The Cytoskeleton

• establishing cell shape
• providing mechanical strength
• locomotion
• chromosome separation in mitosis and meiosis
• intracellular transport of organelles

• Actin filaments (also called microfilaments)
• Intermediate filaments and
• Microtubules

Actin Filaments

• form a band just beneath the plasma membrane that
  • provides mechanical strength to the cell
  • links transmembrane proteins (e.g., cell surface receptors) to cytoplasmic proteins
  • pinches dividing animal cells apart during cytokinesis
• generate cytoplasmic streaming in some cells
• generate locomotion in cells such as white blood cells and the amoeba
• interact with myosin ("thick") filaments in skeletal muscle fibers to provide the force of muscular contraction

Intermediate Filaments

These cytoplasmic fibers average 10 nm in diameter (and thus are "intermediate" in size between actin filaments (8 nm) and microtubules (25 nm)(as well as of the thick filaments of skeletal muscle fibers).

There are four major types of intermediate filament, each constructed from one or more proteins characteristic of it.

Despite their chemical diversity, intermediate filaments play similar roles in the cell: providing a supporting framework within the cell. For example, the nucleus is held within the cell by a basketlike network of intermediate filaments made of the protein keratin. (photo at right)

In the photo (courtesy of W. W. Franke), a fluorescent stain has been used to show the intermediate filaments of keratin in epithelial cells.

Some other functions of intermediate filaments:

• anchor the thick and thin filaments of muscle cells in a fixed position
• provide mechanical strength to the long axons found in some neurons

Microtubules

• are straight, hollow cylinders
• have a diameter of about 25 nm
• are variable in length but can grow 1000 times as long as they are thick
• are built by the assembly of dimers of alpha tubulin and beta tubulin.
• are found in both animal and plant cells

• grow at each end by the polymerization of tubulin dimers, and
• shrink at each end by the release of tubulin dimers (depolymerization)

However, both processes always occur more rapidly at one end, called the plus end. The other, less active, end is the minus end.

Microtubules participate in a wide variety of cell activities. Most involve motion. The motion is provided by protein "motors" that use the energy of ATP to move along the microtubule.

• kinesins (which usually move toward the plus end of the microtubules) and
• dyneins (which usually move toward the minus end).

• The rapid transport of organelles, like vesicles, along the axons of neurons takes place along microtubules with their plus ends pointed toward the end of the axon. The motors are kinesins.
• The migration of chromosomes in mitosis and meiosis is powered by microtubules that make up the spindle. Both kinesins and dyneins are used as motors as we shall see.

Spindle fibers arise from a microtubule organizing center (MTOC). The MTOC in animal cells is the centrosome.

The centrosome

• located in the cytoplasm just outside the nucleus.
• Just before mitosis, the centrosome duplicates.
• The two centrosomes move apart until they are on opposite sides of the nucleus.
• As mitosis proceeds, microtubules grow out from each centrosome with their plus ends growing toward the equatorial plate forming spindle fibers.

The photo (courtesy of Tim Mitchison) shows microtubules growing in vitro from an isolated centrosome. The centrosome was supplied with a mixture of alpha and beta tubulin monomers. These spontaneously assembled into microtubules only in the presence of centrosomes.

Spindle fibers have three destinations:

• Some attach to one kinetochore of a dyad with those growing from the opposite centrosome binding to the other kinetochore of that dyad.
• Some bind to the arms of the chromosomes.
• Other microtubules continue growing from the two centrosomes until they extend between each other in a region of overlap.

All three groups of spindle fibers participate in the separation of the chromosomes at anaphase. Which motor proteins are used is still uncertain, but

• kinetochores move along the microtubules powered by minus-end motors, probably dyneins, while
• the overlapping spindle fibers move past each other (pushing the poles farther apart) powered by plus-end motors, perhaps kinesins.

Chromosome movement in mitosis also involves polymerization and depolymerization of the microtubules. Taxol, a drug found in the bark of the Pacific yew, prevents depolymerization of the microtubules of the spindle fiber. This, in turn, stops chromosome movement, and thus prevents the completion of mitosis. Small wonder, then, that taxol is being eagerly tested as an anticancer drug.

Centrioles

Centrioles are built from a cylindrical array of 9 microtubules, each of which has attached to it 2 partial microtubules.

The photo (courtesy of E. deHarven) is an electron micrograph showing a cross section of a centriole with its array of nine triplets of microtubules. The magnification is approximately 305,000.

Centrioles are needed to make cilia and flagella.

Cilia and flagella

• locomotion for the cells (e.g., sperm) or
• move fluid past the cells (e.g., ciliated epithelial cells that line our air passages and move a film of mucus towards the throat).

• many short ones, we call them cilia or
• only one or a few long ones, we call them flagella.

• a cylindrical array of 9 evenly-spaced microtubules, each with a partial microtubule attached to it. This gives the structure a "figure 8" appearance when view in cross section.
• 2 single microtubules run up through the center of the bundle, completing the so-called "9+2" pattern.
• The entire assembly is sheathed in a membrane that is simply an extension of the plasma membrane.

This electron micrograph (courtesy of Peter Satir) shows the 9+2 pattern of microtubules in a single cilium seen in cross section.

Motion of cilia and flagella is created by the microtubules sliding past one another. This requires:

• motor molecules of dynein, which link adjacent microtubules together, and
• the energy of ATP.