Cytoskeleton: Meaning and Components (With Diagram)

In this article we will discuss about Cytoskeleton:- 1. Meaning of Cytoskeleton 2. Components of Cytoskeleton.

Meaning of Cytoskeleton:

Earlier idea of cell was that it was a collection of some cell organelles suspended in cell sap. But with the advancement of microscopic tech­niques and the discovery of electron microscopy the idea of cell has been changed radically.

Now the idea is that the cell sap is not a liquid but has network of many interconnected fibres and filaments having similarity with the bony skeleton of the animal body, i.e., an internal scaffolding of the cell.

These thread-like struc­tures can be seen under the electron micro­scope or under the fluorescence microscope by tagging them with antibodies and fluorescent dyes. These network of fibres found in a cell are known as cytoskeleton.

The fibre of the cytoskeleton extends throughout the cell having interconnection with cell membrane and cell organelles. It represents some fibrous proteins of the cytoplasm which help to maintain cell shape and give contractibility to the cell.

It also helps to facilitate communication among intracellular organelles. It also helps in cell locomotion or the movement of protoplasm, i.e., cyclosis. It also helps in the movement of cellu­lar components like chromosomes, membranes and granules, with formation of membrane protrusions (microvilli).

Components of Cytoskeleton:

On the basis of the electron microscopically observations, cytoskeleton components can be divided into three types:

i. The thickest tubular components are known as Microtubules.

ii. The thinnest fibres are called Micro­filaments.

iii. The fibres of intermediate thickness are known as Intermediate filaments.

The network of microtubules becomes denser towards the nucleus, i.e., towards the nuclear envelope and then the fibres radiate towards the surface. Microfilaments, consisting of actin fibres, were found crisscrossing the cell outline.

These, microfilaments can be seen by using antibody to actin under immunofluorescence microscopy. Microfilament bundles cross­over each other and also run parallel over long distances. Sometimes these filaments pass over the nucleus.

This type of microfilament organisation is sometimes known as stress fibres. The main function of these microfilaments is to help in the communication between the main cell components.

(i) Microtubules:

Microtubules were first noted in a number of eukaryotic cells by electron microscopic observations (Fig. 18.1). It is a long rod-like structure of 25 nm diameter and up to several millimeters in length.

It has two main characteristics that help to perform diverse type of functions of the cell:

i. Long rigid shape facilitates in support­ing and anchoring different cellular constituents.

ii. Can generate movement in the cellular components as well as in the total cell.

Till the refinement of fixation technique in electron microscopy, microtubules were observed only in some subcellular structures like cilia, flagella, centrioles, mitotic spindle etc. using osmium tetroxide in the fixative. With the use of Glutaraldehyde in the fixative, the network of microtubules in the cytoplasm of the cell, i.e., cytoskeleton, was detected.

Electron microscopy and X-ray diffraction studies show that microtubules contain some longitudinally arranged assemblies of filaments. These filaments are known as protofilaments. About 13 protofilaments form a hollow cylinder that are recognised as microtubules (Fig. 18.2).

The chemical nature of these protofilaments is of tubulin molecules that are different in their amino acid sequence and are known as α and β tubulins of approximately 110,000 molecular weight. These assemblies have an outer diam­eter of 30 nm and an inner diameter of 14 nm with a wall thickness of 8 nm. This beaded structure of protofilaments can be observed under the electron microscope.

By treating the cells with colchicine or any inhibitors of protein synthesis or at low tem­perature, the organisation of the mitotic spindle was hampered (Fig. 18.3) showing the decrease in birefringence of the spindle under polarised light.

Again, with the removal of colchicine or other chemicals, the spindle is again re­formed showing thereby that the microtubule organisation does not require the synthesis of new components, i.e., tubulin.

It has also been noted that microtubules remain organised when the tubulin molecules are in equilibrium with non-polymerised tubulin. So the joining and disassembly of microtubule is regulated by changes in this equilibrium.

Colchicine binds to tubulin dimers in a 1: 1 ratio in protofilaments in which the lateral tubulin-tubulin interacting sites are involved. The tubulin-colchicine complex binds only with the ends of microtubules and initiates mi­crotubule dissociation. The tubulin-colchicine complex activates the guanosine triphosphatase (GTPase) activity. This activation is responsi­ble for the conformational change in the tubulin molecule.

Microtubule assembly is also inhib­ited by the vinca alkaloids and podophyllotoxin. In presence of these alkaloids lateral assembly of tubulin molecules GTPase of the colchicine-tubulin complex are inhibited.

But the stabilisation of micro tubular structure and the extent of microtubule assembly is increased by the interaction of taxol. Taxol is produced from the bark of Taxus baccata. Taxol promotes microtubule assembly and inhibits dissociation.

The binding of taxol to the tubulin remains on the assembled microtubule but not at the end of microtubule like that of colchicine. Thus taxol interferes with the normal course of cell division by interfering in the dynamic state of microtubules, i.e., stabilisation of microtubules.

With regard to microtubule-organisation some microtubule associated proteins and Ca⁺⁺ ions play an important role besides tubulin and GTP. These proteins help in polymerisation of tubulin and other interactions with different cellular components.

The action of these proteins are again regulated by the activities of Protein kinases, the protein phosphorylation enzymes. Again, the activities of kinases depends on the presence of Ca⁺⁺ and another Ca-binding protein called calmodulin.

Of the different microtubule associated pro­teins, two groups of high molecular weight proteins are noted. One group varies from 350KD-280KD in size. The other group in­cludes 55 KD-80 KD. The first group of proteins has been identified as MAP-1 (350 KD) and MAP-2 (280KD).

The second group includes ‘T’ proteins and ‘chartins’. The association of MAP proteins with microtubules has been observed by staining with fluorescent MAP antibodies. It has been remarked by Vallee, Bloom and Theurkauf (1984) that phospho­rylation at MAP proteins may help in inter­actions with separate cytoskeletal elements in the cell.

This phosphorylation mechanism is regulated by calmodulin dependent or calmod­ulin independent mechanism. The presence of calmodulin is found in the site of microtubule disassembly as in the mitotic spindle.

Microtubule Organisation Centre (MTOC):

On the basis of the regular assembly and dis­assembly of cytoskeletal elements, it has been thought that there have some organisation cen­tres which control the assembly process. It has been found that microtubules start assembling from a distinct structure in a highly regulated pattern.

These are clearly seen in case of flagella and cilia of lower eukaryotes and during the formation of mitotic spindles from centrioles in animal cells.

From these recognisable centres, microtubule organisation starts which is known as nucleation process. Since each centre pro­duces many microtubules it has been assumed that each has many nucleation sites.

In this centre also, dense amorphous material is found which may have some role in the organisation of microtubules, although no direct relationship is found between the structure of the dense structure and the highly organised structure of the microtubule.

Flagella and cilia are organised from basal bodies. Each basal body contains nine sets of triplet tubules. These three tubules are known as A-tubule, B-tubule and C-tubule (Fig. 18.5). A-tubule has 13 protofilaments and B-tubule is horse shoe shaped structure having part of the wall of the A-tubule. C-tubule is also horse shoe shaped structure having part of the wall of the B-tubule.

The A- and B-tubules are attached with the tubules of cilia or flagella, while the C-tubules are present only in the basal body. Again, the two central tubules are present in the cilia or flagella but these are absent in the basal body.

It has been found that the organisation of cilia or flagella is controlled by centrioles present near the nucleus. Detailed mechanism for the origin of centrioles and the organisation of microtubules is not discussed here.

High voltage electron micrographic studies revealed that microtubules, microfilaments and other components form an interconnected cytoskeletal network for the cell. Besides giving architectural and mechanical support to the cell, it also helps in different metabolic processes.

One such example is in the synthesis of protein molecules that are not attached with endoplasmic reticulum. Some enzymes and substrates of the glycolysis float floated freely in the solution or loosely bound to the cytoskeletal network.

Some of the ribosomes have also been found to be attached to the cytoskeletal network in the form of multiple aggregates or polymer which are actively engaged in protein synthesis.

Any alterations in the plasma membrane can cause some response to cytoskeletal which transmits message to specific cellular target. It can also regulate the translocation of mem­brane components such as vesicles from the cell surface to different regions of the cell or to other organelles.

Thus the cytoskeleton has a regulatory control over the membrane process. It also gives rigidity to the cell in having connections with the plasma membrane.

(ii) Microfilaments:

The main component of these filaments is the protein actin, which is usually found in muscle. But these proteins have also been detected in many eukaryotic cells. Some actin-like proteins are also found in prokaryotic cells.

These are globular proteins of molecular weight of 42 KD. The different forms of actin (α, β and y) can be separated by special electrophoretic methods known as electrofocussing. The non-muscle actins differ from muscle actin by amino acid sequences.

The association of actin into cytoskeleton network has been found to be of four types:

i. Association of actin molecules into actin filaments.

ii. Association with non-actin proteins into microfilaments.

iii. Joining of microfilaments with network.

iv. Association of actin fibres with other cell components like membranes.

The role of actin as a supporting aid of various cytoplasmic structures gives an idea that the assembly of actin filaments and their associations with cell components are respon­sive to cellular controls. Cytochalasins metabo­lites from fungus Helminthosporium, have a profound effect on the actin filaments.

At low concentration of 1 x 10^-9 M, it inhibits further addition of actin molecules to microfilaments. In addition, cytochalasins induce fragmentation in actin filaments.

These effects show that there may be some proteins associated with actin in the cell. Large number of such proteins have been observed that affect the state of actin. Some proteins inhibit elongation of actin filaments, others promote disassembly and nucleation.

There are some proteins which inhibit cross-linking between actin filaments or between actin and membranes. A protein which bind to the actin monomers is known as profiling of 12-15 KD in size. It inhibits ATPase activity and the polymerisation of actin.

Again, some proteins bind to the ends of the microfilaments thus inhibiting the growth of the filaments. These are known as capping proteins Fragmin, spectrin, β-actinin, Villin etc. Most of the actin-binding proteins play an important role in the binding of microfilaments to membrane.

Detailed studies have been made in the microvilli of mammalian intestine to find out the relationship between membrane and microfilaments. Longitudinal section through microvilli shows the presence of long thread­like microfilaments in the villi.

They contain actin as evidenced by the use of actin antibodies labelled with some fluorescent markers. These actin filaments are again associated with a number of proteins—Villin, Fimbrin, Calmod­ulin etc. These proteins help in binding of microfilaments with membrane.

(iii) Intermediate Filaments:

On the basis of size, another division of cytoskeletal components has been made which has a diameter (10 nm) smaller than that of microtubules. These filaments have a cen­tral highly conserved portion of 311-324 amino acids.

This portion has 4 segments having two a-helical structures wound around each other in each segment. This central portion is flanked by end domains—N-terminal domain and C- terminal domain (Fig. 18.4).

Four segments of different size are present in central domain of the microfilaments. These are designated as 1A, IB, 2A and 2B. 1A and 2A are short— 35 residues long. 2A and 2B are large—101 and 121 residues.

These segments are linked by some linkers, such as L_x (8-14 residues), L 12 (16 or 17 residues) and L₂ (8 residues). These central segments are then coupled with end domains which are again subdivided on the basis of charge.

Examples of Intermediate Filaments are:

Keratin fibres present in epithelial cells, Desmin filaments found in muscle cells, glial filaments and neural filaments in the cells of the nervous system, vimentin filaments present in many types of cells. Intermediate filaments help in the change of cell shape.

During culture in suspension, round-off cells cease to Synthesize intermediate filaments but when the cells firmly adhere to the substratum, synthesis of intermediate filaments go on. In case of plant cells, where cell shape is controlled by cell wall, intermediate filaments are not so common.