We turn now to the assembly of plant cells into tissues. The overall structural organization of plants is generally simpler than that of animals. For instance, plants have only four broad types of cells, which in mature plants form four basic classes of tissue: dermal tissue interacts with the environment, vascular tissue transports water and dissolved substances such as sugars and ions, space-filling ground tissue constitutes the major sites of metabolism, and sporogenous tissue forms the reproductive organs. Plant tissues are organized into just four main organ systems: stems have support and transport functions, roots provide anchorage and absorb and store nutrients, leaves are the sites of photosynthesis, and flowers enclose the reproductive structures. Thus, at the cell, tissue, and organ levels, plants are generally less complex than most animals.
Moreover, unlike animals, plants do not replace or repair old or damaged cells or tissues; they simply grow new organs. Indeed, the developmental fate of any given plant cell is primarily based on its position in the organism rather than on its lineage, whereas both are important in animals. In both plants and animals, a cell’s direct communication with its neighbors is important. Most important for this chapter, and in contrast with animals, few cells in plants contact one another directly through molecules incorporated into their plasma membranes. Instead, plant cells are typically surrounded by a rigid cell wall that contacts the cell walls of adjacent cells (Figure 1a). Also in contrast with animal cells, a plant cell rarely changes its position in the organism relative to other cells. These features of plants and their organization have determined the distinctive molecular mechanisms by which plant cells are incorporated into tis sues and communicate with one another.
Fig1. Structure of the plant cell wall. (a) Overview of the organization of a typical plant cell, in which the organelle-filled cell with its plasma membrane is surrounded by a well-defined extra cellular matrix called the cell wall. (b) Schematic representation of the cell wall of an onion. Cellulose and hemicellulose are arranged into at least three layers in a matrix of pectin. The sizes of the polymers and their separations are drawn to scale. To simplify the diagram, most of the hemicellulose cross-links and other matrix constituents (e.g., extensin, lignin) are not shown. See M. McCann and K. R. Roberts, 1991, in C. Lloyd, ed., The Cytoskeletal Basis of Plant Growth and Form, Academic Press, p. 126. (c) Quick-freeze deep-etch electron micrograph of the cell wall of a garden pea in which some of the pectin molecules were removed by chemical treatment. The abundant thicker fibers are cellulose microfibrils, and the thinner fibers are hemicellulose cross-links (red arrowheads). [Part (b) courtesy Maureen C. McCann. Part (c) republished with permission of Oxford University Press, from Fujino, T., et al., “Characterization of cross-links between cellulose microfibrils, and their occurrence during elongation growth in pea epicotyl,” Plant Cell Physiol. 2000, 41(4):486–94; permission conveyed through the Copyright Clearance Center, Inc.]
The Plant Cell Wall Is a Laminate of Cellulose Fibrils in a Matrix of Glycoproteins
The plant cell wall, an extracellular matrix that is mainly composed of polysaccharides and is about 0.2 μm thick, completely coats the outside of the plant cell’s plasma mem brane. This structure serves some of the same functions as the ECM produced by animal cells, even though the two structures are composed of entirely different macromolecules and have a different organization. About 1000 genes in the plant Arabidopsis, a small flowering plant also called “thale cress”, are devoted to the synthesis and functioning of its cell wall, including approximately 414 glycosyltransferase genes and more than 316 glycosyl hydrolase genes. Similar to animal ECMs, the plant cell wall organizes cells into tissues, signals a plant cell to grow and divide, and controls the shapes of plant organs. It is a dynamic structure that plays important roles in controlling the differentiation of plant cells during embryogenesis and growth, and it provides a barrier to protect against pathogen infection. Just as the ECM helps define the shapes of animal cells, the cell wall defines the shapes of plant cells. When the cell wall is digested away from plant cells by hydrolytic enzymes, spherical cells enclosed by a plasma membrane are left.
Because a major function of the plant cell wall is to with stand the turgor pressure of the cell (between 14.5 and 435 pounds per square inch; see Chapter 11), the cell wall is built for lateral strength. It is arranged into layers of cellulose mi crofibrils: bundles of 30–36 parallel chains of extensively hydrogen-bonded, long (as much as 7 μm or greater), linear polymers of glucose in β glycosidic linkages. The cellulose microfibrils are embedded in a matrix composed of pectin, a negatively charged polymer of d-galacturonic acid and other monosaccharides, and hemicellulose, a short, highly branched polymer of several five- and six-carbon monosaccharides. The mechanical strength of the cell wall depends on cross-linking of the microfibrils by hemicellulose chains (Figure 1b, c). The layers of microfibrils prevent the cell wall from stretching laterally. Cellulose microfibrils are synthesized on the exoplasmic face of the plasma membrane from UDP-glucose and ADP-glucose formed in the cytosol. The polymerizing enzyme, called cellulose synthase, moves within the plane of the plasma membrane along tracks of intracellular microtubules as cellulose is formed, providing a distinctive mechanism for intracellular-extracellular communication and ensuring that the cellulose microfibrils are oriented properly to permit cell-wall, and thus whole-cell, growth.
Unlike cellulose, pectin and hemicellulose are synthesized in the Golgi complex and transported to the cell sur face, where they form an interlinked network that helps bind the walls of adjacent cells to one another and cushions them. When purified, pectin binds water and forms a gel in the presence of Ca2+ and borate ions—hence the use of pectins in many processed foods. As much as 15 percent of the cell wall may be composed of extensin, a glycoprotein that contains abundant hydroxyproline and serine. Most of the hydroxyproline residues are linked to short chains of arabinose (a five-carbon monosaccharide), and the serine residues are linked to galactose. Carbohydrate accounts for about 65 percent of extensin by weight, and its protein backbone forms an extended rodlike helix with the hydroxyl or O-linked carbohydrates protruding outward. Lignin—a complex, insoluble polymer of phenolic residues—associates with cellulose and is a strengthening material. Like cartilage proteoglycans, lignin resists compression forces.
The cell wall is a selective filter whose permeability is controlled largely by pectins. Whereas water and ions diffuse freely across cell walls, the diffusion of large molecules, including proteins larger than 20 kDa, is limited. This limitation may explain why many plant hormones are small, water-soluble molecules, which can diffuse across the cell wall and interact with receptors in the plasma membrane of plant cells.
Loosening of the Cell Wall Permits Plant Cell Growth
Because the cell wall surrounding a plant cell prevents it from expanding, the wall’s structure must be loosened when the cell grows. The amount, type, and direction of plant-cell growth are regulated by small-molecule hormones called auxins. The auxin-induced weakening of the cell wall permits the expansion of the intracellular vacuole (see Figure 1a) by uptake of water, leading to elongation of the cell. We can grasp the magnitude of this phenomenon by considering that, if all cells in a redwood tree were reduced to the size of a typical liver cell, the tree would have a maximum height of only 1 meter, about a hundredfold less than normal.
The cell wall undergoes its greatest changes at the meri stem in a root or shoot tip. Meristems are where cells divide and grow. Young meristematic cells are connected by thin primary cell walls, which can be loosened and stretched to allow subsequent cell elongation. After cell elongation ceases, the cell wall is generally thickened, either by the secretion of additional macromolecules into the primary wall or, more usually, by the formation of a secondary cell wall composed of several layers. In mature tissues such as the xylem—the tubes that conduct salts and water from the roots through the stems to the leaves—most of the cell eventually degenerates, leaving only the cell wall. The unique properties of wood and of plant fibers such as cotton are due to the molecular properties of the cell walls in the tissues of origin.
Plasmodesmata Directly Connect the Cytosols of Adjacent Cells The presence of a cell wall separating cells in plants imposes barriers to cell-cell communication—and thus cell differentiation—not faced by animals. One distinctive mechanism used by plant cells to communicate directly is specialized cell junctions called plasmodesmata, which extend through the cell wall (Figure 2). Like gap junctions, plasmodesmata are channels that connect the cytosol of a cell with that of an adjacent cell. The diameter of the channel is about 30–60 nm, and its length can vary, but may be greater than 1 μm. The density of plasmodesmata varies depending on the plant and cell type, and even the smallest meristematic cells have more than a thousand connections with their neighbors. An adapter protein called NET1A is thought to link the plasmodesmata to the actin cytoskeleton. Although a variety of proteins and polysaccharides that are physically or functionally associated with plasmodesmata have been identified, the key structural protein components of plasmodesmata and the detailed mechanisms underlying their biogenesis remain to be identified.
Fig2. Plasmodesmata. (a) Schematic model of plasmodesmata, showing the desmotubule, an extension of the endoplasmic reticulum (ER), and the annulus, a plasma-membrane-lined channel filled with cytosol that interconnects the cytosols of adjacent cells. The regulated deposition of a glucose polymer called callose (cyan) in the extracellular spaces in the cell wall adjacent to the entrances of the channels has the potential to block intercellular transport through the plasmodesmata, apparently by forcing the closing of the channels by narrowing the annulus. (b) Electron micrographs of thin sections of a sugarcane leaf (brackets indicate individual plasmodesmata). Left: Longitudinal view, showing ER and desmotubule running through each annulus. Right: Perpendicular cross-sectional views of plasmodesmata, in some of which spoke structures connecting the plasma membrane to the desmotubule can be seen. [Part (b) republished with permission of Springer, from Robinson-Beers, K. and Evert, R.F., “Fine structure of plasmodesmata in mature leaves of sugarcane,” Planta, 1991, 184(3):307–18; permission conveyed through the Copyright Clearance Center, Inc.]
Molecules smaller than about 1000 Da, including a variety of metabolic and signaling compounds (ions, sugars, amino acids), can generally diffuse through plasmodesmata. However, the size of the channel through which molecules pass is highly regulated. In some circumstances, the channel is clamped shut; in others, it is dilated sufficiently to permit the passage of molecules larger than 10,000 Da. The deposition and breakdown of a glucose polymer called callose in the extracellular spaces adjacent to the entrances of the channels (see Figure 2a) is thought to regulate the closing and opening of the channels, respectively. Among the factors that affect the permeability of plasmodesmata is the cytosolic Ca2+ concentration: an increase in cytosolic Ca2+ reversibly inhibits movement of molecules through these structures.
Although plasmodesmata and gap junctions resemble each other functionally with respect to forming channels for small-molecule diffusion, their structures differ dramatically in two significant ways (see Figure 2). In plasmodesmata, the plasma membranes of the adjacent plant cells merge to form a continuous channel, called the annulus, whereas the plasma membranes of animal cells at a gap junction are not continuous with each other. There are simple plasmodesmata (with a single pore, like those in Figure 2) and complex plasmodesmata that branch into multiple channels. In addition, plasmodesmata exhibit many additional complex structural and functional characteristics. For example, they contain within the channel an extension of the endoplasmic reticulum, called a desmotubule, that passes through the annulus. They also have a variety of specialized proteins at the entrance of the channel and running throughout the length of the channel, including cytoskeletal, motor, and docking proteins that regulate the sizes and types of molecules that can pass through the channel. Many types of molecules spread from cell to cell through plasmodesmata, including some transcription fac tors, nucleic acid/protein complexes, metabolic products, and plant viruses. It appears that some of these require special chaperones to facilitate transport. Specialized kinases may also phosphorylate plasmodesmal components to regulate their activities (e.g., opening of the channels). Soluble molecules pass through the cytosolic annulus, about 3–4 nm in diameter, that lies between the plasma membrane and desmotubule, whereas membrane-bound molecules or certain proteins within the ER lumen can pass from cell to cell via the desmotubule. Plasmodesmata appear to play an especially important role in protection from pathogens and in regulating the development of plant cells and tis sues, as is suggested by their ability to mediate intracellular movement of transcription factors and ribonuclear protein complexes.
Tunneling Nanotubes Resemble Plasmodesmata and Transfer Molecules and Organelles Between Animal Cells
Tunneling nanotubes are tubelike projections of the plasma membrane that form a continuous channel connecting the cytosols of animal cells (Figure 3) and can transfer chemical and electrical signals between cells in a manner analogous to plasmodesmata in plants. Tunneling nano tubes are typically unbranched, straight tubes and can have a wide variety of diameters (50–300 nm) and lengths (extending between cells from 100 μm, they can thus can be longer than several cell diameters). All tunneling nanotubes have actin filaments passing through the central channel, and in some types of cells they also contain microtubules. There is no evidence for endoplasmic reticulum passing through tunneling nanotubes, as is the case for plasmodesmata. Remarkably, functional mitochondria can travel between cells by passing through tunneling nanotubes in cell culture (see Figure3) and in vivo, thereby rescuing receiving cells that have mitochondrial defects or deficiencies. Thus the concept of metabolic coupling can be extended to include the movement of small molecules and organelles through tunneling nano tubes. Pathogens may also use tunneling nanotubes to spread between cells.
Fig3. Microscopic visualization of a tunneling nanotube and mitochondria in cultured human cells. Cultured human retinal pigment epithelial cells (ARPE-19 cell line) were incubated with a fluorescent dye (JC-1) that specifically stains mitochondria and then examined by a combination of conventional bright field microscopy (see Chapter 4) to visualize the cells and fluorescence microscopy to visualize mitochondria (green intracellular fluorescence). A typical tunneling nanotube can be seen connecting cells 1 and 2. Inset (a) shows a higher magnification of the bright-field-only image with two bulges in the tunneling nanotube highlighted by dashed circles. Inset (b) shows a higher magnification of the same region of the combination image indicating two likely mitochondria within the tunneling nanotube at the positions of those bulges. [Wittig, D., Xiang, W., Walter, C. Hans-Hermman, G., Fun, R. H. W., Roehlecke, C. (2012) “Multi-level communication of human retinal pigment epithelial cells via tunneling nano tubes,” PLoSOne 7(3): e33195. doi:10.1371/journal.pone.0033195.]
Only a Few Adhesion Molecules Have Been Identified in Plants
Systematic analyses of the Arabidopsis genome and bio chemical analyses of other plant species have provided no evidence for the existence of plant homologs of most ani mal CAMs, adhesion receptors, and ECM components. This finding is not surprising, given the dramatically different nature of cell-cell and cell-ECM interactions in animals and plants.
Among the adhesive proteins apparently unique to plants are five wall-associated kinases (WAKs) and WAK-like proteins expressed in the plasma membrane of Arabidopsis cells. These transmembrane proteins have a cytoplasmic serine/ threonine kinase domain, and their extracellular regions contain multiple epidermal growth factor (EGF) repeats, frequently found in animal cell-surface receptors. Some WAKs have an extracellular pectin-binding domain that can recognize and bind full-length pectin and pectin degradation fragments. Such binding has been proposed to help cells monitor and respond to the status of the cell wall during normal growth and in the context of cell-wall damage (wounding) or infection by pathogens. Thus some WAKS in plant cells appear to be analogous to adhesion receptors in animal cells, binding and sensing the ECM and mediating outside-in signaling.
The results of in vitro binding assays, combined with in vivo studies and analyses of plant mutants, have identified several macromolecules in the ECM that are important for adhesion. For example, normal adhesion of pollen, which contains sperm cells, to the stigma or style in the female reproductive organ of the Easter lily requires a cysteine-rich protein called stigma/stylar cysteine-rich adhesin (SCA) and a specialized pectin that can bind to SCA (Figure 4). A small, probably ECM-embedded, 10-kDa protein called chymocyanin works in conjunction with SCA to help direct the movement of the sperm-containing pollen tube (chemotaxis) to the ovary.
Disruption of the gene encoding glucuronyltransferase 1, a key enzyme in pectin biosynthesis, has provided a striking illustration of the importance of pectins in intercellular adhesion in plant meristems. Normally, specialized pectin molecules help hold the cells in meristems tightly together. When grown in culture as a cluster of relatively undifferentiated cells, called a callus, normal meristematic cells adhere tightly and can differentiate into chlorophyll-producing cells, giving the callus a green color. Eventually the callus will generate shoots. In contrast, mutant cells with an inactivated glucuronyltransferase 1 gene are large, associate loosely with one another, and do not differentiate normally, forming a yellow callus. The introduction of a normal glucuronyltransferase 1 gene into the mutant cells restores their ability to adhere and differentiate normally.
The paucity of plant adhesion molecules identified to date, in contrast to the many well-defined animal adhesion molecules, may be due to the technical difficulties of working with the ECM/cell wall of plants. Adhesive interactions are likely to play different roles in plant and animal biology, at least in part because of the differences in development and physiology between plants and animals.
Fig4. An in vitro assay was used to identify molecules required for adherence of pollen tubes to the stylar ECM. In this assay, ECM collected from lily styles (SE) or an artificial ECM was dried on nitrocellulose membranes (NC). Pollen tubes containing sperm were then added, and their binding to the dried ECM was assessed. In this scanning electron micrograph, the tips of pollen tubes (arrows) can be seen binding to dried stylar ECM. This type of assay has shown that pollen adherence depends on stigma/stylar cysteine-rich adhesin (SCA) and a pectin that binds to SCA. [Republished with permission of Springer, from Guang Yuh, J., et al., “Adhesion of lily pollen tubes on an artificial matrix,” Sex. Plant Reprod., 1997, 10:3, pp. 173–180.]
