H. Lodish - Molecular Cell Biology (5ed, Freeman, 2003) (796244), страница 73
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See text fordiscussion. [Part (a) from T. J. Byers and D. Branton, 1985, Proc. Nat’l.Acad. Sci. USA 82:6153. Courtesy of D. Branton. Part (b) adapted fromS. E. Lux, 1979, Nature 281:426, and E. J. Luna and A. L. Hitt, 1992,Science 258:955.]5.4 • The Cytoskeleton: Components and Structural FunctionsThe richest area of actin filaments in many cells lies in thecortex, a narrow zone just beneath the plasma membrane.In this region, most actin filaments are arranged in a networkthat excludes most organelles from the cortical cytoplasm.Perhaps the simplest cytoskeleton is the two-dimensionalnetwork of actin filaments adjacent to the erythrocyteplasma membrane. In more complicated cortical cytoskeletons, such as those in platelets, epithelial cells, and muscle,actin filaments are part of a three-dimensional network thatfills the cytosol and anchors the cell to the substratum.A red blood cell must squeeze through narrow blood capillaries without rupturing its membrane.
The strength andflexibility of the erythrocyte plasma membrane depend on adense cytoskeletal network that underlies the entire membrane and is attached to it at many points. The primary component of the erythrocyte cytoskeleton is spectrin, a200-nm-long fibrous protein. The entire cytoskeleton isarranged in a spoke-and-hub network (Figure 5-31a).
Eachspoke is composed of a single spectrin molecule, which extends from two hubs and cross-links them. Each hub comprises a short (14-subunit) actin filament plus adducin,tropomyosin, and tropomodulin (Figure 5-31b, inset). Thelast two proteins strengthen the network by preventing theactin filament from depolymerizing. Six or seven spokes radiate from each hub, suggesting that six or seven spectrinmolecules are bound to the same actin filament.To ensure that the erythrocyte retains its characteristicshape, the spectrin-actin cytoskeleton is firmly attached tothe overlying erythrocyte plasma membrane by two peripheral membrane proteins, each of which binds to a specificintegral membrane protein and to membrane phospholipids.Ankyrin connects the center of spectrin to band 3 protein, ananion-transport protein in the membrane. Band 4.1 protein,a component of the hub, binds to the integral membrane protein glycophorin, whose structure was discussed previously(see Figure 5-12).
Both ankyrin and band 4.1 protein alsocontain lipid-binding motifs, which help bind them to themembrane (see Table 5-3). The dual binding by ankyrin andband 4.1 ensures that the membrane is connected to both thespokes and the hubs of the spectrin-actin cytoskeleton (seeFigure 5-31b).Intermediate Filaments Support the NuclearMembrane and Help Connect Cells into TissuesIntermediate filaments typically crisscross the cytosol, forming an internal framework that stretches from the nuclear envelope to the plasma membrane (Figure 5-32). A network ofintermediate filaments is located adjacent to some cellularmembranes, where it provides mechanical support.
For example, lamin A and lamin C filaments form an orthogonallattice that is associated with lamin B. The entire supportingstructure, called the nuclear lamina, is anchored to the innernuclear membrane by prenyl anchors on lamin B.At the plasma membrane, intermediate filaments are attached by adapter proteins to specialized cell junctions called177▲ FIGURE 5-32 Fluorescence micrograph of a PtK2fibroblast cell stained to reveal keratin intermediatefilaments. A network of filaments crisscrosses the cell from thenucleus to the plasma membrane. At the plasma membrane, thefilaments are linked by adapter proteins to two types ofanchoring junctions: desmosomes between adjacent cells andhemidesmosomes between the cell and the matrix.
[Courtesy ofR. D. Goldman.]desmosomes and hemidesmosomes, which mediate cell–celladhesion and cell–matrix adhesion, respectively, particularlyin epithelial tissues. In this way, intermediate filaments in onecell are indirectly connected to intermediate filaments in aneighboring cell or to the extracellular matrix.
Because of theimportant role of cell junctions in cell adhesion and the stability of tissues, we consider their structure and relation tocytoskeletal filaments in detail in Chapter 6.Microtubules Radiate from Centrosomesand Organize Certain Subcellular StructuresLike microfilaments and intermediate filaments, microtubules are not randomly distributed in cells. Rather, microtubules radiate from the centrosome, which is the primarymicrotubule-organizing center (MTOC) in animal cells (Figure 5-33).
As detailed in Chapter 20, the two ends of a microtubule differ in their dynamic properties and arecommonly designated as the () and () ends. For this reason, microtubles can have two distinct orientations relativeto one another and to other cell structures. In many nondividing animal cells, the MTOC is located at the center of thecell near the nucleus, and the radiating microtubules are alloriented with their () ends directed toward the cell periphery.
Although most interphase animal cells contain a singleperinuclear MTOC, epithelial cells and plant cells containhundreds of MTOCs. Both of these cell types exhibit distinct178CHAPTER 5 • Biomembranes and Cell ArchitectureMicrofilaments are assembled from monomeric actinsubunits; microtubules, from ,-tubulin subunits; and intermediate filaments, from lamin subunits and other tissuespecific proteins.■In all animal and plant cells, the cytoskeleton providesstructural stability for the cell and contributes to cell movement. Some bacteria have a primitive cytoskeleton.■Actin bundles form the core of microvilli and other fingerlike projections of the plasma membrane.■Cortical spectrin-actin networks are attached to the cellmembrane by bivalent membrane–microfilament bindingproteins such as ankyrin and band 4.1 (see Figure 5-31).■Intermediate filaments are assembled into networks andbundles by various intermediate filament–binding proteins,which also cross-link intermediate filaments to the plasmaand nuclear membranes, microtubules, and microfilaments.■In some animal cells, microtubules radiate out from asingle microtubule-organizing center lying at the cell center (see Figure 5-33).
Intact microtubules appear to be necessary for endoplasmic reticulum and Golgi membranes toform into organized structures.■▲ FIGURE 5-33 Fluorescence micrograph of a Chinesehamster ovary cell stained to reveal microtubles and theMTOC. The microtubules (green), detected with an antibody totubulin, are seen to radiate from a central point, the microtubuleorganizing center (MTOC), near the nucleus. The MTOC (yellow)is detected with an antibody to a protein localized to thecentrosome. [Courtesy of R. Kuriyame.]functional or structural properties or both in different regions of the cell. The functional and structural polarity ofthese cells is linked to the orientation of microtubules withinthem.Findings from studies discussed in Chapter 20 show thatthe association of microtubules with the endoplasmic reticulum and other membrane-bounded organelles may be critical to the location and organization of these organelleswithin the cell.
For instance, if microtubules are destroyed bydrugs such as nocodazole or colcemid, the ER loses its networklike organization. Microtubules are also critical to theformation of the mitotic apparatus—the elaborate, transientstructure that captures and subsequently separates replicatedchromosomes in cell division.KEY CONCEPTS OF SECTION 5.4The Cytoskeleton: Components and StructuralFunctionsThe cytosol is the internal aqueous medium of a cell exclusive of all organelles and the cytoskeleton. It containsnumerous soluble enzymes responsible for much of thecell’s metabolic activity.■Three major types of protein filaments—actin filaments,microtubules, and intermediate filaments—make up the cytoskeleton (see Figure 5-29).■5.5 Purification of Cellsand Their PartsMany studies on cell structure and function require samplesof a particular type of cell or subcellular organelle.
Most animal and plant tissues, however, contain a mixture of celltypes; likewise, most cells are filled with a variety of organelles. In this section, we describe several commonly usedtechniques for separating different cell types and organelles.The purification of membrane proteins presents some uniqueproblems also considered here.Flow Cytometry Separates Different Cell TypesSome cell types differ sufficiently in density that they can beseparated on the basis of this physical property.
White bloodcells (leukocytes) and red blood cells (erythrocytes), for instance, have very different densities because erythrocyteshave no nucleus; thus these cells can be separated by equilibrium density centrifugation (described shortly). Becausemost cell types cannot be differentiated so easily, other techniques such as flow cytometry must be used to separatethem.A flow cytometer identifies different cells by measuringthe light that they scatter and the fluorescence that they emitas they flow through a laser beam; thus it can sort out cells ofa particular type from a mixture. Indeed, a fluorescenceactivated cell sorter (FACS), an instrument based on flow cytometry, can select one cell from thousands of other cells(Figure 5-34).
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