Ultrastructural studies revealed that most, if not all eukaryotic cells and tissue contain several types of cytoplasmic fiber. Electron microscopy distinguishes three major classes. The thicker fibers are the MICROTUBULES with a diameter of about 25nm and
an apparently hollow core. MICROFILAMENTS are the second class at 6nm in diameter. Microfilaments usually run close to the plasma membrane and are reminiscent of the thin filaments found in skeletal muscle. The third type of filaments have a diameter of about 10nm and are known as INTERMEDIATE (sized) filaments or 10nm filaments. The
densely interwoven networks of the three classes of filaments are quite resistant to mild detergent treatment, usually used to extract membrane constituents and soluble components of the cytoplasm. These filamentous "ghosts" left after detergent treatment are commonly referred to as the CYTOSKELETON or cytoskeletal framework.
This definition is misleading since the functions of the various filaments are not merely "skeletal" and certainly include "non-skeletal" properties such as force generation and movement, cell division and uptake of materials from the exterior
of the cell.
Of the three filament classes, at least one was known to investigators from well characterized model systems. Microfilaments have a similar appearance in electron
micrographs to the actin-containing thin filaments of muscle. This observation was
supported by direct biochemical analysis and experiments showing that microfilaments
in the cells specifically bind myosin sub-fragment. Detailed information on the contractile
mechanism of muscle and direct involvement of actin in this process led investigators
to suggest that actin participates in the construction of a mechanochemical apparatus
in non-muscle cells. It was realized that actin, by itself cannot contract and attempts were
made to find additional proteins (such as myosin) which might interact with actin by
mechanisms similar to those found in muscle. During the last several years a large number
of proteins were described which can interact with actin in non-muscle cells. These
include proteins which bind to the unpolymerized form of actin (G-actin) and prevent
filament formation. Other proteins may induce fragmentation of F-actin filaments into
G-actin or short fragment. They may also induce filament bundle formation, or cause
cross-linking and gelation. It is now clear that the number of actin associated proteins
within every single cell is quite high. Therefore the various interactions of actin in
such systems must be well coordinated and spatially aligned to generate force
efficiently, at the right time and the right place.(Geiger, p.1)
Remarkable progress in the study of cytoskeletal structure was made with
the introduction of immunofluorescence microscopy for visualization of cytoskeletal
elements within cells. Initially antibodies to actin were used for immunolabeling of the
microfilament system of cultured cells. The patterns revealed by such labeling were
of elaborate networks of filament bundles (stress fibers). Actin was also detected in
the leading open areas (lamella) of the cells, which is actively involved in motility,
and in perinuclear areas (narrow space between the outer and inner membranes
that make up the nuclear envelope). Since then an ever increasing number of
actin associated proteins have been localized in cells.
Microtubules appear under the electron microscope as tubes, 25nm in
diameter, composed of 13 protofilaments. The major protein constituent of microtubules
is TUBULIN. The two polypeptide chains of tubulin become assembled into the microtubular
backbone. Electron microscopy reveals several patterns of microtubule organization.
In flagella and cilia the familiar 9+2 arrangement construct the motile apparatus responsible
for movement. In mitotic cells tubulin is reorganized into the mitotic spindle, while during
interphase an elaborate network of microtubules is distributed throughout the cytoplasm.
The filament network most compatible with the term "cyto-skeleton" is
that of the intermediate filaments. These are usually the most insoluble filaments within cells.
There are five major classes of intermediate filament subunits.
- Desmin (or skeletin) is about 55,000 daltons, occurring in muscle cells, including
skeletal, cardiac and smooth muscle.
- Vimentin, at approximately 58,000 daltons, is found in mesenchymal cells and
melanocytic cells, and most cultured cells.
- Prekaratin occurs in epithelial cells. This family of poly-peptides ranging in
molecular weight from 40,000-69,000 daltons constitute the filaments of epithelial cells
and exhibit typical diversity in different epithelia.
- Glial acidic fibrillary protein, at 51,000 daltons, is found in astrocytes.
- Neurofilaments contain three polypeptides of approximately 68,000, 120,000,
and 220,000 daltons, found in many nerve cells.
Some examples of intermediate filament research include the expression of
intermediate filaments in tumor cells and the possible use of specific antibodies for tumor
diagnosis. More specifically, it appears that tumor cells retain the same type of intermediate
filaments as the tissue of origin. Thus differential diagnosis may be applied. They may also
be used for identification of cell types in amniotic fluid. This may prove to be important in
cases such as neural tube disorders in which large numbers of glial cells (positive for
glial filaments) are present and may be detected by antibody labeling.(Geiger, p.10-11)