Molecular Biology of the Cell - Volume 2 - Alberts B., Bray D., Lewis J., Raff M., Roberts K., Watson J. 1993

Cell adhesion, cell junctions, and the extracellular matrix
Cell recognition and adhesion

So far, we have examined how Cell junctions and the Extracellular matrix hold Cells together in mature Tissues and Organs. But how do cells assemble with one another during the Cytology/cytology/16.html">Early stages of tissue formation? At least two fundamentally different mechanisms exist. Most commonly, a tissue arises from "founder cells" whose descendants remain together simply because they are attached to extracellular matrix macromolecules and/or to other cells (Fig. 14-55). The specific properties of these junctions ultimately determine The Structure of the cellular ensemble. Epithelial cell sheets typically form in this manner, and Animal Embryonic Development largely consists of the formation, bending, and differentiation of such cell sheets to generate the tissues and Organs of the adult Organism. As a rule, all Cells of the early embryo are organized into epithelia, and only later do certain cells alter their adhesive properties, leave the sheets, and form Other types of tissues (Sections 16.1.4–16.1.11).

A different strategy of tissue formation appears more complex and involves cell migration: one cell population invades another and integrates with it (and sometimes with other migrating cells) to form a tissue of mixed origin. For example, in vertebrate embryos, neural crest cells delaminate from the epithelial (neural) tube of which they were initially a part and migrate along defined pathways to numerous other locations. There they aggregate and differentiate into various tissues, including elements of the Peripheral Nervous system (Fig. 14-56). Such a process requires a mechanism to guide cells to their destination, such as the secretion of a soluble chemical agent that attracts the migrating cells (via chemotaxis), or the deposition of adhesive molecules like Fibronectin within the extracellular matrix (Section 14.2.13), which directs cell migration along specific pathways (via contact guidance).

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Fig. 14-55. The simplest mechanism of tissue formation from cells. The descendants of founder cells are held within the epithelial layer by the basal lamina and cell-adhesion mechanisms (including specialized Intercellular junctions).

Fig. 14-56. An example of a more complex mechanism of tissue construction from cells. Neural crest cells migrate away from the epithelium on the dorsal surface of the neural tube and are directed to many other Regions of the embryo, where they form various cell groups and tissues. Here, these cells are shown assembling and differentiating into two clusters of Neurons of the peripheral nervous system, known as ganglia. Other neural crest cells within the ganglia differentiate into supporting (satellite) cells that surround the neurons.

Fig. 14-57. Light micrograph of a crawling plasmodium of the cellular slime mold Dictyostelium discoideum. (Courtesy of David Francis.)

Upon reaching its destination, a migrating cell must recognize other cells of the appropriate type in order to form a tissue with them. Even in tissues that develop without migration, their constituent cells appear to specifically recognize one another: if such a developing tissue is dissociated into single cells, they will preferentially reassociate with each other rather than with cells from a different tissue (Section 14.3.4). Apparently, this specific cell-Cell Recognition helps ensure that cells of a developing tissue remain in contact with one another while remaining segregated from the cells of neighboring tissues.

In attempts to understand how cells recognize one another in developing animal tissues, ingenious experiments have been performed on certain simple microorganisms capable of transitioning between unicellular and multicellular existence and back. Regardless of their significance as potential models for cell-cell interactions in animals, these organisms are fascinating in their own right.

14.3.1. Starving Slime Mold Myxamoebae Aggregate to Form Multicellular Fruiting Bodies [28]

The cellular slime mold Dictyostelium discoideum is a eukaryotic organism whose genome is only 10 times larger than that of a bacterium and 100 times smaller than that of a human. These organisms live in forest leaf litter as individual, motile cells called myxamoebae, which feed on Bacteria and Yeast and divide every few hours under optimal conditions (in the laboratory, they can be cultured in a liquid synthetic medium). When food supplies become depleted, myxamoebae stop dividing and aggregate to form tiny (1–2 mm) multicellular worm-like creatures (pseudoplasmodia or slugs) that crawl along, leaving a trail of slime behind them (Fig. 14-57). Each slug is formed by the aggregation of up to 100,000 cells and displays behavioral traits not exhibited by free-living myxamoebae. For example, the pseudoplasmodium is extremely sensitive to light and heat and can migrate toward a weak light source, such as the fluorescent dial of a watch; this behavior presumably helps it move toward more favorable conditions. As they move, the cells begin to differentiate, resulting about 30 hours after the onset of aggregation in The formation of a miniature, plant-like structure consisting of a stalk and a fruiting body (Fig. 14-58). The fruiting body contains numerous spores that can survive for long periods even under extremely harsh conditions. Figure 14-59 schematically illustrates the complex cell Migrations that occur during the Formation of the stalk and fruiting body. Cells at the anterior end of the slug become the stalk tissue, those following them differentiate into spores, and the trailing cells form the basal disc. Both the stalk cells and the spores become coated with an extracellular matrix (in the form of Cellulose walls), and eventually all cells except the spores die. Only when favorable conditions return do the spores germinate into free-living myxamoebae, reinitiating the cycle (Fig. 14-60).

Fig. 14-58. Various stages of fruiting body formation in Dictyostelium discoideum. (Light micrographs; courtesy of John Bonner.)

Fig. 14-59. Cell migrations during fruiting body formation in Dictyostelium discoideum. Cells from the anterior region of the slime mold move downward to form the stalk, whereas cells from the middle region migrate upward and differentiate into the spores that make up the fruiting body.

14.3.2. Slime Mold Amoebae Aggregate via Chemotaxis [29]

During pseudoplasmodium formation, individual slime mold cells aggregate via chemotaxis, which we must examine before discussing The Role of cell-Cell adhesion. One of the responses of myxamoebae to starvation is that they begin to produce and secrete cAMP, which serves as a chemotactic signal attracting other myxamoebae. (As we know from Chapter 12, cAMP functions as an intracellular signal in prokaryotic and animal cells; Dictyostelium is the only organism in which it also acts as an extracellular signaling molecule.) Aggregation appears to be initiated randomly: whichever cells happen to begin secreting cyclic AMP first attract other cells and thereby become aggregation centers. The cyclic AMP produced by these "initiator cells" is secreted in discrete "pulses" and binds to specific receptors On the surface of neighboring starving amoebae, thereby directing their movement toward the source of cyclic AMP. Such a chemotactic response can be demonstrated by applying a minute amount of cAMP via a micropipette to any point on The surface of a starving myxamoeba. The response is the immediate formation of a pseudopodium growing toward the pipette (Fig. 14-61); the pseudopodium attaches to the substratum on which The Cell rests and pulls the cell in that same direction. Once an aggregation center is established, its zone of influence rapidly expands because the aggregating cells not only respond to the cyclic AMP signal but also Relay it from cell to cell. Each pulse of cyclic AMP prompts neighboring cells not only to move toward the source of the pulse but also to emit a pulse of cyclic AMP of their own. This new pulse, released with a slight delay, in turn orients nearby cells, triggering them to release cAMP as well, and so on. In this way, regular, alternating waves of cyclic AMP ripple outward from each aggregation center, causing more distant myxamoebae to move inward in concentric or spiral waves that can be seen in time-lapse cinematography (Fig. 14-62). The advantage of such a relay system is that as the signal propagates outward from the center, it is continually renewed without attenuating over long distances. In contrast, a signal that spreads solely by diffusion gradually weakens as it travels. This difference can be clearly seen by comparing the aggregation process in Dictyostelium discoideum and D. minutum, a species that lacks a relay system. In D. minutum, the signal strength originating from each aggregation center decays rapidly, resulting in the formation of very small pseudoplasmodia and fruiting bodies.

Fig. 14-60. The life cycle of Dictyostelium discoideum. Upon starvation, free-living myxamoebae aggregate to form a motile pseudoplasmodium, which subsequently develops into a fruiting body. Under favorable conditions, spores released from the fruiting body germinate and transform back into amoebae.

Fig. 14-61. Applying a small amount of cyclic AMP to any point on the surface of a starving Dictyostelium cell (amoeba) immediately induces the formation of a pseudopodium at that spot. This mechanism enables the amoeba to move toward the source of cAMP. To affect the cell, cyclic AMP must bind to specific receptors on its surface.

14.3.3. Intercellular Adhesion in Slime Molds Depends on Specific Cell-Surface Glycoproteins [30]

In addition to activating the cyclic AMP signaling system, starvation of Dictyostelium myxamoebae induces the expression of hundreds of new genes, some of which encode cell-adhesion molecules involved in cell aggregation. For example, it is believed that one carbohydrate-binding protein (i.e., a lectin, see Section 6.3.1)—discicodin-I—is secreted by starving cells to facilitate primitive forms of contact guidance. By binding to the myxamoeba surface and to the substratum over which it migrates, it could help promote the formation of streams of myxamoebae moving toward aggregation centers, much like fibronectin directs cell migration during Gastrulation. Indeed, the binding of cells to discoidin-I depends on the same RGD tripeptide found in fibronectin and many other adhesive Proteins (Section 14.2.13).

Various Newly synthesized proteins facilitate cell-cell adhesion, enabling migrating myxamoebae to adhere tightly to one another and form a multicellular organism. During the first 8 hours of starvation, cells adhere via a Ca²⁺-dependent mechanism involving an adhesion molecule called contact site B. After 8 hours, another adhesion system comes into play, in which cell adhesion is mediated by a Ca²⁺-independent mechanism involving a cell-adhesion molecule known as contact site A. Contact sites A and B have been isolated and identified as integral Plasma Membrane glycoproteins using an ingenious immunological method illustrated in Fig. 14-63. Later, this method was also used to identify cell-adhesion molecules in vertebrates.

How do cell-surface glycoproteins, such as contact sites A and B, bind cells to one another? Figure 14-64 illustrates three possible mechanisms: 1) molecules on one cell can bind to identical molecules on neighboring cells (so-called homophilic binding); 2) molecules on one cell can bind to Different types of molecules on neighboring cells (heterophilic binding); and 3) cell-surface receptors on neighboring cells can bind to each other by secreting multivalent linker molecules. As it turns out, all three of these mechanisms operate in animals.

It is believed that contact site A mediates cell cohesion via a homophilic mechanism, because following the attachment of the protein to synthetic aggregates, these aggregates bind exclusively to cells expressing contact site A, and this binding is blocked if the cells are pretreated with Antibodies against contact site A. DNA Sequencing reveals that contact site A is a single-pass transmembrane protein, apparently unrelated to any previously known cell adhesion molecules (see below).

14.3.4. Dissociated Vertebrate Cells Can Reassociate into Organized Tissues Through Selective Cell Adhesion

The appeal of using microorganisms such as Dictyostelium in The Study of cell aggregation is that this process occurs normally in a culture dish.

Fig. 14-62. "Waves" of starving Dictyostelium amoebae moving toward an aggregation center. At such low magnification, individual amoebae are not visible. (Light micrograph; courtesy of Günter Gerisch.)

Fig. 14-63. Immunological method for identifying plasma Membrane Proteins involved in Cell-to-Cell Adhesion. In step 1, antibodies (usually rabbit) are raised against the cells under investigation or their isolated Plasma Membranes. In step 2, monovalent fragments are isolated and tested to yield an antibody preparation that blocks cell adhesion. (Monovalent fragments obtained using proteases are used—see Section 18.2.4—because they do not cross-link cells and thus do not cause "false" adhesion.)

To identify cell-surface molecules involved in cell adhesion, plasma membrane proteins are solubilized, separated from one another, and each fraction is tested for its ability to neutralize the cell aggregation-blocking activity of antibody fragments (steps 3 and 4). Fractions showing this activity are then purified and retested until a pure protein is obtained (this process is not shown in the diagram). Another immunological approach involves generating A large number of Monoclonal Antibodies (Section 4.5.4) against cell-surface Antigens and screening them to identify those that block cell adhesion. Both immunological Methods are based on an important general observation: simply applying antibodies to the cell surface does not in itself prevent normal cell adhesion; adhesion is blocked only when specific cell-surface molecules involved in adhesion serve as the targets for antibody binding.

Unfortunately, such opportunities to study cell recognition processes during The Development of Multicellular animals are rare. Usually, we can at best dissociate the cells of a developing tissue and then test their ability to reassociate in vitro. Unlike adult tissues, which are difficult to separate into individual cells, embryonic tissues of vertebrates dissociate readily upon Treatment with low concentrations of the proteolytic enzyme Trypsin, sometimes combined with the removal of extracellular calcium using a suitable chelator (e.g., EDTA). These treatments disrupt the Protein-Protein Interactions (many of which are Ca²⁺-dependent, see Section 14.3.7) that hold cells together. Remarkably, such dissociated cells often reassociate in vitro into structures resembling the original tissue. Thus, tissue structure is not merely the end product of a developmental process, but is actively maintained and stabilized by a system of mutual affinities between cells and between cells and the extracellular matrix. Therefore, it is hoped that studying the reaggregation of dissociated cells in culture will help clarify the role of cell-cell and cell-matrix adhesion in establishing and maintaining tissue Organization.

In this respect, experiments on cultured epidermal cells (Skin epithelium) are instructive. In this tissue, Ca²⁺-dependent adhesion systems play a crucial role in holding cells called keratinocytes together in a multilayered sheet on the basal lamina. Keratinocytes in the basal layer of the skin are relatively undifferentiated cells; they proliferate rapidly and supply new cells to the upper layers, where Cell Division ceases and terminal differentiation takes place (Section 17.4.2). Placed on a suitable substrate in culture, dissociated keratinocytes will divide and differentiate in much the same way. However, if the Ca²⁺ concentration in the culture is kept below normal, the Ca²⁺-dependent adhesion systems cannot function, and the keratinocytes will grow as a monolayer in which dividing and differentiating cells are intermingled. If the Ca²⁺ concentration is subsequently raised, the Spatial Organization of the cells soon changes; the monolayer transforms into a Stratified Epithelium, where proliferating cells form the basal layer adjacent to the substrate, and differentiating cells are segregated into the upper layers, just as in normal skin. This suggests that the stratification of keratinocytes according to their state of differentiation is maintained by Ca²⁺-dependent MECHANISMS OF CELL-cell adhesion (see Fig. 14-68).

14.3.5. Reaggregation of Dissociated Vertebrate Cells Depends on Tissue-Specific Recognition Systems [32]

During normal development of most tissues, sorting out of randomly mixed cells of different types does not occur (Section 16.4). Nevertheless, if dissociated embryonic cells from two different vertebrate tissues, such as The Liver and retina, are mixed together, these mixed cell aggregates will gradually sort out according to their tissue origin. This assay apparently reveals tissue-specific systems of cell recognition that hold cells together in developing tissue. Such recognition systems can also be demonstrated in another way. As shown in Figure 14-65, dissociated cells adhere more readily to aggregates of their own tissue than to aggregates of other tissues. Thus, two different assays—determining cell affinity in long-term incubation experiments (cell sorting) and evaluating the tendency of cells to attach to pre-existing aggregates—yield similar results.

What is the molecular basis for such selective cell adhesion in vertebrates? Apparently, here as in slime molds, two distinct mechanisms of cell adhesion are responsible, one of which is Ca²⁺-independent and the other Ca²⁺-dependent, with each involving a distinct family of homologous cell-surface glycoproteins.

Fig. 14-64. Three possible modes of interaction between cell-surface molecules during cell-cell adhesion.

Fig. 14-65. Tissue-specific adhesiveness of dissociated embryonic vertebrate cells as measured by the radioactive cell-binding assay. Adhesiveness can be evaluated by determining the number of labeled cells that bind to cell aggregates over a given time interval. The tendency for adhesion is greater between cells of the same type. In a commonly used modification of this assay, cells are labeled with a fluorescent or radioactive marker and examined for their binding to a monolayer of unlabeled cells in culture.

14.3.6. Plasma Membrane Glycoproteins from the Immunoglobulin Superfamily Mediate Ca²⁺-Independent Cell Adhesion in Vertebrates; Examples Include Neural Cell Adhesion Molecules (N-CAM)

To identify certain cell-surface glycoproteins involved in cell adhesion in vertebrates, the immunological method illustrated in Figure 14-63 was used. In one of the most thoroughly studied examples, monovalent antibody fragments were raised against embryonic chick retina cells. Antibodies that inhibit the reaggregation of these cells in vitro were then selected. Retina cell membrane proteins were subsequently fractionated and tested for their ability to neutralize the blocking activity of the antibodies. In this way, a large transmembrane glycoprotein (about 1000 amino acid residues) was identified and named the neural cell adhesion molecule (N-CAM). N-CAM is expressed on the surface of nerve and glial cells (Section 19.1.6), "gluing" them together via a Ca²⁺-independent mechanism. If such membrane proteins are purified and incorporated into synthetic phospholipid vesicles, these vesicles will adhere to each other as well as to cells bearing N-CAM on their surface; however, this adhesion is blocked if the cells are pretreated with monovalent antibodies against N-CAM. This indicates that N-CAM binds cells to one another through homophilic interactions that directly link two N-CAM molecules (see Fig. 14-64).

Antibodies against N-CAM disrupt normal retinal development in tissue culture, and when injected into a developing chick eye, they prevent the normal axon outgrowth of retinal Nerve Cells. As we shall see later (Section 19.7.8), this suggests that N-CAM plays a vital role in the Development of the Central Nervous System by promoting cell-cell adhesion. In addition, neural crest cells, which form the peripheral nervous system, carry abundant N-CAM on their surface while residing within the neural tube and lose it upon migration. However, when they aggregate to form ganglia, N-CAM reappears (see Fig. 14-56), indicating an essential role for N-CAM in ganglion assembly. N-CAM is also expressed during critical stages in the development of many non-neural tissues, where these molecules are thought to help hold specific cells together.

Several forms of N-CAM exist, each encoded by a separate mRNA. Different mRNAs are generated by Alternative Splicing of the RNA transcript of the same large Gene. In most forms of N-CAM, the large extracellular portion of the polypeptide chain (about 680 amino acid residues) is identical and organized into five domains homologous to the immunoglobulin domains characteristic of antibody molecules (Section 18.3.3). Thus, N-CAM belongs to the same ancient superfamily of "recognition proteins" as antibodies (Section 18.6.20). The Various Forms of N-CAM differ primarily in their membrane-associated segments and cytoplasmic domains and may therefore interact with the Cytoskeleton in different ways; in fact, one form does not span The Lipid Bilayer and is attached to The Plasma Membrane solely via a covalent linkage to phosphatidylinositol (Section 8.6.13) (Fig. 14-66), whereas another is secreted and incorporated into the extracellular matrix. The functional differences among all these forms remain unknown.

An increasing number of cell-surface glycoproteins mediating Ca²⁺-independent cell adhesion are being discovered in vertebrates, all belonging to the immunoglobulin superfamily. However, not all surface proteins involved in such adhesion belong to this superfamily; for instance, those that function only in the presence of extracellular Ca²⁺ ions belong to a different family.

14.3.7. Cadherins—A Family of Homologous Cell-Surface Glycoproteins—Mediate Ca²⁺-Dependent Cell Adhesion in Vertebrates [34]

The immunological methods presented in Figure 14-63 also played a decisive role in the discovery of three related cell-surface glycoproteins, termed cadherins, which are involved in Ca²⁺-dependent cell-cell adhesion in vertebrates. E-cadherin is found on the surface of many epithelial cells (as well as in preimplantation mammalian embryo cells), N-cadherin on the surface of nerve, Heart, and lens cells, and P-cadherin on placental and epidermal cells; like N-CAM, all of these are occasionally found in other tissues during development. These three cadherins are homologous single-pass transmembrane glycoproteins (each consisting of roughly 700 amino acid residues), and in this respect they resemble N-CAM. However, in the absence of Ca²⁺, the conformation of cadherins changes drastically, rendering them susceptible to rapid Cleavage by Proteolytic Enzymes. Because certain cells, such as endothelial cells, exhibit Ca²⁺-dependent adhesion yet express none of the three known cadherins, one can expect that new members of the cadherin family will yet be discovered.

Fig. 14-66. Schematic representation of three forms of N-CAM. In all three cases, the extracellular portion of the polypeptide chain is identical and organized into five domains resembling immunoglobulin domains. Each such domain forms a loop whose ends are linked by disulfide bridges. (Based on V. A. Cunningham et al., Science 236: 799-800, 1987. Copyright 1987 by the AAAS.)

E-cadherin, also known as liver cell adhesion molecule (L-CAM) or uvomorulin, has been characterized in the most detail. The large extracellular region of its polypeptide chain forms three homologous domains that apparently lack Homology with immunoglobulin domains. It presumably plays a crucial role in binding together the cells of various epithelia. For instance, the Са²⁺-dependent reaggregation of dissociated embryonic liver epithelial cells is blocked by antibodies against E-cadherin. In mature Epithelial Tissues, E-cadherin is typically concentrated in adhesion belts (zonulae adhaerentes), where it is believed to act as a transmembrane linker connecting the cortical Actin cytoskeletons of adjacent cells and holding them together (see Section 14.1.3). It is also involved in the compaction of blastomeres in early mouse embryos (see Section 16.2.4). During compaction, loosely arranged blastomeres flatten against one another, pack tightly, and become linked by intercellular junctions. Antibodies to E-cadherin block blastomere compaction, whereas antibodies directed against many other surface molecules of these cells have no such effect.

It seems likely that cadherins also play a pivotal role at later stages of vertebrate development, as their appearance and disappearance correlate with major morphogenetic events during which tissues segregate from one another. For example, during the formation of the neural tube and its Separation from the surface ectoderm (see Section 16.1.9), E-cadherin disappears from the cells of the developing neural epithelium and is replaced by N-cadherin (along with N-CAM) (Fig. 14-67). When neural crest cells migrate away from the neural tube, they lose N-cadherin (as well as N-CAM, see above), but re-express it later during the formation of neural ganglia (see Fig. 14-56).

The Biological Significance of the striking dependence of cadherin cell adhesion proteins on calcium ion concentration remains unknown. For instance, there is currently no evidence that extracellular calcium concentration is actively regulated to control cell-cell adhesion during development.

14.3.8. Cell surface molecules involved in cell-cell and cell-matrix adhesion can be viewed as elements of a morphogenetic code [35]

Cytophysiological, morphological, and biochemical studies indicate that even a single defined cell type employs numerous molecular mechanisms to attach to other cells and to the extracellular matrix. Some of these mechanisms are associated with specialized intercellular junctions, whereas others are not (Fig. 14-68). Because an individual cell utilizes a large array of adhesive systems, almost every cell type shares at least one cell-adhesion system with any other cell type, ensuring that all cells possess a baseline affinity for one another. Typically, cells from different tissues (and even from evolutionarily distant species) are capable of forming desmosomes, Gap Junctions, and adherens junctions with each other. This suggests that the proteins involved in such junctions are highly conserved across different tissues and species. However, just as every cell of a multicellular animal contains a specific set of surface receptors enabling it to respond in a tailored manner to a complementary set of soluble signaling molecules (Hormones or local mediators), every cell within a tissue exhibits a distinct combination (or concentration) of receptors that allow it to bind specifically to other cells or to the extracellular matrix.

Fig. 14-67. Immunofluorescence micrographs of a cross-section through a chick embryo: the developing neural tube is labeled with antibodies against E-cadherin (A) and N-cadherin (B). Note that the cells of the overlying ectoderm contain only E-cadherin, whereas the neural tube cells have lost E-cadherin and acquired N-cadherin. (Courtesy of Kohei Hatta and Masatoshi Takeichi.)

Fig. 14-68. GENERALIZED SCHEME OF the adhesive mechanisms used by typical epithelial cells to attach to each other and to the extracellular matrix (basal lamina). The left side illustrates mechanisms involving specialized regions visible in conventional electron micrographs and/or freeze-fracture replicas. The right side shows other mechanisms. In some cases, the same cell-surface glycoproteins participate in cell-cell or cell-matrix adhesion via both types of mechanisms. As noted in the text, all specialized adhesion mechanisms except gap junctions are Са²⁺-dependent; among the remaining adhesion mechanisms, only some are Са²⁺-dependent.

Unlike receptors for soluble factors, which bind their specific ligands with high affinity, receptors that bind cell-surface or extracellular-matrix molecules do so with relatively low affinity. Consequently, the action of these receptors relies on a cumulative increase in binding strength resulting from the simultaneous engagement of many receptors with many ligands on a neighboring cell or the extracellular matrix. Because any two cells possess a specific repertoire of adhesion receptors for other cells and the matrix, as well as characteristic concentrations and surface distributions of these receptors, this will determine the net affinity with which the cells bind to each other and to the matrix. It is thought that this very repertoire constitutes the "morphogenetic code" that dictates how cells organize into tissues. Because animal cells—even from closely related types—correctly sort out in vitro, they must be capable of detecting relatively small differences in adhesive properties and utilizing these differences to establish the most favorable contacts among the many possible interactions with other cells and the matrix. Observations of motile cells in culture offer insights into how this might be accomplished.

14.3.9. Highly motile cells act as sensitive detectors of small differences in adhesiveness [36]

Cells participating in embryonic morphogenetic processes are often highly motile. When such cells are dissociated and placed in a culture dish, they initially extend microspikes and lamellipodia in all directions and then actively crawl across the dish surface. This motility often coincides with The Emergence of cellular differentiation and, consequently, with the period when cell recognition processes must play a crucial role. For example, in Xenopus embryos, cells suddenly become highly motile at the midblastula transition stage, when gene METABOLISM/31.html">Transcription begins (see Section 16.1.2).

Intensive studies of cell motility using cultures of fibroblasts, neutrophils, and regenerating neurons—summarized in Chapter 11—indicate that motile cells function as extraordinarily sensitive detectors of subtle differences in adhesiveness. The microspikes and lamellipodia extended in all directions apparently participate in a "tug-of-war" process, As a result of which the cell polarizes and reliably moves toward the more adhesive region of the substrate, even when the differences in adhesiveness are very slight (see Section 11.6.3). Fibroblasts, for instance, will steadily crawl up a shallow adhesiveness gradient established on a culture dish surface. Studies of neutrophil chemotaxis suggest that a motile cell can detect differences in adhesiveness across its length as small as 1%. Similarly, cells within tissues could decode the surface "morphogenetic code" with high sensitivity, migrating decisively to establish close contact with those neighboring cells to which they exhibit the highest adhesiveness.

14.3.10. Transient contacts can initiate tissue-specific cell-cell adhesion, which is subsequently stabilized by junctional complexes [37]

Which of the numerous types of intercellular junctions described at the beginning of this chapter might operate during cell migration and mutual recognition in the formation of tissues and organs? To find out, Electron Microscopy can be used to examine contacts between neighboring cells as they move within a developing embryo or in mature tissues undergoing repair. Such studies reveal that these contacts typically do not result in the formation of organized intercellular junctions. Nevertheless, the contacting membranes are often closely apposed and run parallel, separated by a gap of 10–20 nm. It is precisely across such a distance (about 13 nm) that the Influenza virus hemagglutinin protrudes from the plasma membrane—the first plasma membrane glycoprotein whose three-dimensional structure was resolved (see Section 8.6.12). Glycoproteins on two adjacent plasma membranes can interact across a 10–20 nm gap to mediate adhesion. This type of transient contact may be optimal for cell locomotion: tight enough to provide traction, yet loose enough to allow continued cell movement.

Because junctional complexes between motile embryonic cells are generally not visible (except perhaps for small gap junctions), the formation of stable intercellular junctions may serve primarily as a mechanism to immobilize cells within an already organized tissue. A plausible hypothesis is that transient adhesion mediated by cell-surface proteins initiates tissue-specific cell-cell adhesion, which is then stabilized by the subsequent assembly of intercellular junctions. Because many of the transmembrane glycoproteins involved in this process can diffuse freely in the plane of the plasma membrane, they can accumulate at sites of cell-cell contact, thereby serving both for initial transient adhesion and for the subsequent formation of specialized junctional structures. Thus, certain cell-adhesion proteins, such as E-cadherins (see Section 14.3.7), may promote the initiation of cell adhesion and later become Structural components of intercellular junctions.

To decipher the recognition and binding rules operating during the morphogenesis of complex tissues, an ideal approach would be to inactivate various classes of cell-cell and cell-matrix adhesion receptors individually and in various combinations. As the repertoire of characterized MONOCLONAL ANTIBODIES AND protein fragments—each blocking a single type of cell adhesion molecule or matrix receptor—continues to grow, and as the genes encoding these cell surface proteins become accessible for manipulation in vitro and in Transgenic Animals, this dream of developmental biologists is becoming a reality.

Summary

Dissociated and mixed cells from different embryonic vertebrate tissues reassociate preferentially with cells of their own tissue type. Initial difficulties in analyzing the molecular mechanisms underlying the normal organization of cells into complex tissues in higher animals prompted a shift toward simpler model systems. Free-living myxamoebae of the cellular slime mold Dictyostelium discoideum aggregate upon starvation to form multicellular fruiting bodies. Their intercellular adhesion is mediated by at least two cell-surface glycoproteins: one functions early in development and is dependent on extracellular calcium concentration, whereas the other acts at a later stage and is calcium-independent. Tissue-specific recognition in vertebrates is similarly mediated by cell-surface glycoproteins belonging to at least two major families: members of one family are Са²⁺-dependent (cadherins), while those of the other are Са²⁺-independent (represented by N-CAM and other members of the immunoglobulin superfamily). Both families of cell adhesion molecules appear to play vital roles in governing vertebrate morphogenesis. Because even a single cell type employs multiple molecular mechanisms to bind to others (and to the extracellular matrix), the Specificity of cell adhesion observed during embryonic development must be the net outcome of affinities from a vast array of different adhesion systems. The capacity of motile cells to detect minute differences in adhesiveness illustrates how the specific combinations, concentrations, and distributions of cell adhesion molecules and matrix receptors characteristic of each cell type could serve as an essential "morphogenetic code."

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