MECHANICAL MOLECULES THE CYTOSKELETON IS BOTH STRONG AND MOTILE Eukaryotic cells are supported by a network of struts and

cables called the cytoskeleton. In animal cells, which lack a rigid cell wall, it is the cytoskeleton that determines cell shape. In addition the cytoskeleton is responsible for cell locomotion, for moving components from place to place within the cell, and for cell division. The cytoskeleton is composed of three cytoplasmic lament networks: microtubules, microlaments, and intermediate laments. Although all three are physically strong, microtubules and microlaments have the additional role of organizing movement, both of entire cells and of structures them. The term cytoskeleton implies a rigid set of bones within the cell. Nothing could be further from the truth, and all three lament systems are highly dynamic, altering their organization in response to the needs of the cell. The molecules that make up the three lament systems have been highly conserved throughout evolution; the cytoskeletal proteins present in the cells of complex organisms such as humans are much the same as those in a simple organism such as a yeast. Although the individual molecules making up the cytoskeleton are below the limit of resolution of the light microscope, the cytoskeleton itself can be readily observed within the cell by using uorescence microscopy . MICROTUBULES Microtubules possess a combination of physical properties that allows them to participate multiple cellular functions. They can form bundles of rigid bers that make excellent structural scaffolds and hence serve an important role in the determination of cell shape. They have an inherent structural polarity that denes a polarity (front vs. back) to the cell. They provide a system of intracellular highways that supports a two-way trafc of organelles and small vesicles powered by enzymes that interact with the microtubule surface. They can be rapidly formed and broken down, a property that allows the cell to respond to subtle environmental changes. Finally, they play a role in one of the most exquisite and precise of all movements within the cell, the segregation of chromosomes at mitosis and meiosis. Animal cells contain a network of several thousand microtubules, each 25 nm in diameter. All the cells microtubules can be traced back to a single structure called the centrosome, which is tightly attached to the surface of the nucleus at the cell centre. The centrosome is the microtubule organizing center of the cell and consists of amorphous material enclosing a pair of centrioles. Centrioles have a characteristic nine-way pattern that we will meet again in cilia and agella. Microtubules in plant cells have a quite different organization, lying immediately beneath the cell membrane, oriented at right angles to the direction of cell expansion. The role of microtubules in plants is todirect the deposition of cellulose bers on the outside of the cell membrane. Cellulose synthas e, a multiprotein enzyme complex that spans the cell membrane, is thought to move along microtubules forming new parallel tracks of cellulose microbrils on the cell surface. As a consequence of

the position of the microtubules beneath the membrane, cellulose is laid down in hoops that encase the plant cell in a rigid corset and allow it to expand only in one direction. Plant cells do not contain centrioles, and it remains unclear whether their microtubules arise from a dened organizing center. Microtubules are composed of a protein called tubulin that consists of two dissimilar subunits designated and . In the human genome there are about ve -tubulin genes and roughly the same number for tubulin. There is a third member of the tubulin superfamily, tubulin, which does not itself contribute to microtubule structure but which is found at the centrosome and plays a role in initiating microtubule assembly. -Tubulin/ -tubulin dimers assemble into chains called protolaments, 13 of which make up the microtubule wall (Fig. 18.4). Within each protolament the tubulin dimers are arranged in a head-to-tail manner, and so on. This gives the microtubule an built-in molecular polarity that is reected in the way it grows. Tubulin subunits are added to, and lost from, one end much more rapidly than the other. By convention, the fast growing end is referred to as the (+) end and the slow growing end as the () end. In cells, the minus end of each microtubule is embedded in the centrosome so that only the plus ends are free to grow or shrink. This process is surprisingly complex. Individual microtubules undergo periods of slow growth followed by rapid shrinkage, sometimes disappearing completely. This phenomenon is referred to as dynamic instability. By chance, the growing ends of certain microtubules may be captured by sites at the cell membrane and stabilized, so that they are protected from shrinkage. Their further growth inuences the shape of the cell. Groups of microtubules having a common orientation make an excellent structural framework. Because microtubules are dynamic, the framework can be continually remodeled as the needs of the cell change. One of the most important tools in establishing microtubule function in cells has been the plant alkaloid colchicine. Extracted from the corms of the autumn crocus, Colchicum autumnale, colchicine has been used since Roman times as a treatment for gout. Cells exposed to colchicine lose their shape, and the movement of organelles within the cytoplasm ceases. When the drug is washed away, microtubules reassemble from the centrosome and normal functions are resumed. Another drug, taxol, obtained from the bark of the Pacic yew, Taxus brevifolia, has the opposite effect, causing large numbers of very stable microtubules to form in the cell, an effect that is difcult to reverse. Unlike animal cells, plant cells do not use their cytoskeletons to dene their cell shape, but instead use their extracellular cell walls. The direction in which a growing plant cell extends is determined in the rst instance by the direction in which the cellulose microlaments run. However, these are laid down by cellulose synthase following microtubule tracks so in fact the cytoskeleton has dened the direction of cell growth, but indirectly rather than by itself acting as a load-bearing component. MICROTUBULE-BASED MOTILITY

Cells move for a variety of reasons. Human spermatozoa in their millions swim frantically toward an ovum; the soil amoeba, Acanthamoeba (said to be the most abundant organism on Earth), crawls over and between soil particles, engulng bacteria and small organic particles as it does so. Cells in the early human embryo show similar crawling movement as they reorganize to form tissues and organs; the invasive properties of some cancer cells is due to their reverting to this highly motile embryonic state. Of course, not all cells show these obvious forms of motility, but careful observation of even the most sedentary eukaryotic cells often reveals a remarkable repertoire of intracellular movements. Both microtubules and actin laments play important roles in cell motility. We will describe microtubule-based motility rst; actin-based motility will be covered following the section on microlaments. Cilia and Flagella Cilia and agella appeared very early in the evolution of eukaryotic cells and have remained essentially unchanged to the present day. The terms cilium (meaning an eyelash) and agellum (meaning a whip) are often used arbitrarily. Generally, cilia are shorter than agella (<10 m compared to >40 m) and are present on the surface of the cell in much greater numbers (ciliated cells often have hundreds of cilia but agellated cells usually have a single agellum). The real difference, however, lies in the nature of their movement. Cilia row like oars. The movement is biphasic, consisting of an effective stroke in which the cilium is held rigid and bends only at its base and a recovery stroke in which the bend formed at the base passes out to the tip. Flagella wriggle like eels. They generate waves that pass along their length, usually from base to tip at constant amplitude. Thus the movement of water by a agellum is parallel to its axis while a cilium moves water perpendicular to its axis and, hence, perpendicular to the surface of the cell. Cilia are such a conspicuous feature of some protists that they are called ciliates. The swimming of a paramecium, for example, is generated by the coordinated motion of several thousand cilia on the cell surface. Cilia also play a number of important roles in the human body. The respiratory tract, for example, is lined with about 0.5 m2 of ciliated epithelium, bearing something like 1012 cilia. The beating of these cilia moves a belt of mucus containing inhaled particles and microorganisms away from the lungs. This activity is paralyzed by cigarette smoke, causing mucus to accumulate in the smokers lung and causing the typical cough. Despite their different pattern of beating, cilia and agella are indistinguishable structurally. Both consist of a regular arrangement of microtubules and associated proteins with the nine-way pattern we have also seen in centrioles. Unlike centrioles, cilia and agella have a central pair of microtubules, so that the overall structure is called the 9 + 2 axoneme. The axoneme is enclosed by an extension of the cell membrane. Attached to the nine outer doublet microtubules are projections, or arms, composed of a motor protein dynein. Dynein is an ATPase that converts the energy released by ATP hydrolysis into the mechanical work of ciliary and agellar beating. Using ATP produced by mitochondria near the base of the cilium or agellum as fuel, the dynein arms push on the adjacent outer doublets, forcing a

sliding movement to occur between adjacent outer doublets. Because the arms are activated in a strict sequence both around and along the axoneme and because the amount of sliding is restricted by the radial spokes and interdoublet links, sliding is converted into bending. Bacterial agella (page 264) use a fundamentally different mechanism. Like the propellor of a boat, the motion of the bacterial agellum is entirely driven by the rotary motor at its base. The bacterial agellum itself is a specialized piece of extracellular cell wall, made of one protein (agellin) that has no similarity to tubulin or dynein. Cilia and agella full of cytosol all the way to their tips, and use the ATP in that cytosol to generate force all the way along their length. microlaments.