic">Hieracium</Emphasis> and <Emphasis Type="Italic">Pilosella</Emphasis> (Asteraceae)

Members of the genera Hieracium and Pilosella are model plants that are used to study the mechanisms of apomixis. In order to have a proper understanding of apomixis, knowledge about the relationship between the maternal tissue and the gametophyte is needed. In the genus Pilosella, previous authors have described the specific process of the “liquefaction” of the integument cells that surround the embryo sac. However, these observations were based on data only at the light microscopy level. The main aim of our paper was to investigate the changes in the integument cells at the ultrastructural level in Pilosella officinarum and Hieracium alpinum. We found that the integument peri-endothelial zone in both species consisted of mucilage cells. The mucilage was deposited as a thick layer between the plasma membrane and the cell wall. The mucilage pushed the protoplast to the centre of the cell, and cytoplasmic bridges connected the protoplast to the plasmodesmata through the mucilage layers. Moreover, an elongation of the plasmodesmata was observed in the mucilage cells. The protoplasts had an irregular shape and were finally degenerated. After the cell wall breakdown of the mucilage cells, lysigenous cavities that were filled with mucilage were formed.     

Tobacco mosaic virus movement protein associates with the cytoskeleton in tobacco cells.

Tobacco mosaic virus movement protein P30 complexes with genomic viral RNA for transport through plasmodesmata, the plant intercellular connections. Although most research with P30 focuses on its targeting to and gating of plasmodesmata, the mechanisms of P30 intracellular movement to plasmodesmata have not been defined. To examine P30 intracellular localization, we used tobacco protoplasts, which lack plasmodesmata, for transfection with plasmids carrying P30 coding sequences under a constitutive promoter and for infection with tobacco mosaic virus particles. In both systems, P30 appears as filaments that colocalize primarily with microtubules. To a lesser extent, P30 filaments colocalize with actin filaments, and in vitro experiments suggested that P30 can bind directly to actin and tubulin. This association of P30 with cytoskeletal elements may play a critical role in intracellular transport of the P30-viral RNA complex through the cytoplasm to and possibly through plasmodesmata.     

Plasmodesmata: composition,structure and trafficking

Plasmodesmata are highly specialized gatable trans-wall channels that interconnect contiguous cells and function in direct cytoplasm-to-cytoplasm intercellular transport. Computer-enhanced digital imaging analysis of electron micrographs of plasmodesmata has provided new information on plasmodesmatal fine structure. It is now becoming clear that plasmodesmata are dynamic quasi-organelles whose conductivity can be regulated by environmental and developmental signals. New findings suggest that signalling mechanisms exist which allow the plasmodesmatal pore to dilate to allow macromolecular transport. Plant viruses spread from cell to cell via plasmodesmata. Two distinct movement mechanisms have been elucidated. One movement mechanism involves the movement of the complete virus particle along virus-induced tubular structures within a modified plasmodesma. Apparently two virus-coded movement proteins are involved. A second movement mechanism involves the movement of a non-virion form through existing plasmodesmata. In this mechanism, the viral movement protein causes a rapid dilation of existing plasmodesmata to facilitate protein and nucleic acid movement. Techniques for the isolation of plasmodesmata have been developed and information on plasmodesma-associated proteins is now becoming available. New evidence is reviewed which suggests that plasmodesmatal composition and regulation may differ in different cells and tissues.