Cell_Biology_Alberts

Cell_Biology_Alberts. The classes of macromolecules constituting the extracellular matrix in different animal tissues are broadly similar, but variations in the relative amounts of these different classes of molecules and in the ways in which they are organized give rise to an amazing diversity of materials. The matrix can become calcified to form the rock-hard structures of bone or teeth, or it can form the transparent substance of the cornea, or it can adopt the ropelike organization that gives tendons their enormous tensile strength. It forms the jelly in a jellyfish. Covering the body of a beetle or a lobster, it forms a rigid carapace. Moreover, the extracellular matrix is more than a passive scaffold to provide physical support. It has an active and complex role in regulating the behavior of the cells that touch it, inhabit it, or crawl through its meshes, influencing their survival, development, migration, proliferation, shape, and function.


Cell_Biology_Alberts

Cell_Biology_Alberts. In this section, we describe the major features of the extracellular matrix in animal tissues, with an emphasis on vertebrates. We begin with an overview of the major classes of macromolecules in the matrix, after which we turn to the structure and function of the basal lamina, the thin layer of specialized extracellular matrix that lies beneath all epithelial cells. In the sections that follow, we then describe the varied types of cell–matrix junctions through which cells are connected to the matrix. the Extracellular Matrix Is Made and Oriented by the Cells Within It


Cell_Biology_Alberts

Cell_Biology_Alberts. the Extracellular Matrix Is Made and Oriented by the Cells Within It The macromolecules that constitute the extracellular matrix are mainly produced locally by cells in the matrix. As we discuss later, these cells also help to organize the matrix: the orientation of the cytoskeleton inside the cell can control the orientation of the matrix produced outside. In most connective tissues, the matrix macromolecules are secreted by cells called fibroblasts (Figure 19–30). In certain specialized types of connective tissues, such as cartilage and bone, however, they are secreted by cells of the fibroblast family that have more specific names: chondroblasts, for example, form cartilage, and osteoblasts form bone.


Cell_Biology_Alberts

Cell_Biology_Alberts. The extracellular matrix is constructed from three major classes of macromolecules: (1) glycosaminoglycans (GAGs), which are large and highly charged polysaccharides that are usually covalently linked to protein in the form of proteoglycans; (2) fibrous proteins, which are primarily members of the collagen family; and (3) a large class of noncollagen glycoproteins, which carry conventional asparagine-linked oligosaccharides (described in Chapter 12). All three classes of macromolecule have many members and come in a great variety of shapes and


Cell_Biology_Alberts

Cell_Biology_Alberts. Figure 19–30 Fibroblasts in connective tissue. this scanning electron micrograph shows tissue from the cornea of a rat. the extracellular matrix surrounding the fibroblasts is here composed largely of collagen fibrils. the glycoproteins, hyaluronan, and proteoglycans, which normally form a hydrated gel filling the interstices of the fibrous network, have been removed by enzyme and acid treatment. (Courtesy of t. Nishida.) sizes (Figure 19–31). Mammals are thought to have almost 300 matrix proteins, including about 36 proteoglycans, about 40 collagens, and over 200 glycoproteins, which usually contain multiple subdomains and self-associate to form multimers. Add to this the large number of matrix-associated proteins and enzymes that can modify matrix behavior by cross-linking, degradation, or other mechanisms, and one begins to see that the matrix is an almost infinitely variable material. Each tissue contains its own unique blend of matrix components, resulting in an extracellular


Cell_Biology_Alberts

Cell_Biology_Alberts. or other mechanisms, and one begins to see that the matrix is an almost infinitely variable material. Each tissue contains its own unique blend of matrix components, resulting in an extracellular matrix that is specialized for the needs of that tissue.


Cell_Biology_Alberts

Cell_Biology_Alberts. The proteoglycan molecules in connective tissue typically form a highly hydrated, gel-like “ground substance” in which collagens and glycoproteins are embedded. The polysaccharide gel resists compressive forces on the matrix while permitting the rapid diffusion of nutrients, metabolites, and hormones between the blood and the tissue cells. The collagen fibers strengthen and help organize the matrix, while other fibrous proteins, such as the rubberlike elastin, give it resilience. Finally, the many matrix glycoproteins help cells migrate, settle, and differentiate in the appropriate locations. Glycosaminoglycan (GaG) Chains Occupy Large amounts of Space and Form hydrated Gels


Cell_Biology_Alberts

Cell_Biology_Alberts. Glycosaminoglycan (GaG) Chains Occupy Large amounts of Space and Form hydrated Gels Glycosaminoglycans (GAGs) are unbranched polysaccharide chains composed of repeating disaccharide units. One of the two sugars in the repeating disaccharide is always an amino sugar (N-acetylglucosamine or N-acetylgalactosamine), which in most cases is sulfated. The second sugar is usually a uronic acid (glucuronic or iduronic). Because there are sulfate or carboxyl groups on most of their sugars, GAGs are highly negatively charged (Figure 19–32). Indeed, they are the most anionic molecules produced by animal cells. Four main groups of GAGs are distinguished by their sugars, the type of linkage between the sugars, and the number and location of sulfate groups: (1) hyaluronan, (2) chondroitin sulfate and dermatan sulfate, (3) heparan sulfate, and (4) keratan sulfate.


Cell_Biology_Alberts

Cell_Biology_Alberts. Polysaccharide chains are too stiff to fold into compact globular structures, and they are strongly hydrophilic. Thus, GAGs tend to adopt highly extended conformations that occupy a huge volume relative to their mass (Figure 19–33), and they form hydrated gels even at very low concentrations. The weight of GAGs in connective tissue is usually less than 10% of the weight of proteins, but GAG chains fill most of the extracellular space. Their high density of negative charges attracts a cloud of cations, especially Na+, that are osmotically active, causing large amounts of water to be sucked into the matrix. This creates a swelling pressure, or turgor, that enables the matrix to withstand compressive forces (in contrast to collagen fibrils, which resist stretching forces). The cartilage matrix that lines the knee joint, for example, can support pressures of hundreds of atmospheres in this way.


Cell_Biology_Alberts

Cell_Biology_Alberts. Defects in the production of GAGs can affect many different body systems. In one rare human genetic disease, for example, there is a severe deficiency in the synthesis of dermatan sulfate disaccharide. The affected individuals have a short stature, a prematurely aged appearance, and generalized defects in their skin, joints, muscles, and bones. Figure 19–31 The comparative shapes and sizes of some of the major extracellular matrix macromolecules. protein is shown in green, and glycosaminoglycan (GaG) in red.


Cell_Biology_Alberts

Cell_Biology_Alberts. Figure 19–31 The comparative shapes and sizes of some of the major extracellular matrix macromolecules. protein is shown in green, and glycosaminoglycan (GaG) in red. Figure 19–32 The repeating disaccharide sequence of a heparan sulfate glycosaminoglycan (GAG) chain. these chains can consist of as many as 200 disaccharide units, but are typically less than half that size. there is a high density of negative charges along the chain due to the presence of both carboxyl and sulfate groups. the molecule is shown here with its maximal number of sulfate groups. In vivo, the proportion of sulfated and nonsulfated groups is variable. heparin typically has >70% sulfation, while heparan sulfate has <50%. hyaluronan acts as a Space Filler During tissue Morphogenesis and repair


Cell_Biology_Alberts

Cell_Biology_Alberts. hyaluronan acts as a Space Filler During tissue Morphogenesis and repair Hyaluronan (also called hyaluronic acid or hyaluronate) is the simplest of the GAGs (Figure 19–34). It consists of a regular repeating sequence of up to 25,000 disaccharide units, is found in variable amounts in all tissues and fluids in adult animals, and is especially abundant in early embryos. Hyaluronan is not a typical GAG because it contains no sulfated sugars, all its disaccharide units are identical, its chain length is enormous, and it is not generally linked covalently to any core protein. Moreover, whereas other GAGs are synthesized inside the cell and released by exocytosis, hyaluronan is spun out directly from the cell surface by an enzyme complex embedded in the plasma membrane.


Cell_Biology_Alberts

Cell_Biology_Alberts. Hyaluronan is thought to have a role in resisting compressive forces in tissues and joints. It is also important as a space filler during embryonic development, where it can be used to force a change in the shape of a structure, as a small quantity expands with water to occupy a large volume. Hyaluronan synthesized locally from the basal side of an epithelium can deform the epithelium by creating a cell-free space beneath it, into which cells subsequently migrate. In the developing heart, for example, hyaluronan synthesis helps in this way to drive formation of the valves and septa that separate the heart’s chambers. Similar processes occur in several other organs. When cell migration ends, the excess hyaluronan is generally degraded by the enzyme hyaluronidase. Hyaluronan is also produced in large quantities during wound healing, and it is an important constituent of joint fluid, in which it serves as a lubricant.


Cell_Biology_Alberts

Cell_Biology_Alberts. proteoglycans are Composed of GaG Chains Covalently Linked to a Core protein


Cell_Biology_Alberts

Cell_Biology_Alberts. Except for hyaluronan, all GAGs are covalently attached to protein as proteoglycans, which are produced by most animal cells. Membrane-bound ribosomes make the polypeptide chain, or core protein, of a proteoglycan, which is then threaded into the lumen of the endoplasmic reticulum. The polysaccharide chains are mainly assembled on this core protein in the Golgi apparatus before delivery to the exterior of the cell by exocytosis. First, a special linkage tetrasaccharide is attached to a serine side chain on the core protein to serve as a primer for polysaccharide growth; then, one sugar at a time is added by specific glycosyl transferases (Figure 19–35). While still in the Golgi apparatus, many of the polymerized sugars are covalently modified by a sequential and coordinated series of reactions. Epimerizations alter the configuration of the substituents around individual carbon atoms in the sugar molecule; sulfations increase the negative charge.


Cell_Biology_Alberts

Cell_Biology_Alberts. Proteoglycans are clearly distinguished from other glycoproteins by the nature, quantity, and arrangement of their sugar side chains. By definition, at least one of the sugar side chains of a proteoglycan must be a GAG. Whereas glycoproteins generally contain relatively short, branched oligosaccharide chains that contribute only a small fraction of their weight, proteoglycans can contain as much as globular protein (MW 50,000) glycogen (MW ~400,000) spectrin (MW 460,000) collagen (MW 290,000) Figure 19–33 The relative dimensions and volumes occupied by various macromolecules. Several proteins, a glycogen granule, and a single hydrated molecule of hyaluronan are shown. Figure 19–34 The repeating disaccharide sequence in hyaluronan, a relatively simple GAG. this ubiquitous molecule in vertebrates consists of a single long chain of up to 25,000 sugar monomers. Note the absence of sulfate groups.


Cell_Biology_Alberts

Cell_Biology_Alberts. 95% carbohydrate by weight, mostly in the form of long, unbranched GAG chains, each typically about 80 sugars long. In principle, proteoglycans have the potential for almost limitless heterogeneity. Even a single type of core protein can carry highly variable numbers and types of attached GAG chains. Moreover, the underlying repeating sequence of disaccharides in each GAG can be modified by a complex pattern of sulfate groups. The core proteins, too, are diverse, though many of them share some characteristic domains such as the LINK domain, involved in binding to GAGs.


Cell_Biology_Alberts

Cell_Biology_Alberts. Proteoglycans can be huge. The proteoglycan aggrecan, for example, which is a major component of cartilage, has a mass of about 3 × 106 daltons with over 100 GAG chains. Other proteoglycans are much smaller and have only 1–10 GAG chains; an example is decorin, which is secreted by fibroblasts and has a single GAG chain (Figure 19–36). Decorin binds to collagen fibrils and regulates fibril assembly and fibril diameter; mice that cannot make decorin have fragile skin that has reduced tensile strength. The GAGs and proteoglycans of these various types can associate to form even larger polymeric complexes in the extracellular matrix. Molecules of aggrecan, for example, assemble with hyaluronan in cartilage matrix to form aggregates that are as big as a bacterium (Figure 19–37). Moreover, besides associating with one another, GAGs and proteoglycans associate with fibrous matrix proteins such as collagen and with protein meshworks such as the basal lamina, creating extremely complex


Cell_Biology_Alberts

Cell_Biology_Alberts. besides associating with one another, GAGs and proteoglycans associate with fibrous matrix proteins such as collagen and with protein meshworks such as the basal lamina, creating extremely complex composites (Figure 19–38).


Cell_Biology_Alberts

Cell_Biology_Alberts. Not all proteoglycans are secreted components of the extracellular matrix. Some are integral components of plasma membranes and have their core protein either inserted across the lipid bilayer or attached to the lipid bilayer by a glycosylphosphatidylinositol (GPI) anchor. Among the best-characterized plasma membrane proteoglycans are the syndecans, which have a membrane-spanning core protein whose intracellular domain is thought to interact with the actin cytoskeleton and with signaling molecules in the cell cortex. Syndecans are located on the surface of many types of cells, including fibroblasts and epithelial cells. In DECORIN AGGRECAN (MW ~40,000) (MW ~3 x 106)


Cell_Biology_Alberts

Cell_Biology_Alberts. DECORIN AGGRECAN (MW ~40,000) (MW ~3 x 106) Figure 19–35 The linkage between a GAG chain and its core protein in a proteoglycan molecule. a specific link tetrasaccharide is first assembled on a serine side chain. the rest of the GaG chain, consisting mainly of a repeating disaccharide unit, is then synthesized, with one sugar being added at a time. In chondroitin sulfate, the disaccharide is composed of D-glucuronic acid and N-acetyl-D-galactosamine; in heparan sulfate, it is either D-glucuronic acid or L-iduronic acid and N-acetyl-Dglucosamine; in keratan sulfate, it is D-galactose and N-acetyl-D-glucosamine.


Cell_Biology_Alberts

Cell_Biology_Alberts. Figure 19–36 examples of a small (decorin) and a large (aggrecan) proteoglycan found in the extracellular matrix. the figure compares these two proteoglycans with a typical secreted glycoprotein molecule, pancreatic ribonuclease B. all three are drawn to scale. the core proteins of both aggrecan and decorin contain oligosaccharide chains as well as the GaG chains, but these are not shown. aggrecan typically consists of about 100 chondroitin sulfate chains and about 30 keratan sulfate chains linked to a serine-rich core protein of almost 3000 amino acids. Decorin “decorates” the surface of collagen fibrils, hence its name.


Cell_Biology_Alberts

Cell_Biology_Alberts. RIBONUCLEASE (MW ~15,000) short, branched oligosaccharide side chain fibroblasts, syndecans can be found in cell–matrix adhesions, where they modulate integrin function by interacting with fibronectin on the cell surface and with cytoskeletal and signaling proteins inside the cell. As we discuss later, syndecan and other proteoglycans also interact with soluble peptide growth factors, influencing their effects on cell growth and proliferation. Collagens are the Major proteins of the Extracellular Matrix The collagens are a family of fibrous proteins found in all multicellular animals. They are secreted in large quantities by connective-tissue cells, and in smaller quantities by many other cell types. As a major component of skin and bone, collagens are the most abundant proteins in mammals, where they constitute 25% of the total protein mass.


Cell_Biology_Alberts

Cell_Biology_Alberts. The primary feature of a typical collagen molecule is its long, stiff, triple-stranded helical structure, in which three collagen polypeptide chains, called α chains, are wound around one another in a ropelike superhelix (Figure 19–39). 0.5 µm aggrecan aggregate Figure 19–37 An aggrecan aggregate from fetal bovine cartilage. (a) an electron micrograph of an aggrecan aggregate shadowed with platinum. Many free aggrecan molecules are also visible.


Cell_Biology_Alberts

Cell_Biology_Alberts. Figure 19–37 An aggrecan aggregate from fetal bovine cartilage. (a) an electron micrograph of an aggrecan aggregate shadowed with platinum. Many free aggrecan molecules are also visible. (B) a drawing of the giant aggrecan aggregate shown in (a). It consists of about 100 aggrecan monomers (each like the one shown in Figure 19–36) noncovalently bound through the N-terminal domain of the core protein to a single hyaluronan chain. a link protein binds both to the core protein of the proteoglycan and to the hyaluronan chain, thereby stabilizing the aggregate. the link proteins are members of a family of hyaluronan-binding proteins, some of which are cell-surface proteins. the molecular mass of such a complex can be 108 daltons or more, and it occupies a volume equivalent to that of a bacterium, which is about 2 × 10–12 cm3. (a, courtesy of Lawrence rosenberg.) Figure 19–38 Proteoglycans in the extracellular matrix of rat cartilage.


Cell_Biology_Alberts

Cell_Biology_Alberts. Figure 19–38 Proteoglycans in the extracellular matrix of rat cartilage. the tissue was rapidly frozen at –196°C, and fixed and stained while still frozen (a process called freeze substitution) to prevent the GaG chains from collapsing. In this electron micrograph, the proteoglycan molecules are seen to form a fine filamentous network in which a single striated collagen fibril is embedded. the more darkly stained parts of the proteoglycan molecules are the core proteins; the faintly stained threads are the GaG chains. (reproduced from E.B. hunziker and r.K. Schenk, J. Cell Biol. 98:277–282, 1984. With permission from the rockefeller University press.)


Cell_Biology_Alberts

Cell_Biology_Alberts. Figure 19–39 The structure of a typical collagen molecule. (a) a model of part of a single collagen α chain, in which each amino acid is represented by a sphere. the chain is about 1000 amino acids long. It is arranged as a left-handed helix, with three amino acids per turn and with glycine as every third amino acid. therefore, an α chain is composed of a series of triplet Gly-X-Y sequences, in which X and Y can be any amino acid (although X is commonly proline and Y is commonly hydroxyproline, a form of proline that is chemically modified during collagen synthesis in the cell). (B) a model of part of a collagen molecule, in which three α chains, each shown in a different color, are wrapped around one another to form a triple-stranded helical rod. Glycine is the only amino acid small enough to occupy the crowded interior of the triple helix. Only a short length of the molecule is shown; the entire molecule is 300 nm long. (From a model by B.L. trus.)


Cell_Biology_Alberts

Cell_Biology_Alberts. Collagens are extremely rich in proline and glycine, both of which are important in the formation of the triple-stranded helix.


Cell_Biology_Alberts

Cell_Biology_Alberts. The human genome contains 42 distinct genes coding for different collagen α chains. Different combinations of these genes are expressed in different tissues. Although in principle thousands of types of triple-stranded collagen molecules could be assembled from various combinations of the 42 αchains, only a limited number of triple-helical combinations are possible, and roughly 40 types of collagen molecules have been found. Type I is by far the most common, being the principal collagen of skin and bone. It belongs to the class of fibrillar collagens, or fibril-forming collagens: after being secreted into the extracellular space, they assemble into higher-order polymers called collagen fibrils, which are thin structures (10–300 nm in diameter) many hundreds of micrometers long in mature tissues, where they are clearly visible in electron micrographs (Figure 19–40; see also Figure 19–38). Collagen fibrils often aggregate into larger, cablelike bundles, several micrometers in diameter,


Cell_Biology_Alberts

Cell_Biology_Alberts. where they are clearly visible in electron micrographs (Figure 19–40; see also Figure 19–38). Collagen fibrils often aggregate into larger, cablelike bundles, several micrometers in diameter, that are visible in the light microscope as collagen fibers.


Cell_Biology_Alberts

Cell_Biology_Alberts. Collagen types IX and XII are called fibril-associated collagens because they decorate the surface of collagen fibrils. They are thought to link these fibrils to one another and to other components in the extracellular matrix. Type IV is a network-forming collagen, forming a major part of basal laminae, while type VII molecules form dimers that assemble into specialized structures called anchoring fibrils. Anchoring fibrils help attach the basal lamina of multilayered epithelia to the underlying connective tissue and therefore are especially abundant in the skin. There are also a number of “collagen-like” proteins containing short collagen-like segments. These include collagen type XVII, which has a transmembrane domain and is found in hemidesmosomes, and type XVIII, the core protein of a proteoglycan in basal laminae.


Cell_Biology_Alberts

Cell_Biology_Alberts. Many proteins appear to have evolved by repeated duplications of an original DNA sequence, giving rise to a repetitive pattern of amino acids. The genes that encode the αchains of most of the fibrillar collagens provide a good example: they are very large (up to 44 kilobases in length) and contain about 50 exons. Most of 1.5 nm Figure 19–40 A fibroblast surrounded by collagen fibrils in the connective tissue of embryonic chick skin. In this electron micrograph, the fibrils are organized into bundles that run approximately at right angles to one another. therefore, some bundles are oriented longitudinally, whereas others are seen in cross section. the collagen fibrils are produced by fibroblasts. (From C. ploetz, E.I. Zycband and


Cell_Biology_Alberts

Cell_Biology_Alberts. D.E. Birk, J. Struct. Biol. 106:73–81, 1991. With permission from Elsevier.) the exons are 54, or multiples of 54, nucleotides long, suggesting that these collagens originated through multiple duplications of a primordial gene containing 54 nucleotides and encoding exactly six Gly-X-Y repeats (see Figure 19–39). Table 19–2 provides additional details for some of the collagen types discussed in this chapter. Secreted Fibril-associated Collagens help Organize the Fibrils


Cell_Biology_Alberts

Cell_Biology_Alberts. Table 19–2 provides additional details for some of the collagen types discussed in this chapter. Secreted Fibril-associated Collagens help Organize the Fibrils In contrast to GAGs, which resist compressive forces, collagen fibrils form structures that resist tensile forces. The fibrils have various diameters and are organized in different ways in different tissues. In mammalian skin, for example, they are woven in a wickerwork pattern so that they resist tensile stress in multiple directions; leather consists of this material, suitably preserved. In tendons, collagen fibrils are organized in parallel bundles aligned along the major axis of tension. In mature bone and in the cornea, they are arranged in orderly plywoodlike layers, with the fibrils in each layer lying parallel to one another but nearly at right angles to the fibrils in the layers on either side. The same arrangement occurs in tadpole skin (Figure 19–41).


Cell_Biology_Alberts

Cell_Biology_Alberts. The connective-tissue cells themselves determine the size and arrangement of the collagen fibrils. The cells can express one or more genes for the different types of fibrillar collagen molecules. But even fibrils composed of the same mixture of collagens have different arrangements in different tissues. How is this achieved? Part of the answer is that cells can regulate the disposition of the collagen


Cell_Biology_Alberts

Cell_Biology_Alberts. Figure 19–41 Collagen fibrils in the tadpole skin. this electron micrograph shows the plywoodlike arrangement of the fibrils: successive layers of fibrils are laid down nearly at right angles to each other. this organization is also found in mature bone and in the cornea. (Courtesy of Jerome Gross.) 5 µm (A) type IX collagen molecule fbril of type II collagen (B) (C) 100 nm Figure 19–42 Type IX collagen. (a) type IX collagen molecules binding in a periodic pattern to the surface of a fibril containing type II collagen. (B) Electron micrograph of a rotary-shadowed type-II-collagen-containing fibril in cartilage, decorated by type IX collagen molecules. (C) an individual type IX collagen molecule. (B and C, from L. Vaughan et al., J. Cell Biol. 106:991–997, 1988. With permission from the rockefeller University press.) molecules after secretion by guiding collagen fibril formation near the plasma membrane. In addition, cells can influence this organization by secreting, along with


Cell_Biology_Alberts

Cell_Biology_Alberts. the rockefeller University press.) molecules after secretion by guiding collagen fibril formation near the plasma membrane. In addition, cells can influence this organization by secreting, along with their fibrillar collagens, different kinds and amounts of other matrix macromolecules. In particular, they secrete the fibrous protein fibronectin, as we discuss later, and this precedes the formation of collagen fibrils and helps guide their organization.


Cell_Biology_Alberts

Cell_Biology_Alberts. Fibril-associated collagens, such as types IX and XII collagens, are thought to be especially important in organizing collagen fibrils. They differ from fibrillar collagens in the following ways. First, their triple-stranded helical structure is interrupted by one or two short nonhelical domains, which makes the molecules more flexible than fibrillar collagen molecules. Second, they do not aggregate with one another to form fibrils in the extracellular space. Instead, they bind in a periodic manner to the surface of fibrils formed by the fibrillar collagens. Type IX molecules bind to type-II-collagen-containing fibrils in cartilage, the cornea, and the vitreous of the eye (Figure 19–42), whereas type XII molecules bind to type-I-collagen-containing fibrils in tendons and various other tissues.


Cell_Biology_Alberts

Cell_Biology_Alberts. Fibril-associated collagens are thought to mediate the interactions of collagen fibrils with one another and with other matrix macromolecules to help determine the organization of the fibrils in the matrix. Cells help Organize the Collagen Fibrils they Secrete by Exerting tension on the Matrix Cells interact with the extracellular matrix mechanically as well as chemically, and studies in culture suggest that the mechanical interaction can have dramatic effects on the architecture of connective tissue. Thus, when fibroblasts are mixed with a meshwork of randomly oriented collagen fibrils that form a gel in a culture dish, the fibroblasts tug on the meshwork, drawing in collagen from their surroundings and thereby causing the gel to contract to a small fraction of its initial volume. By similar activities, a cluster of fibroblasts surrounds itself with a capsule of densely packed and circumferentially oriented collagen fibers.


Cell_Biology_Alberts

Cell_Biology_Alberts. If two small pieces of embryonic tissue containing fibroblasts are placed far apart on a collagen gel, the intervening collagen becomes organized into a compact band of aligned fibers that connect the two explants (Figure 19–43). The fibroblasts subsequently migrate out from the explants along the aligned collagen fibers. Thus, the fibroblasts influence the alignment of the collagen fibers, and the collagen fibers in turn affect the distribution of the fibroblasts. Fibroblasts may have a similar role in organizing the extracellular matrix inside the body. First they synthesize the collagen fibrils and deposit them in the correct orientation. Then they work on the matrix they have secreted, crawling over it and tugging on it so as to create tendons and ligaments and the tough, dense layers of connective tissue that surround and bind together most organs.


Cell_Biology_Alberts

Cell_Biology_Alberts. Figure 19–43 The shaping of the extracellular matrix by cells. this micrograph shows a region between two pieces of embryonic chick heart (rich in fibroblasts as well as heart muscle cells) that were cultured on a collagen gel for 4 days. a dense tract of aligned collagen fibers has formed between the explants, presumably as a result of the fibroblasts in the explants tugging on the collagen. (From D. Stopak and a.K. harris, Dev. Biol. 90:383–398, 1982. With permission from academic press.)


Cell_Biology_Alberts

Cell_Biology_Alberts. Many vertebrate tissues, such as skin, blood vessels, and lungs, need to be both strong and elastic in order to function. A network of elastic fibers in the extracellular matrix of these tissues gives them the resilience to recoil after transient stretch (Figure 19–44). Elastic fibers are at least five times more extensible than a rubber band of the same cross-sectional area. Long, inelastic collagen fibrils are interwoven with the elastic fibers to limit the extent of stretching and prevent the tissue from tearing.


Cell_Biology_Alberts

Cell_Biology_Alberts. The main component of elastic fibers is elastin, a highly hydrophobic protein (about 750 amino acids long), which, like collagen, is unusually rich in proline and glycine but, unlike collagen, is not glycosylated. Soluble tropoelastin (the biosynthetic precursor of elastin) is secreted into the extracellular space and assembled into elastic fibers close to the plasma membrane, generally in cell-surface infoldings. After secretion, the tropoelastin molecules become highly cross-linked to one another, generating an extensive network of elastin fibers and sheets.


Cell_Biology_Alberts

Cell_Biology_Alberts. The elastin protein is composed largely of two types of short segments that alternate along the polypeptide chain: hydrophobic segments, which are responsible for the elastic properties of the molecule; and alanineand lysine-rich α-helical segments, which are cross-linked to adjacent molecules by covalent attachment of lysine residues. Each segment is encoded by a separate exon. There is still uncertainty concerning the conformation of elastin molecules in elastic fibers and how the structure of these fibers accounts for their rubberlike properties. However, it seems that parts of the elastin polypeptide chain, like the polymer chains in ordinary rubber, adopt a loose “random coil” conformation, and it is the random coil nature of the component molecules cross-linked into the elastic fiber network that allows the network to stretch and recoil like a rubber band (Figure 19–45).


Cell_Biology_Alberts

Cell_Biology_Alberts. Elastin is the dominant extracellular matrix protein in arteries, comprising 50% of the dry weight of the largest artery—the aorta (see Figure 19–44). Mutations in the elastin gene causing a deficiency of the protein in mice or humans result in narrowing of the aorta and other arteries and excessive proliferation of smooth muscle cells in the arterial wall. Apparently, the normal elasticity of an artery is required to restrain the proliferation of these cells.


Cell_Biology_Alberts

Cell_Biology_Alberts. Elastic fibers do not consist solely of elastin. The elastin core is covered with a sheath of microfibrils, each of which has a diameter of about 10 nm. The microfibrils appear before elastin in developing tissues and seem to provide scaffolding to guide elastin deposition. Arrays of microfibrils are elastic in their own right, and in some places they persist in the absence of elastin: they help to hold the lens in its place in the eye, for example. Microfibrils are composed of a number of distinct glycoproteins, including the large glycoprotein fibrillin, which binds to


Cell_Biology_Alberts

Cell_Biology_Alberts. Figure 19–44 elastic fibers. these scanning electron micrographs show (a) a low-power view of a segment of a dog’s aorta and (B) a high-power view of the dense network of longitudinally oriented elastic fibers in the outer layer of the same blood vessel. all the other components have been digested away with enzymes and formic acid. (From K.S. haas et al., Anat. Rec. 230:86–96, 1991. With permission from Wiley-Liss.) elastin and is essential for the integrity of elastic fibers. Mutations in the fibrillin gene result in Marfan’s syndrome, a relatively common human disorder. In the most severely affected individuals, the aorta is prone to rupture; other common effects include displacement of the lens and abnormalities of the skeleton and joints. Affected individuals are often unusually tall and lanky: Abraham Lincoln is suspected to have had the condition. Fibronectin and Other Multidomain Glycoproteins help Organize the Matrix


Cell_Biology_Alberts

Cell_Biology_Alberts. Fibronectin and Other Multidomain Glycoproteins help Organize the Matrix In addition to proteoglycans, collagens, and elastic fibers, the extracellular matrix contains a large and varied assortment of glycoproteins that typically have multiple domains, each with specific binding sites for other matrix macromolecules and for receptors on the surface of cells (Figure 19–46). These proteins therefore contribute to both organizing the matrix and helping cells attach to it. Like the proteoglycans, they also guide cell movements in developing tissues, by serving Figure 19–45 Stretching a network of elastin molecules. the molecules are joined together by covalent bonds (red) to generate a cross-linked network. In this model, each elastin molecule in the network can extend and contract in a manner resembling a random coil, so that the entire assembly can stretch and recoil like a rubber band.


Cell_Biology_Alberts

Cell_Biology_Alberts. Figure 19–46 Complex glycoproteins of the extracellular matrix. Many matrix glycoproteins are large scaffold proteins containing multiple copies of specific protein-interaction domains. Each domain is folded into a discrete globular structure, and many such domains are arrayed along the protein like beads on a string. this diagram shows four representative proteins among the roughly 200 matrix glycoproteins that are found in mammals. Each protein contains multiple repeat domains, with the names listed in the key at the bottom. Fibronectin, for example, contains numerous copies of three different fibronectin repeats (types I–III, labeled here as FN1, FN2, and FN3). two type III repeats near the C-terminus contain important binding sites for cell-surface integrins, whereas other FN repeats are involved in binding fibrin, collagen, and heparin, as indicated (see Figure 19–47). Other matrix proteins contain repeated sequences resembling those of epidermal growth factor (EGF), a major


Cell_Biology_Alberts

Cell_Biology_Alberts. are involved in binding fibrin, collagen, and heparin, as indicated (see Figure 19–47). Other matrix proteins contain repeated sequences resembling those of epidermal growth factor (EGF), a major regulator of cell growth and proliferation; these repeats might serve a similar signaling function in matrix proteins. Other proteins contain domains, such as the insulin-like growth factor-binding protein (IGFBp) repeat, that bind and regulate the function of soluble growth factors. to add more structural diversity, many of these proteins are encoded by rNa transcripts that can be spliced in different ways, adding or removing exons, such as those in fibronectin. Finally, the scaffolding and regulatory functions of many matrix proteins are further expanded by assembly into multimeric forms, as shown at the right: fibronectin forms dimers linked at the C-termini, whereas tenascin and thrombospondin form N-terminally linked hexamers and trimers, respectively. Other domains include four repeats


Cell_Biology_Alberts

Cell_Biology_Alberts. at the right: fibronectin forms dimers linked at the C-termini, whereas tenascin and thrombospondin form N-terminally linked hexamers and trimers, respectively. Other domains include four repeats from thrombospondin (tSpN, tSp1, tSp3, tSp_C). VWC, von Willebrand type C; FBG, fibrinogen-like. (adapted from r.O. hynes and a. Naba, Cold Spring Harb. Perspect. Biol. 4:a004903, 2012.) as tracks along which cells can migrate or as repellents that keep cells out of forbidden areas. They can also bind and thereby influence the function of peptide growth factors and other small molecules produced by nearby cells.


Cell_Biology_Alberts

Cell_Biology_Alberts. The best-understood member of this class of matrix proteins is fibronectin, a large glycoprotein found in all vertebrates and important for many cell– matrix interactions. Mutant mice that are unable to make fibronectin die early in embryogenesis because their endothelial cells fail to form proper blood vessels. The defect is thought to result from abnormalities in the interactions of these cells with the surrounding extracellular matrix, which normally contains fibronectin.


Cell_Biology_Alberts

Cell_Biology_Alberts. Fibronectin is a dimer composed of two very large subunits joined by disulfide bonds at their C-terminal ends. Each subunit contains a series of small repeated domains, or modules, separated by short stretches of flexible polypeptide chain (Figure 19–47). Each domain is usually encoded by a separate exon, suggesting that the fibronectin gene, like the genes encoding many matrix proteins, evolved by multiple exon duplications. In the human genome, there is only one fibronectin gene, containing about 50 exons of similar size, but the transcripts can be spliced in different ways to produce multiple fibronectin isoforms (see Figure 19–46). The major repeat domain in fibronectin is called the type III fibronectin repeat, which is about 90 amino acids long and occurs at least 15 times in each subunit. This repeat is among the most common of all protein domains in vertebrates. Fibronectin Binds to Integrins


Cell_Biology_Alberts

Cell_Biology_Alberts. Fibronectin Binds to Integrins One way to analyze a complex multifunctional protein molecule such as fibronectin is to synthesize individual regions of the protein and test their ability to bind other proteins. By these and other methods, it was possible to show that one region of fibronectin binds to collagen, another to proteoglycans, and another to specific integrins on the surface of various types of cells (see Figure 19–47B). Synthetic peptides corresponding to different segments of the integrin-binding domain were then used to show that binding depends on a specific tripeptide sequence (Arg-Gly-Asp, or RGD) that is found in one of the type III repeats (see Figure 19–47C). Even very short peptides containing this RGD sequence can compete with fibronectin for the binding site on cells, thereby inhibiting the attachment of the cells to a fibronectin matrix.


Cell_Biology_Alberts

Cell_Biology_Alberts. Several extracellular proteins besides fibronectin also have an RGD sequence that mediates cell-surface binding. Many of these proteins are components of the extracellular matrix, while others are involved in blood clotting. Peptides


Cell_Biology_Alberts

Cell_Biology_Alberts. Figure 19–47 The structure of a fibronectin dimer. (a) Electron micrographs of individual fibronectin dimer molecules shadowed with platinum; red arrows mark the joined C-termini. (B) the two polypeptide chains are similar but generally not identical (being made from the same gene but from differently spliced mrNas). they are joined by two disulfide bonds near the C-termini. Each chain is almost 2500 amino acids long and is folded into multiple domains (see Figure 19–46). as indicated, some domains are specialized for binding to a particular molecule. For simplicity, not all of the known binding sites are shown. (C) the three-dimensional structure of the ninth and tenth type III fibronectin repeats, as determined by x-ray crystallography. Both the arg-Glyasp (rGD) and the “synergy” sequences shown in red are important for binding to integrins on cell surfaces. (a, from J. Engel et al., J. Mol. Biol. 150:97–120, 1981. With permission from academic press; C, from Daniel J. Leahy, Annu.


Cell_Biology_Alberts

Cell_Biology_Alberts. shown in red are important for binding to integrins on cell surfaces. (a, from J. Engel et al., J. Mol. Biol. 150:97–120, 1981. With permission from academic press; C, from Daniel J. Leahy, Annu. Rev. Cell Dev. Biol. 13:363–393, 1997. With permission from annual reviews.) 1068 Chapter 19: Cell Junctions and the Extracellular Matrix containing the RGD sequence have been useful in the development of anti-clotting drugs. Some snakes use a similar strategy to cause their victims to bleed: they secrete RGD-containing anti-clotting proteins called disintegrins into their venom.


Cell_Biology_Alberts

Cell_Biology_Alberts. The cell-surface receptors that bind RGD-containing proteins are members of the integrin family, which we describe in detail later. Each integrin specifically recognizes its own small set of matrix molecules, indicating that tight binding requires more than just the RGD sequence. Moreover, RGD sequences are not the only sequence motifs used for binding to integrins: many integrins recognize and bind to other motifs instead. tension Exerted by Cells regulates the assembly of Fibronectin Fibrils


Cell_Biology_Alberts

Cell_Biology_Alberts. Fibronectin can exist both in a soluble form, circulating in the blood and other body fluids, and as insoluble fibronectin fibrils, in which fibronectin dimers are cross-linked to one another by additional disulfide bonds and form part of the extracellular matrix. Unlike fibrillar collagen molecules, however, which can self-assemble into fibrils in a test tube, fibronectin molecules assemble into fibrils only on the surface of cells, and only where those cells possess appropriate fibronectin-binding proteins—in particular, integrins. The integrins provide a linkage from the fibronectin outside the cell to the actin cytoskeleton inside it. The linkage transmits tension to the fibronectin molecules—provided that they also have an attachment to some other structure—and stretches them, exposing cryptic binding sites in the fibronectin molecules (Figure 19–48). This allows them to bind directly to one another and to recruit additional fibronectin molecules to form a fibril (Figure 19–49).


Cell_Biology_Alberts

Cell_Biology_Alberts. cryptic binding sites in the fibronectin molecules (Figure 19–48). This allows them to bind directly to one another and to recruit additional fibronectin molecules to form a fibril (Figure 19–49). This dependence on tension and interaction with cell surfaces ensures that fibronectin fibrils assemble where there is a mechanical need for them and not in inappropriate locations such as the bloodstream.


Cell_Biology_Alberts

Cell_Biology_Alberts. Many other extracellular matrix proteins contain multiple copies of the type III fibronectin repeat (see Figure 19–46), and it is possible that tension exerted on these proteins also uncovers cryptic binding sites and thereby influences their behavior. the Basal Lamina Is a Specialized Form of Extracellular Matrix


Cell_Biology_Alberts

Cell_Biology_Alberts. the Basal Lamina Is a Specialized Form of Extracellular Matrix Thus far in this section we have reviewed the general principles underlying the structure and function of the major classes of extracellular matrix components. We now describe how some of these components are assembled into a specialized type of extracellular matrix called the basal lamina (also known as the basement membrane). This exceedingly thin, tough, flexible sheet of matrix molecules is an essential underpinning of all epithelia. Although small in volume, it has a critical role in the architecture of the body. Like the cadherins, it seems to be one of the defining features common to all multicellular animals, and it seems to have appeared very early in their evolution. The major molecular components of the basal lamina are among the most ancient extracellular matrix macromolecules.


Cell_Biology_Alberts

Cell_Biology_Alberts. Basal laminae are typically 40–120 nm thick. A sheet of basal lamina not only lies beneath epithelial cells but also surrounds individual muscle cells, fat cells, Figure 19–48 Tension-sensing by fibronectin. Some type III fibronectin repeats are thought to unfold when fibronectin is stretched. the unfolding exposes cryptic binding sites that interact with other fibronectin molecules resulting in the formation of fibronectin filaments like those shown in Figure 19–49. (From V. Vogel and M. Sheetz, Nat. Rev. Mol. Cell Biol. 7:265–275, 2006. With permission from Macmillan publishers Ltd.)


Cell_Biology_Alberts

Cell_Biology_Alberts. Figure 19–49 Organization of fibronectin into fibrils at the cell surface. this fluorescence micrograph shows the front end of a migrating mouse fibroblast. Extracellular fibronectin is stained green and intracellular actin filaments are stained red. the fibronectin is initially present as small dotlike aggregates near the leading edge of the cell. It accumulates at focal adhesions (sites of anchorage of actin filaments, discussed later) and becomes organized into fibrils parallel to the actin filaments. Integrin molecules spanning the cell membrane link the fibronectin outside the cell to the actin filaments inside it (see Figure 19–55). tension exerted on the fibronectin molecules through this linkage is thought to stretch them, exposing binding sites that promote fibril formation. (Courtesy of roumen pankov and Kenneth Yamada.) and Schwann cells (which wrap around peripheral nerve cell axons to form myelin). The basal lamina thus separates these cells and epithelia from the


Cell_Biology_Alberts

Cell_Biology_Alberts. (Courtesy of roumen pankov and Kenneth Yamada.) and Schwann cells (which wrap around peripheral nerve cell axons to form myelin). The basal lamina thus separates these cells and epithelia from the underlying or surrounding connective tissue and forms the mechanical connection between them. In other locations, such as the kidney glomerulus, a basal lamina lies between two cell sheets and functions as a selective filter (Figure 19–50). Basal laminae have more than simple structural and filtering roles, however. They are able to determine cell polarity; influence cell metabolism; organize the proteins in adjacent plasma membranes; promote cell survival, proliferation, or differentiation; and serve as highways for cell migration.


Cell_Biology_Alberts

Cell_Biology_Alberts. The mechanical role is nevertheless essential. In the skin, for example, the epithelial outer layer—the epidermis—depends on the strength of the basal lamina to keep it attached to the underlying connective tissue—the dermis. In people with genetic defects in certain basal lamina proteins or in a special type of collagen that anchors the basal lamina to the underlying connective tissue, the epidermis becomes detached from the dermis. This causes a blistering disease called junctional epidermolysis bullosa, a severe and sometimes lethal condition. Laminin and type IV Collagen are Major Components of the Basal Lamina


Cell_Biology_Alberts

Cell_Biology_Alberts. Laminin and type IV Collagen are Major Components of the Basal Lamina The basal lamina is synthesized by the cells on each side of it: the epithelial cells contribute one set of basal lamina components, while cells of the underlying bed of connective tissue (called the stroma, Greek for “bedding”) contribute another set (Figure 19–51). Although the precise composition of the mature basal lamina varies from tissue to tissue and even from region to region in the same lamina, it


Cell_Biology_Alberts

Cell_Biology_Alberts. Figure 19–50 Three ways in which basal laminae are organized. Basal laminae (yellow) surround certain cells (such as skeletal muscle cells), underlie epithelia, and are interposed between two cell sheets (as in the kidney glomerulus). Note that, in the kidney glomerulus, both cell sheets have gaps in them, and the basal lamina has a filtering as well as a supportive function, helping to determine which molecules will pass into the urine from the blood. the filtration also depends on other protein-based structures, called slit diaphragms, that span the intercellular gaps in the epithelial sheet.


Cell_Biology_Alberts

Cell_Biology_Alberts. Figure 19–51 The basal lamina in the cornea of a chick embryo. In this scanning electron micrograph, some of the epithelial cells have been removed to expose the upper surface of the matlike basal lamina. a network of collagen fibrils in the underlying connective tissue interacts with the lower face of the lamina. (Courtesy of robert trelstad.) typically contains the glycoproteins laminin, type IV collagen, and nidogen, along with the proteoglycan perlecan. Other common basal lamina components are fibronectin and type XVIII collagen (an atypical member of the collagen family, forming the core protein of a proteoglycan).


Cell_Biology_Alberts

Cell_Biology_Alberts. Laminin is the primary organizer of the sheet structure, and, early in development, basal laminae consist mainly of laminin molecules. Laminins comprise a large family of proteins, each composed of three long polypeptide chains (α, β, and γ) held together by disulfide bonds and arranged in the shape of an asymmetric bouquet, like a bunch of three flowers whose stems are twisted together at the foot but whose heads remain separate (Figure 19–52). These heterotrimers can self-assemble in vitro into a network, largely through interactions between their heads, although interaction with cells is needed to organize the network into an orderly sheet. Since there are several isoforms of each type of chain, and these can associate in different combinations, many different laminins can be produced, creating basal laminae with distinctive properties. The laminin γ1 chain is, however, a component of most laminin heterotrimers; mice lacking it die during embryogenesis because they are unable to


Cell_Biology_Alberts

Cell_Biology_Alberts. creating basal laminae with distinctive properties. The laminin γ1 chain is, however, a component of most laminin heterotrimers; mice lacking it die during embryogenesis because they are unable to make basal laminae.


Cell_Biology_Alberts

Cell_Biology_Alberts. Type IV collagen is a second essential component of mature basal laminae, and it, too, exists in several isoforms. Like the fibrillar collagens that constitute the bulk of the protein in connective tissues such as bone or tendon, type IV collagen molecules consist of three separately synthesized long protein chains that twist together to form a ropelike superhelix; however, they differ from the fibrillar collagens in that the triple-stranded helical structure is interrupted in more than 20 regions, allowing multiple bends. Type IV collagen molecules interact via their terminal domains to assemble extracellularly into a flexible, feltlike network that gives the basal lamina tensile strength.


Cell_Biology_Alberts

Cell_Biology_Alberts. Laminin and type IV collagen interact with other basal lamina components, such as the glycoprotein nidogen and the proteoglycan perlecan, resulting in a highly cross-linked network of proteins and proteoglycans (Figure 19–53). The laminin molecules that generate the initial sheet structure first join to each other while bound to receptors on the surface of the cells that produce laminin. The cell-surface receptors are primarily members of the integrin family, but another important type of laminin receptor is dystroglycan, a proteoglycan with a core protein that spans the cell membrane, dangling its GAG chains in the extracellular space. Together, these receptors organize basal lamina assembly: they hold the laminin molecules by their feet, leaving the laminin heads positioned to interact so as to form a two-dimensional network. This laminin network then coordinates the assembly of the other basal lamina components.


Cell_Biology_Alberts

Cell_Biology_Alberts. In the kidney glomerulus, an unusually thick basal lamina acts as one of the layers of a molecular filter, helping to prevent the passage of macromolecules from the blood into the urine as urine is formed (see Figure 19–50). The proteoglycan in Figure 19–52 The structure of laminin.


Cell_Biology_Alberts

Cell_Biology_Alberts. (a) the best-understood family member is laminin-111, shown here with some of its binding sites for other molecules (yellow boxes). Laminins are multidomain glycoproteins composed of three polypeptides (α, β, and γ) that are disulfide-bonded into an asymmetric crosslike structure. Each of the polypeptide chains is more than 1500 amino acids long. Five types of α chains, four types of β chains, and three types of γ chains are known, and various combinations of these subunits can assemble to form a large variety of different laminins, which are named according to numbers assigned to each of their three subunits: laminin-111, for example, contains α1, β1, and γ1 subunits. Each isoform tends to have a specific tissue distribution: laminin-332 is found in skin, laminin-211 in muscle, and laminin-411 in endothelial cells of blood vessels. through their binding sites for other proteins, laminin molecules play a central part in organizing basal laminae and anchoring them to cells. (B)


Cell_Biology_Alberts

Cell_Biology_Alberts. laminin-411 in endothelial cells of blood vessels. through their binding sites for other proteins, laminin molecules play a central part in organizing basal laminae and anchoring them to cells. (B) Electron micrographs of laminin molecules shadowed with platinum. (B, from J. Engel et al., J. Mol. Biol. 150:97–120, 1981. With permission from academic press.) the basal lamina is important for this function: when its GAG chains are removed by specific enzymes, the filtering properties of the lamina are destroyed. Type IV collagen also has a role: in a human hereditary kidney disorder (Alport syndrome), mutations in a type IV collagen gene result in an irregularly thickened and dysfunctional glomerular filter. Laminin mutations, too, can disrupt the function of the kidney filter, but in a different way—by interfering with the differentiation of the cells that contact it and support it.


Cell_Biology_Alberts

Cell_Biology_Alberts. The basal lamina can act as a selective barrier to the movement of cells, as well as a filter for molecules. The lamina beneath an epithelium, for example, usually prevents fibroblasts in the underlying connective tissue from making contact with the epithelial cells. It does not, however, stop macrophages, lymphocytes, or nerve processes from passing through it, using specialized protease enzymes to cut a hole for their transit. The basal lamina is also important in tissue regeneration after injury. When cells in tissues such as muscles, nerves, and epithelia are damaged or killed, the basal lamina often survives and provides a scaffold along which regenerating cells can migrate. In this way, the original tissue architecture is readily reconstructed.


Cell_Biology_Alberts

Cell_Biology_Alberts. A particularly striking example of the role of the basal lamina in regeneration comes from studies of the neuromuscular junction, the site where the nerve terminals of a motor neuron form a chemical synapse with a skeletal muscle cell (discussed in Chapter 11). In vertebrates, the basal lamina that surrounds the muscle cell separates the nerve and muscle cell plasma membranes at the synapse, and the synaptic region of the lamina has a distinctive chemical character,


Cell_Biology_Alberts

Cell_Biology_Alberts. Figure 19–53 A model of the molecular structure of a basal lamina. (a) the basal lamina is formed by specific interactions (B) between the proteins laminin, type IV collagen, and nidogen, and the proteoglycan perlecan. Arrows in (B) connect molecules that can bind directly to each other. there are various isoforms of type IV collagen and laminin, each with a distinctive tissue distribution. transmembrane laminin receptors (integrins and dystroglycan) in the plasma membrane are thought to organize the assembly of the basal lamina; only the integrins are shown. (Based on h. Colognato and p.D. Yurchenco, Dev. Dyn. 218:213–234, 2000. With permission from Wiley-Liss.) shell of REGENERATED residual returns to site of original junction nerve cut new acetylcholine receptors become concentrated at site of originalDEGENERATED MUSCLE AND NERVE REGENERATED junctionMUSCLE FIBER with special isoforms of type IV collagen and laminin and a proteoglycan called agrin. After a nerve or muscle injury,


Cell_Biology_Alberts

Cell_Biology_Alberts. at site of originalDEGENERATED MUSCLE AND NERVE REGENERATED junctionMUSCLE FIBER with special isoforms of type IV collagen and laminin and a proteoglycan called agrin. After a nerve or muscle injury, the basal lamina at the synapse has a central role in reconstructing the synapse at the correct location (Figure 19–54). Defects in components of the basal lamina at the synapse are responsible for some forms of muscular dystrophy, in which muscles develop normally but then degenerate later in life.


Cell_Biology_Alberts

Cell_Biology_Alberts. Cells have to Be able to Degrade Matrix, as Well as Make It The ability of cells to degrade and destroy extracellular matrix is as important as their ability to make it and bind to it. Rapid matrix degradation is required in processes such as tissue repair, and even in the seemingly static extracellular matrix of adult animals there is a slow, continuous turnover, with matrix macromolecules being degraded and resynthesized. This allows bone, for example, to be remodeled so as to adapt to changes in the stresses on it. From the point of view of individual cells, the ability to cut through matrix is crucial in two ways: it enables them to divide while embedded in matrix, and it enables them to travel through it. Cells in connective tissues generally need to be able to stretch out in order to divide. If a cell lacks the enzymes needed to degrade the surrounding matrix, it is strongly inhibited from dividing, as well as being hindered from migrating.


Cell_Biology_Alberts

Cell_Biology_Alberts. Localized degradation of matrix components is also required wherever cells have to escape from confinement by a basal lamina. It is needed during normal branching growth of epithelial structures such as glands, for example, to allow the population of epithelial cells to increase, and needed also when white blood cells migrate across the basal lamina of a blood vessel into tissues in response to infection or injury. Matrix degradation is important both for the spread of cancer cells through the body and for their ability to proliferate in the tissues that they invade (discussed in Chapter 20).


Cell_Biology_Alberts

Cell_Biology_Alberts. In general, matrix components are degraded by extracellular proteolytic enzymes (proteases) that act close to the cells that produce them. Many of these proteases belong to one of two general classes. The largest group, with about 50 members in vertebrates, is the matrix metalloproteases, which depend on bound Ca2+ or Zn2+ for activity. The second group is the serine proteases, which have a highly reactive serine in their active site. Together, metalloproteases and serine


Cell_Biology_Alberts

Cell_Biology_Alberts. Figure 19–54 Regeneration experiments demonstrating the special character of the junctional basal lamina at a neuromuscular junction. If a frog muscle and its motor nerve are destroyed, the basal lamina around each muscle cell remains intact and the sites of the old neuromuscular junctions are still recognizable. When the nerve, but not the muscle, is allowed to regenerate (upper right), the junctional basal lamina directs the regenerating nerve to the original synaptic site. When the muscle, but not the nerve, is allowed to regenerate (lower right), the junctional basal lamina causes newly made acetylcholine receptors (blue) to accumulate at the original synaptic site. these experiments show that the junctional basal lamina controls the localization of synaptic components on both sides of the lamina. Some of the molecules responsible for these effects have been identified. Motor neuron axons, for example, deposit agrin in the junctional basal lamina, where it regulates the assembly


Cell_Biology_Alberts

Cell_Biology_Alberts. of the lamina. Some of the molecules responsible for these effects have been identified. Motor neuron axons, for example, deposit agrin in the junctional basal lamina, where it regulates the assembly of acetylcholine receptors and other proteins in the junctional plasma membrane of the muscle cell. reciprocally, muscle cells deposit a particular isoform of laminin in the junctional basal lamina, and this molecule is likely to interact with specific ion channels on the presynaptic membrane of the neuron.


Cell_Biology_Alberts

Cell_Biology_Alberts. proteases cooperate to degrade matrix proteins such as collagen, laminin, and fibronectin. Some metalloproteases, such as the collagenases, are highly specific, cleaving particular proteins at a small number of sites. In this way, the structural integrity of the matrix is largely retained, while the limited amount of proteolysis that occurs is sufficient for cell migration. Other metalloproteases may be less specific, but, because they are anchored to the plasma membrane, they can act just where they are needed; it is this type of matrix metalloprotease that is crucial for a cell’s ability to divide when embedded in matrix.


Cell_Biology_Alberts

Cell_Biology_Alberts. Clearly, the activities of the proteases that degrade the matrix must be tightly controlled, if the fabric of the body is not to collapse in a heap. Numerous mechanisms are therefore employed to ensure that matrix proteases are activated only at the correct time and place. Protease activity is generally confined to the cell surface by specific anchoring proteins, by membrane-associated activators, and by the production of specific protease inhibitors in regions where protease activity is not needed. Matrix proteoglycans and Glycoproteins regulate the activities of Secreted proteins


Cell_Biology_Alberts

Cell_Biology_Alberts. Matrix proteoglycans and Glycoproteins regulate the activities of Secreted proteins The physical properties of extracellular matrix are important for its fundamental roles as a scaffold for tissue structure and as a substrate for cell anchorage and migration. The matrix also has an important impact on cell signaling. Cells communicate with each other by secreting signal molecules that diffuse through the extracellular fluid to influence other cells (discussed in Chapter 15). En route to their targets, the signal molecules encounter the tightly woven meshwork of the extracellular matrix, which contains a high density of negative charges and protein-interaction domains that can interact with the signal molecules, thereby altering their function in a variety of ways.


Cell_Biology_Alberts

Cell_Biology_Alberts. The highly charged heparan sulfate chains of proteoglycans, for example, interact with numerous secreted signal molecules, including fibroblast growth factors (FGFs) and vascular endothelial growth factor (VEGF), which (among other effects) stimulate a variety of cell types to proliferate. By providing a dense array of growth factor binding sites, proteoglycans are thought to generate large local reservoirs of these factors, limiting their diffusion and focusing their actions on nearby cells. Similarly, proteoglycans might help generate steep growth factor gradients in an embryo, which can be important in the patterning of tissues during development. FGF activity can also be enhanced by proteoglycans, which oligomerize the FGF molecules, enabling them to cross-link and activate their cell-surface receptors more effectively.


Cell_Biology_Alberts

Cell_Biology_Alberts. The importance of proteoglycans as regulators of the distribution and activity of signal molecules is illustrated by the severe developmental defects that can occur when specific proteoglycans are inactivated by mutation. In Drosophila, for example, the function of several signal proteins during development is governed by interactions with the membrane-associated proteoglycans Dally and Dally-like. These members of the glypican family are thought to concentrate signal proteins in specific locations and act as co-receptors that collaborate with the conventional cell-surface receptor proteins; as a result, they promote signaling in the correct location and prevent it in the wrong locations. In the Drosophila ovary, for example, Dally is partly responsible for the restricted localization and function of a signaling protein called Dpp, which blocks differentiation of the germ-line stem cells: when the gene encoding Dally is mutated, Dpp activity is greatly reduced and oocyte development


Cell_Biology_Alberts

Cell_Biology_Alberts. function of a signaling protein called Dpp, which blocks differentiation of the germ-line stem cells: when the gene encoding Dally is mutated, Dpp activity is greatly reduced and oocyte development is abnormal.


Cell_Biology_Alberts

Cell_Biology_Alberts. Several matrix proteins also interact with signal proteins. The type IV collagen of basal laminae interacts with Dpp in Drosophila, for example. Fibronectin contains a type III fibronectin repeat that interacts with VEGF, and another domain that interacts with another growth factor called hepatocyte growth factor (HGF), thereby promoting the activities of these factors. As discussed earlier, many matrix glycoproteins contain extensive arrays of binding domains, and the arrangement of these domains is likely to influence the presentation of signal proteins to their target cells (see Figure 19–46). Finally, many matrix glycoproteins contain domains that bind directly to specific cell-surface receptors, thereby generating signals that influence the behavior of the cells, as we describe in the next section.


Cell_Biology_Alberts

Cell_Biology_Alberts. Cells are embedded in an intricate extracellular matrix, which not only binds the cells together but also influences their survival, development, shape, polarity, and migratory behavior. The matrix contains various protein fibers interwoven in a network of glycosaminoglycan (GAG) chains. GAGs are negatively charged polysaccharide chains that (except for hyaluronan) are covalently linked to protein to form proteoglycan molecules. GAGs attract water and occupy a large volume of extracellular space. Proteoglycans are also found on the surface of cells, where they often function as co-receptors to help cells respond to secreted signal proteins. Fiber-forming proteins give the matrix strength and resilience. The fibrillar collagens (types I, II, III, V, and XI) are ropelike, triple-stranded helical molecules that aggregate into long fibrils in the extracellular space, thereby providing tensile strength. They also form structures to which cells can be anchored, often via large multidomain


Cell_Biology_Alberts

Cell_Biology_Alberts. molecules that aggregate into long fibrils in the extracellular space, thereby providing tensile strength. They also form structures to which cells can be anchored, often via large multidomain glycoproteins, such as laminin and fibronectin, that bind to integrins on the cell surface. Elasticity is provided by elastin molecules, which form an extensive cross-linked network of fibers and sheets that can stretch and recoil.


Cell_Biology_Alberts

Cell_Biology_Alberts. The basal lamina is a specialized form of extracellular matrix that underlies epithelial cells or is wrapped around certain other cell types, such as muscle cells. Basal laminae are organized on a framework of laminin molecules, which are linked together by their side-arms and bind to integrins and other receptors in the basal plasma membrane of overlying epithelial cells. Type IV collagen molecules, together with the protein nidogen and the large heparan sulfate proteoglycan perlecan, assemble into a sheetlike mesh that is an essential component of all mature basal laminae. Basal laminae provide mechanical support for epithelia; they form the interface and attachment between epithelia and connective tissue; they serve as filters in the kidney; they act as barriers to keep cells in their proper compartments; they influence cell polarity and cell differentiation; and they guide cell migration during development and tissue regeneration.


Cell_Biology_Alberts

Cell_Biology_Alberts. Cells make extracellular matrix, organize it, and degrade it. The matrix in its turn exerts powerful influences on the cells. The influences are exerted chiefly through transmembrane cell adhesion proteins that act as matrix receptors. These proteins tie the matrix outside the cell to the cytoskeleton inside it, but their role goes far beyond simple passive mechanical attachment. Through them, components of the matrix can affect almost any aspect of a cell’s behavior. The matrix receptors have a crucial role in epithelial cells, mediating their interactions with the basal lamina beneath them. They are no less important in connective-tissue cells, mediating the cells’ interactions with the matrix that surrounds them.


Cell_Biology_Alberts

Cell_Biology_Alberts. Several types of molecules can function as matrix receptors or co-receptors, including the transmembrane proteoglycans. But the principal receptors on animal cells for binding most extracellular matrix proteins are the integrins. Like the cadherins and the key components of the basal lamina, integrins are part of the fundamental architectural toolkit that is characteristic of multicellular animals. The members of this large family of homologous transmembrane adhesion molecules have a remarkable ability to transmit signals in both directions across the plasma membrane. The binding of a matrix component to an integrin can send a message into the interior of the cell, and conditions in the cell interior can send a signal outward to control binding of the integrin to the matrix. Tension applied to an integrin can cause it to tighten its grip on intracellular and extracellular structures, and loss of tension can loosen its hold, so that molecular signaling complexes fall apart on either


Cell_Biology_Alberts

Cell_Biology_Alberts. to an integrin can cause it to tighten its grip on intracellular and extracellular structures, and loss of tension can loosen its hold, so that molecular signaling complexes fall apart on either side of the membrane. In this way, integrins can serve not only to transmit mechanical and molecular signals, but also to convert one type of signal into the other.


Cell_Biology_Alberts

Cell_Biology_Alberts. Integrins are transmembrane heterodimers that Link the Extracellular Matrix to the Cytoskeleton There are many varieties of integrins, but they all conform to a common plan. An integrin molecule is composed of two noncovalently associated glycoprotein subunits called α and β. Both subunits span the cell membrane, with short intracellular C-terminal tails and large N-terminal extracellular domains (Figure 19–55). The extracellular domains bind to specific amino acid sequence motifs in extracellular matrix proteins or, in some cases, in proteins on the surfaces of other cells. The best-understood binding site for integrins is the RGD sequence mentioned earlier (see Figure 19–47), which is found in fibronectin and other extracellular matrix proteins. Some integrins bind a Leu-Asp-Val (LDV) sequence in fibronectin and other proteins. Additional integrin-binding sequences, as yet poorly defined, exist in laminins and collagens.


Cell_Biology_Alberts

Cell_Biology_Alberts. Humans contain 24 types of integrins, formed from the products of 8 different β-chain genes and 18 different α-chain genes, dimerized in different combinations. Each integrin dimer has distinctive properties and functions. Moreover, because the same integrin molecule in different cell types can have different ligand-binding specificities, it seems that additional cell-type-specific factors can interact with integrins to modulate their binding activity. The binding of integrins to their matrix ligands is also affected by the concentration of Ca2+ and Mg2+ in the extracellular medium, reflecting the presence of divalent cation-binding domains in the α and β subunits. The divalent cations can influence both the affinity and the specificity of the binding of an integrin to its extracellular ligands.


Cell_Biology_Alberts

Cell_Biology_Alberts. The intracellular portion of an integrin dimer binds to a complex of several different proteins, which together form a linkage to the cytoskeleton. For all but one of the 24 varieties of human integrins, this intracellular linkage is to actin filaments. These linkages depend on proteins that assemble at the short cytoplasmic tails of the integrin subunits (see Figure 19–55). A large adaptor protein called talin is a component of the linkage in many cases, but numerous additional proteins are also involved. Like the actin-linked cell–cell junctions formed by cadherins, the actin-linked cell–matrix junctions formed by integrins may be small, inconspicuous, and transient, or large, prominent, and durable. Examples of the latter are the focal adhesions that form when fibroblasts have sufficient time to establish strong attachments to the rigid surface of a culture dish, and the myotendinous junctions that attach muscle cells to their tendons.


Cell_Biology_Alberts

Cell_Biology_Alberts. Figure 19–55 The subunit structure of an active integrin molecule, linking extracellular matrix to the actin cytoskeleton. the N-terminal heads of the integrin chains attach directly to an extracellular protein such as fibronectin; the C-terminal intracellular tail of the integrin β subunit binds to adaptor proteins that interact with filamentous actin. the best-understood adaptor is a giant protein called talin, which contains a string of multiple domains for binding actin and other proteins, such as vinculin, that help reinforce and regulate the linkage to actin filaments. One end of talin binds to a specific site on the integrin β subunit cytoplasmic tail; other regulatory proteins, such as kindlin, bind at another site on the tail.


Cell_Biology_Alberts

Cell_Biology_Alberts. In epithelia, the most prominent cell–matrix attachment sites are the hemidesmosomes, where a specific type of integrin anchors the cells to laminin in the basal lamina. Here, uniquely, the intracellular attachment is to keratin intermediate filaments, via the intracellular adaptor proteins plectin and BP230 (Figure 19–56). Integrin Defects are responsible for Many Genetic Diseases Although there is some overlap in the activities of the different integrins—at least five bind laminin, for example—it is the diversity of integrin functions that is more remarkable. Table 19–3 lists some varieties of integrins and the problems that result when individual integrin αor β chains are defective. The β1 subunit forms dimers with at least 12 distinct α subunits and is found on almost all vertebrate cells: α5β1 is a fibronectin receptor and α6β1 is a laminin Figure 19–56 Hemidesmosomes.


Cell_Biology_Alberts

Cell_Biology_Alberts. Figure 19–56 Hemidesmosomes. (a) hemidesmosomes spot-weld epithelial cells to the basal lamina, linking laminin outside the cell to keratin filaments inside it. (B) Molecular components of a hemidesmosome. a specialized integrin (α6β4 integrin) spans the membrane, attaching to keratin filaments intracellularly via adaptor proteins called plectin and Bp230, and to laminin extracellularly. the adhesive complex also contains, in parallel with the integrin, an unusual collagen family member known as collagen type XVII; this has a membrane-spanning domain attached to its extracellular collagenous portion. Defects in any of these components can give rise to a blistering disease of the skin. One such disease, called bullous pemphigoid, is an autoimmune disease in which the immune system develops antibodies against collagen XVII or Bp230.


Cell_Biology_Alberts

Cell_Biology_Alberts. receptor on many types of cells. Mutant mice that cannot make any β1 integrins die early in embryonic development. Mice that are only unable to make the α7 subunit (the partner for β1 in muscle) survive but develop muscular dystrophy (as do mice that cannot make the laminin ligand for the α7β1 integrin).


Cell_Biology_Alberts

Cell_Biology_Alberts. The β2 subunit forms dimers with at least four types of α subunit and is expressed exclusively on the surface of white blood cells, where it has an essential role in enabling these cells to fight infection. The β2 integrins mainly mediate cell– cell rather than cell–matrix interactions, binding to specific ligands on another cell, such as an endothelial cell. The ligands are members of the Ig superfamily of cell–cell adhesion molecules. We have already described an example earlier in the chapter: an integrin of this class (αLβ2, also known as LFA1) on white blood cells enables them to attach firmly to the Ig family protein ICAM1 on vascular endothelial cells at sites of infection (see Figure 19–28B). People with the genetic disease called leukocyte adhesion deficiency fail to synthesize functional β2 subunits. As a consequence, their white blood cells lack the entire family of β2 receptors, and they suffer repeated bacterial infections.


Cell_Biology_Alberts

Cell_Biology_Alberts. The β3 integrins are found on blood platelets (as well as various other cells), and they bind several matrix proteins, including the blood clotting factor fibrinogen. Platelets have to interact with fibrinogen to mediate normal blood clotting, and humans with Glanzmann’s disease, who are genetically deficient in β3 integrins, suffer from defective clotting and bleed excessively.


Cell_Biology_Alberts

Cell_Biology_Alberts. A cell crawling through a tissue—a fibroblast or a macrophage, for example, or an epithelial cell migrating along a basal lamina—has to be able both to make and to break attachments to the matrix, and to do so rapidly if it is to travel quickly. Similarly, a circulating white blood cell has to be able to switch on or off its tendency to bind to endothelial cells in order to crawl out of a blood vessel at a site of inflammation. Furthermore, if force is to be applied where it is needed, the making and breaking of the extracellular attachments in all these cases has to be coupled to the prompt assembly and disassembly of cytoskeletal attachments inside the cell. The integrin molecules that span the membrane and mediate the attachments cannot simply be passive, rigid objects with sticky patches at their two ends. They must be able to switch between an active state, where they readily form attachments, and an inactive state, where they do not.


Cell_Biology_Alberts

Cell_Biology_Alberts. Structural studies, using a combination of electron microscopy and x-ray crystallography, suggest that integrins exist in multiple structural conformations that reflect different states of activity (Figure 19–57). In the inactive state, the external segments of the integrin dimer are folded together into a compact structure that cannot bind matrix proteins. In this state, the cytoplasmic tails of the dimer are Figure 19–57 Integrins exist in two major activity states. Inactive (folded) and active (extended) structures of an integrin molecule, based on data from x-ray crystallography and other methods.


Cell_Biology_Alberts

Cell_Biology_Alberts. Figure 19–57 Integrins exist in two major activity states. Inactive (folded) and active (extended) structures of an integrin molecule, based on data from x-ray crystallography and other methods. Figure 19–58 Activation of integrins by intracellular signaling. Signals received from outside the cell can act through various intracellular mechanisms to stimulate integrin activation. In platelets, as illustrated here, the extracellular signal protein thrombin activates a G-protein-coupled receptor on the cell surface, thereby initiating a signaling pathway that leads to activation of rap1, a member of the monomeric Gtpase family. activated rap1 interacts with the protein rIaM, which then recruits talin to the plasma membrane. together with another protein called kindlin, talin interacts with the integrin β chain to trigger integrin activation. talin then interacts with adaptor proteins such as vinculin, resulting in the formation of an actin linkage (see Figure 19–55).


Cell_Biology_Alberts

Cell_Biology_Alberts. talin regulation depends in part on an interaction between its flexible C-terminal rod domain and the N-terminal head domain that contains the integrin-binding site. this interaction is thought to maintain talin in an inactive state when it is free in the cytoplasm. When talin is recruited by rIaM to the plasma membrane, the talin head domain interacts with a phosphoinositide called pI(4,5)p2 (not shown here, but see Figure 15–28), resulting in dissociation of the rod domain. talin unfolds to expose its binding sites for integrin and other proteins.


Cell_Biology_Alberts

Cell_Biology_Alberts. hooked together, preventing their interaction with cytoskeletal linker proteins. In the active state, the two integrin subunits are unhooked at the membrane to expose the intracellular binding sites for cytoplasmic adaptor proteins, and the external domains unfold and extend, like a pair of legs, to expose a high-affinity matrix-binding site at the tips of the subunits. Thus, the switch from inactive to active states depends on a major conformational change that simultaneously exposes the external and internal ligand-binding sites at the ends of the integrin molecule. External matrix binding and internal cytoskeleton linkages are thereby coupled.


Cell_Biology_Alberts

Cell_Biology_Alberts. Switching between the inactive and active states is regulated by a variety of mechanisms that vary, depending on the needs of the cell. In some cases, activation occurs by an “outside-in” mechanism: the binding of an external matrix protein, such as the RGD sequence of fibronectin, can drive some integrins to switch from the low-affinity inactive state to the high-affinity active state. As a result, binding sites for talin and other cytoplasmic adaptor proteins are exposed on the tail of the β chain. The binding of these adaptor proteins then leads to attachment of actin filaments to the intracellular end of the integrin molecule (see Figure 19–55). In this way, when the integrin catches hold of its ligand outside the cell, the cell reacts by tying the integrin molecule to the cytoskeleton, so that force can be applied at the point of cell attachment.


Cell_Biology_Alberts

Cell_Biology_Alberts. The chain of cause and effect can also operate in reverse, from inside to outside. This “inside-out” integrin-activation process generally depends on intracellular regulatory signals that stimulate the ability of talin and other proteins to interact with the β chain of the integrin. Talin competes with the integrin α chain for its binding site on the tail of the β chain. Thus, when talin binds to the β chain, it blocks the intracellular α–βlinkage, allowing the two legs of the integrin molecule to spring apart. The regulation of “inside-out” integrin activation is particularly well understood in platelets, where an extracellular signal protein called thrombin binds to a specific G-protein-coupled receptor (GPCR) on the cell surface and thereby activates an intracellular signaling pathway that leads to integrin activation (Figure 19–58). It is likely that similar signaling pathways govern integrin activation in numerous other cell types. Integrins Cluster to Form Strong adhesions


Cell_Biology_Alberts

Cell_Biology_Alberts. Integrins Cluster to Form Strong adhesions Integrins, like other cell adhesion molecules, differ from cell-surface receptors for hormones and for other extracellular soluble signal molecules in that they usually bind their ligand with lower affinity and are present at a 10–100-fold higher concentration on the cell surface. The Velcro principle, mentioned earlier in the context of cadherin adhesion (see Figure 19–6C), operates here too. Following their activation, integrins cluster together to create a dense plaque in which many integrin molecules are anchored to cytoskeletal filaments. The resulting protein structure can be remarkably large and complex, as seen in the focal adhesion made by a fibroblast on a fibronectin-coated surface culture dish.


Cell_Biology_Alberts

Cell_Biology_Alberts. The assembly of mature cell–matrix junctional complexes depends on the recruitment of dozens of different scaffolding and signaling proteins. Talin is a major component of many cell–matrix complexes, but numerous other proteins also make important contributions. These include the integrin-linked kinase (ILK) and its binding partners pinch and parvin, which together form a trimeric complex that serves as an organizing hub at many junctions. Cell–matrix junctions also employ several actin-binding proteins, such as vinculin, zyxin, VASP, and α-actinin, to promote the assembly and organization of actin filaments. Another critical component of many cell–matrix junctions is the focal adhesion kinase (FAK), which interacts with multiple components in the junction and serves an important function in signaling, as we describe next. Extracellular Matrix attachments act through Integrins to Control Cell proliferation and Survival


Cell_Biology_Alberts

Cell_Biology_Alberts. Extracellular Matrix attachments act through Integrins to Control Cell proliferation and Survival Like other transmembrane cell adhesion proteins, integrins do more than just create attachments. They also activate intracellular signaling pathways and thereby allow control of almost any aspect of the cell’s behavior according to the nature of the surrounding matrix and the state of the cell’s attachments to it.


Cell_Biology_Alberts

Cell_Biology_Alberts. Many cells will not grow or proliferate in culture unless they are attached to extracellular matrix; nutrients and soluble growth factors in the culture medium are not enough. For some cell types, including epithelial, endothelial, and muscle cells, even cell survival depends on such attachments. When these cells lose contact with the extracellular matrix, they undergo apoptosis. This dependence of cell growth, proliferation, and survival on attachment to a substratum is known as anchorage dependence, and it is mediated mainly by integrins and the intracellular signals they generate. Mutations that disrupt or override this form of control, allowing cells to escape from anchorage dependence, occur in cancer cells and play a major part in their invasive behavior.


Cell_Biology_Alberts

Cell_Biology_Alberts. Our understanding of anchorage dependence has come mainly from studies of cells living on the surface of matrix-coated culture dishes. For connective-tissue cells that are normally surrounded by matrix on all sides, this is a far cry from the natural environment. Walking over a two-dimensional plain is very different from clambering through a three-dimensional jungle. The types of contacts that cells make with a rigid substratum are not the same as those, much less well studied, that they make with the deformable web of fibers of the extracellular matrix, and there are substantial differences in cell behavior in the two contexts. Nevertheless, it is likely that the same basic principles apply. Both in vitro and in vivo, intracellular signals generated at cell–matrix adhesion sites are crucial for cell proliferation and survival. Integrins recruit Intracellular Signaling proteins at Sites of Cell–Matrix adhesion


Cell_Biology_Alberts

Cell_Biology_Alberts. Integrins recruit Intracellular Signaling proteins at Sites of Cell–Matrix adhesion The mechanisms by which integrins signal into the cell interior are complex, involving several pathways, and integrins and conventional signaling receptors often influence one another and work together to regulate cell behavior, as we have already emphasized. The Ras/MAP kinase pathway (see Figure 15–49), for


Cell_Biology_Alberts

Cell_Biology_Alberts. Figure 19–59 Tyrosine phosphorylation at focal adhesions. a fibroblast cultured on a fibronectin-coated substratum and stained with fluorescent antibodies: actin filaments are stained green and activated proteins that contain phosphotyrosine are red, giving orange where the two components overlap. the actin filaments terminate at focal adhesions, where the cell attaches to the substratum by means of integrins. proteins containing phosphotyrosine are also concentrated at these sites, reflecting the local activation of FaK and other protein kinases. Signals generated at such adhesion sites help regulate cell division, growth, and survival. (Courtesy of Keith Burridge.) example, can be activated both by conventional signaling receptors and by integrins, but cells often need both kinds of stimulation of this pathway at the same time to give sufficient activation to induce cell proliferation. Integrins and conventional signaling receptors also cooperate to promote cell survival (discussed


Cell_Biology_Alberts

Cell_Biology_Alberts. of this pathway at the same time to give sufficient activation to induce cell proliferation. Integrins and conventional signaling receptors also cooperate to promote cell survival (discussed in Chapters 15 and 18).


Cell_Biology_Alberts

Cell_Biology_Alberts. One of the best-studied modes of integrin signaling depends on a cytoplasmic protein tyrosine kinase called focal adhesion kinase (FAK). In studies of cells cultured on plastic dishes, focal adhesions are often prominent sites of tyrosine phosphorylation (Figure 19–59), and FAK is one of the major tyrosine-phosphorylated proteins found at these sites. When integrins cluster at cell–matrix contacts, FAK is recruited to the integrin β subunit by intracellular adaptor proteins such as talin or paxillin (which binds to one type of integrin α subunit). The clustered FAK molecules phosphorylate each other on a specific tyrosine, creating a phosphotyrosine docking site for members of the Src family of cytoplasmic tyrosine kinases. In addition to phosphorylating other proteins at the adhesion sites, these kinases then phosphorylate FAK on additional tyrosines, creating docking sites for a variety of additional intracellular signaling proteins. In this way, outside-in signaling from integrins,


Cell_Biology_Alberts

Cell_Biology_Alberts. these kinases then phosphorylate FAK on additional tyrosines, creating docking sites for a variety of additional intracellular signaling proteins. In this way, outside-in signaling from integrins, via FAK and Src family kinases, is relayed into the cell in much the same way as receptor tyrosine kinases generate signals (as discussed in Chapter 15).


Cell_Biology_Alberts

Cell_Biology_Alberts. Cell–Matrix adhesions respond to Mechanical Forces Like the cell–cell junctions we described earlier, cell–matrix junctions can sense and respond to the mechanical forces that act on them. Most cell–matrix junctions, for example, are connected to a contractile actin network that tends to pull the junctions inward, away from the matrix. When cells are attached to a rigid matrix that strongly resists such pulling forces, the cell–matrix junction is able to sense the resulting high tension and trigger a response in which it recruits additional integrins and other proteins to increase the junction’s ability to withstand that tension. Cell attachment to a relatively soft matrix generates less tension and therefore a less robust response. These mechanisms allow cells to sense and respond to differences in the rigidity of extracellular matrices in different tissues.


Cell_Biology_Alberts

Cell_Biology_Alberts. We saw earlier that mechanotransduction at cadherin-based cell–cell junctions likely depends on junctional proteins that change their structure when the junction is stretched by tension (see Figure 19–12). The same is true for cell– matrix junctions. The long C-terminal tail domain of talin, for example, includes a large number of binding sites for the actin-regulatory protein vinculin. Many of these sites are hidden inside folded protein domains but are exposed when those folded talin helices 1 to 12 vinculin as talin unfolds, helix 12 is exposed and can bind to vinculin domains are unfolded by stretching the protein (Figure 19–60). The N-terminal end of talin binds integrin and the C-terminal end binds actin (see Figure 19–55); thus, when actin filaments are pulled by myosin motors inside the cell, the resulting tension stretches the talin rod, thereby exposing vinculin-binding sites. The vinculin molecules then recruit and organize additional actin filaments. Tension thereby


Cell_Biology_Alberts

Cell_Biology_Alberts. inside the cell, the resulting tension stretches the talin rod, thereby exposing vinculin-binding sites. The vinculin molecules then recruit and organize additional actin filaments. Tension thereby increases the strength of the junction.


Cell_Biology_Alberts

Cell_Biology_Alberts. Integrins are the principal cell-surface receptors used by animal cells to bind to the extracellular matrix: they function as transmembrane linkers between the extracellular matrix and the cytoskeleton. Most integrins connect to actin filaments, while those at hemidesmosomes bind to intermediate filaments. Integrin molecules are heterodimers, and the binding of extracellular matrix ligands or intracellular activator proteins such as talin results in a dramatic conformational switch from an inactive to an active state. This creates an allosteric coupling between binding to matrix outside the cell and binding to the cytoskeleton inside it, allowing the integrin to convey signals in both directions across the plasma membrane. Complex assemblies of proteins become organized around the intracellular tails of activated integrins, producing intracellular signals that can influence almost any aspect of cell behavior, from proliferation and survival, as in the phenomenon of anchorage


Cell_Biology_Alberts

Cell_Biology_Alberts. intracellular tails of activated integrins, producing intracellular signals that can influence almost any aspect of cell behavior, from proliferation and survival, as in the phenomenon of anchorage dependence, to cell polarity and guidance of migration. Integrin-based cell–matrix junctions are also capable of mechanotransduction: they can sense and respond to mechanical forces acting across the junction.


Cell_Biology_Alberts

Cell_Biology_Alberts. Each cell in a plant deposits, and is in turn completely enclosed by, an elaborate extracellular matrix called the plant cell wall. It was the thick cell walls of cork, visible in a primitive microscope, that in 1663 enabled Robert Hooke to distinguish and name cells for the first time. The walls of neighboring plant cells, cemented


Cell_Biology_Alberts

Cell_Biology_Alberts. Figure 19–60 Talin is a tension sensor at cell–matrix junctions. tension across cell–matrix junctions stimulates the local recruitment of vinculin and other actin-regulatory proteins, thereby strengthening the junction’s attachment to the cytoskeleton. the experiments presented here tested the hypothesis that tension is sensed by the talin adaptor protein that links integrins to actin filaments (see Figure 19–55). (a) the long, flexible, C-terminal region of talin is divided into a series of folded domains, some of which contain vinculin-binding sites (dark green lines) that are thought to be hidden and therefore inaccessible. One domain near the N-terminus, for example, comprises a folded bundle of 12 α helices containing five vinculin-binding sites. (B) this experiment tested the hypothesis that tension stretches the 12-helix domain, thereby exposing vinculin-binding sites. a fragment of talin containing this domain was attached to an apparatus in which the domain could be


Cell_Biology_Alberts

Cell_Biology_Alberts. hypothesis that tension stretches the 12-helix domain, thereby exposing vinculin-binding sites. a fragment of talin containing this domain was attached to an apparatus in which the domain could be stretched, as shown here. the fragment was labeled at its N-terminus with a tag that sticks to the surface of a glass slide on a microscope stage. the C-terminal end of the fragment was bound to a tiny magnetic bead, so the talin fragment could be stretched using a small magnetic electrode. the solution around the protein contained fluorescently tagged vinculin proteins. after the talin protein was stretched, excess vinculin solution was washed away, and the microscope was used to determine if any fluorescent vinculin proteins were bound to the talin protein. In the absence of stretching (top), most talin molecules did not bind vinculin. When the protein was stretched (bottom), two or three vinculin molecules were bound (only one is shown here for clarity). (adapted from a. del rio et al.,


Cell_Biology_Alberts

Cell_Biology_Alberts. most talin molecules did not bind vinculin. When the protein was stretched (bottom), two or three vinculin molecules were bound (only one is shown here for clarity). (adapted from a. del rio et al., Science 323:638– 641, 2009.) together to form the intact plant (Figure 19–61), are generally thicker, stronger, and, most important of all, more rigid than the extracellular matrix produced by animal cells. In evolving relatively rigid walls, which can be up to many micrometers thick, early plant cells forfeited the ability to crawl about and adopted a sedentary lifestyle that has persisted in all present-day plants.