http://www.collembola.org/publicat/integum/integum.htm Last updated on 2004.12.12 by Frans Janssens
Checklist of the Collembola: Some notes on the Ultrastructure of the Cuticula

Luc De Bruyn, Department of Biology, University of Antwerp (RUCA), Antwerp, B-2020, Belgium
Jean-Auguste Barra, Laboratoire de Zoologie, Université Louis Pasteur, Strasbourg, 67000, France
Frans Janssens, Department of Biology, University of Antwerp (RUCA), Antwerp, B-2020, Belgium

This page is under construction.

Introductory Remark

The major part of the information presented here is common knowledge compiled from the literature and from personal communications with other researchers, and it is not based on research results from our lab. The added value comes from the synergic application of concepts compiled from different disciplines.

Abstract

We propose that the formation of the surface structure of the laminate cuticula of Collembola and more in particular that the formation of the regular hexagonal pattern of the surface of the epicuticula is based on an epidermal by exocytosis regulated receptor mediated deposition of lipocuticulin micellae.

Introduction

The arthropodan integument serves many different functions (Weber, 1974:12): it gives the body its shape and size, colour and pigmentation -- its habitus; it supports movement of the body by segmentation of the body and its appendages and it offers a surface for muscle attachement, required for the body movement -- therefore also called exoskeleton; it protects the body against mechanical, chemical and physical environmental influences; it is a sensory interface with the environment; it supports oxygen transport from the environment; it secretes endproducts of the metabolism.

All different concepts of the disciplines involved are shortly introduced. Specific Collembola related issues are mentioned when applicable.

Overview

Structure of the integument

The Collembolan integument has a stratified structure. It consists of the cellular epithelium, that has an outer extracellular matrix - the cuticula - and an inner extracellular matrix - the basement membrane. In entomology the cell layer of the body wall is commonly called the "hypodermis", but the term epidermis is preferable inasmuch as the integumental epithelium is the homologue of the ectodermal layer of the skin so designated in vertebrate anatomy, though either term is illogical when applied to invertebrates having no accompanying "dermis" (Snodgrass, 1935).

The epidermis is a simple, unstratified epithelial tissue. It consists of a single layer of polyhedral cells. Depending on the life cycle phase of the animal the cells are squamous, cuboidal or columnar, with or without a microvillar apical surface.
The extracellular cuticula is differentiated from outside to inside into the epicuticula, the procuticula or exocuticula, and the subcuticula or endocuticula.

The outer surface of the cuticula is seldom smooth or bare; it presents a great variety of microscopic roughenings in the form of points, pits, ridges, and sculptured designs, and it is covered with larger outgrowths that take the shape of spicules, spines, hairs, and scales. All the external processes of the body wall, however, may be classed in two groups according to whether the epidermal cells take a direct part in their production or do not; that is, they are either cellular or noncellular outgrowths. Of the cellular processes, some are unicellular, others are multicellular (Snodgrass, 1935).

Noncellular Processes.
The noncellular projections of the outer surface of the body wall are purely epicuticular structures. They have the form of minute points or nodules (scobinations), spicules, small spines, hairs, corrugations, and ridges, the last often enclosing regular polygonal areas. The pattern of these surface characters appears in some cases to have a relation to that of the underlying epidermal cells, but in others it seems to be entirely unrelated to the cell arrangement. These surface structures of the mature body wall are probably formed over cytoplasmic processes of the epidermis when the outer layers of the cuticula are being generated, and later become solid (Snodgrass, 1935).
Multicellular Processes.
Cuticular structures of this nature are hollow outgrowths of the entire body wall and are therefore lined by a layer of formative epidermal cells. They are usually large and spine-like in form. Most of them are solidly fixed to the surrounding cuticula, but some are set in a membranous ring and are movable. The immovable varieties are specifically termed spines, the movable ones are distinguished as spurs. Both spines and spurs may themselves bear unicellular processes, or setae (Snodgrass, 1935).
Unicellular Processes.
The typical outgrowths of the body wall in this class are the hairlike processes, termed setae, that constitute the principal body covering of most insects. Some unicellular processes, however, are thick and spinous, such being distinguished as spine-like setae; others are branched or featherlike and are termed plumose hairs; still others are flat squamous structures of various shapes, known as scales. Also there are unicellular outgrowths of many other varieties having the form of cones, pegs, hooks, spatulae, knobbed hairs. etc., but all are fundamentally seta structures (Snodgrass, 1935).

The epidermis is a monolayer of matrixcells responsible for producing at least part of the basement membrane as well as all of the layers of cuticula. It is seen in its full development only when the new cuticula is being produced (Wigglesworth, 1965:25). When it is fully active the apical part of the epidermal cell is striated, with microvillar extensions into the vertical pore canals of the cuticula. In the growing stages of insects the epidermal cells are usually cubical or columnar, with the nuclei near their bases; but in adult insects, after the activity of cuticula formation is over, the matrix cells become more or less degenerate and appear in most places as a thin protoplasmic layer beneath the cuticula, in which cell boundaries are indistinct and the cell areas are marked only by the nuclei (Snodgrass, 1935).
The histological appearance of the cuticula varies somewhat in different insects and in different parts of the integument of the same insect. The endocuticula has a faint horizontally lamellate structure, in which usually there are visible fine vertical striations. The striations appear to be canals left by protoplasmic filaments that, during the formative stage of the cuticula, extend outward from the epidermal cells. The cuticular material is probably laid down in layers between these filaments, which are later retracted. N. Holmgren (1902) has suggested that the protoplasmic strands of the epidermis represent primitive cilia that once may have covered the bodies of the arthropod ancestors (Snodgrass, 1935).
The basement membrane (lamina basalis) serves as a backing for the epidermal cells and effectively separates the hemocoel (the main body cavity) from the integument.
The cuticula is formed by an apical epidermal cell secretion of water, proteins, chitin and lipids.
The cuticula differentiates in a hard exocuticula, an optional mesocuticula and a soft endocuticula. Differentiation of the exocuticula involves a chemical process (called sclerotisation) that occurs shortly after each moult. During sclerotisation, individual protein molecules are linked together by quinone compounds. These reactions "solidify" the protein matrix, creating rigid "plates" of exoskeleton known as sclerites. After moulting a hormone bursicon stimulates the epidermal cells to secrete phenolic compounds which permeate the cuticule, undergo oxidation and due to action of phenolases cross-link cuticular proteins. Quinone cross-linkages do not form in parts of the exoskeleton where resilin (an elastic protein) is present in high concentrations. These areas are membranes -- they remain soft and flexible because they never develop a well-differentiated exocuticula.

The epidermis is primarily a secretory tissue formed by a single layer of epithelial cells. The basement membrane is a supportive extracellular matrix of amorphous mucopolysaccharides, the basal lamina, enforced with an embedded reticular layer of collagen fibers. The cuticula lies immediately above the epidermis. It contains microfibers of chitin surrounded by a matrix of protein that varies in composition from species to species and even from place to place within the body of a single specimen.
Fig.3. Schematic diagram of the integument
After Meyer, J.R. 1998
As the cuticula forms, it is laid down in thin lamellae with chitin microfibers oriented at a slightly different angle in each subsequent layer. In some parts of the body, the cuticula stratifies into a hard, outer exocuticula and a soft, inner endocuticula. The epicuticula is the outermost part of the integument. The dense innermost layer of epicuticula is often missing. The subsequent layer of about 15 nanometer, called the cuticulin layer, is a stratum composed of lipoproteins and chains of fatty acids embedded in a protein-polyphenol complex. An oriented monolayer of wax molecules of about 10 to 100 nanometer lies just above the cuticulin layer; it serves as the chief barrier to movement of water into or out of the body. In many species a cement layer covers the wax and protects it from abrasion. Tiny hair-like projections or surface sculpturing of the cuticle are known as microtrichae. These acellular structures consist of a solid core of exocuticula covered by a thin layer of epicuticula.

Moulting process

The shedding of the cuticula is known as moulting, or ecdysis. Moulting affects the entire body wall and all internal parts that are formed as invaginations of it. The discarded epicuticula constitutes the exuviae (after Snodgrass, 1935).

The succession of ecdyses divides the life span of the animal into a series of stages, while the animal itself appears as a series of instars. The number of moults varies with different species and is frequently different with individuals of the same species reared under the same conditions. It is influenced somewhat by temperature, humidity, and the amount of feeding. Yet, notwithstanding all irregularities, the number of moults is surprisingly constant for each species and may be characteristic of families and even orders (Snodgrass, 1935).

The beginning of an instar is not marked by the discarding of the old cuticula, though in "life-history" studies the length of a developmental stage is usually measured from the time the exuviae are cast. Physiologically, however, it should be reckoned from the time the old cuticula is loosened from the epidermis, which more approximately marks the beginning of the short period of development that is to give the increased size and the characteristics of the following instar. The loosened cuticula may not be shed for several days. When the cuticula begins to separate from the epidermis preparatory to ecdysis, the insect usually ceases to feed and becomes more or less quiescent. Each active stage in the insect's life is thus followed by a sluggish premoulting period (after Snodgrass, 1935).

The moulting process is triggered by hormones released when a Collembola's growth reaches the physical limits of its exoskeleton. Each moult represents the end of one growth stage (instar) and the beginning of another. Moulting does not stop when the Collembola becomes an adult: about 20 moults in Pseudosinella from caves (Barra, 1991:191), about 67 moults in Hypogastrura viatica kept in experimental conditions at 20 degrees Celcius (Mertens & al., 1983).

A new, larger exoskeleton is constructed inside the old one. The moulting process begins when epidermal cells respond to hormonal changes by increasing their rate of protein synthesis. This quickly leads to apolysis -- physical separation of the epidermis from the old endocuticle. In Podura aquatica the old cuticle is lifted away from the epidermal cells by a foam-like secretion (Wiggelesworth, 1965:41). The epidermal cells initially secrete the inactive precursors of chitinase and protease, followed by cuticulin, a highly cross-linked protein that is deposited in the form of a set of layers. Cuticulin itself, formed by phenolic polymerisation of arthropodin, a mixture of soluble proteins, is resistive to enzymatic hydrolisation. The cuticulin matrix insulates and protects the epidermis from the moulting fluid's digestive action that becomes active only when the new cuticulinar multilayer is complete. The hydrolytic enzymes are too large to get through the mazes of the cuticulinar matrix. They hydrolise the proteins of the old endocuticula and the products of this digestion, amino acids and glucosamine units, that are small enough to get through the mazes of the cuticulinar matrix are recycled by the epidermis to form the new cuticula.

The separation of the old cuticula from the epidermis is accomplished by a moulting liquid formed by the epidermal cells, and/or by special exuvial glands of the epidermis, that dissolves the inner layers of the endocuticula and thus frees the rest of the cuticula from the epidermis (after Snodgrass, 1935). In Collembola, which moult during the adult stage, the exuvial glands are said to persist throughout life.
Barra, 1973:(Figure 17.C) confirmed the existance of the exuvial glands in Collembola.

The post-apolysis exuvial fluid of Collembola is characterised by the presence of proteineous polysaccharidic moulting granules (Barra, 1970:3243) and specialised haemocytes - also called ecdysohaemocytes or exuviocytes (Barra, 1991:191). The release of moulting granules into the exuvial fluid by tormogen cells is followed by the protrusion of the the exuvial lumen by thrichogen cells. Two types of ecdysohaemocytes can be distinguished: the typical granulocyte and a granulocyte with bilobed lysosomes. Apparently, the inactive moulting granules are activated by granules released by the granulocytes. The moulting granules and ecdysohaemocytes may play a role in the cuticular digestion (Barra, 1991:191).

The intermoult period can be divided - based onto the moulting granule lifecycle - into a pro-exuvial phase followed by a post-exuvial phase of equal duration (Barra, 1991:191,192(Fig.1)):
- pro-exuvial phase: after apolysis, the moulting granules released from the tormogenic vacuole into the exuvial fluid migrate towards the basis of the old endocuticula; the epicuticula is formed by the epidermal cells
- end of pro-exuvial phase: the old endocuticula and exocuticula is digested and the number and size of the moulting granules is reduced; the new exocuticula is formed by the epidermal cells; the tormogen cells form the setal base membranes
- post-exuvial phase: the microvilli of the tormogen cells retract, creating extracellular vacuoles at the setal bases
- end of post-exuvial phase: the moulting granules produced by the tormogen cells migrate into the extracellular vacuole; the new endocuticula is formed by the epidermal cells

Fig.t. Schematic dynamics of the layered deposition of the cuticula
After Barra, 1973, 1991.
Day Apolysis Pre-exuvial period Exuvation Post-exuvial period
  Moulting granules in exuvial space Deposition of epicuticula Deposition of exocuticula Ecdysis Deposition of endocuticula Secretion of moulting granules
1                          
2                          
3                          
4                          
5                          
6                          
7                          
8                          
9                          
10                          
11                          
12                          
13                          

Table I. Summary of the moulting process
Step 1:Apolysis, during which the old cuticle separates from the epidermis and the created lumen is filled with moulting gel containing moulting granules of inactive digestive enzymes
Step 2:Mitosis and cell divisions of epidermis, followed by apical secretion of a new cuticulin layer that serves as protection from digestive enzymes
Step 3:Moulting gel turns into moulting fluid, the moulting enzymes proteases and chitinases are activated and digest old endocuticle
Step 4:Epidermal cells secrete new exocuticle using "recycled" N-acetylglucosamine and aminoacids
Step 5:Moulting fluid is resorbed and a wax layer is secreted via the pore canals (??? to be verified) and spread over cuticulin rendering it impermeable to water
Step 6:Ecdysis which involves stereotyped sequence of behaviors causes splitting of the old exocuticle along ecdysial sutures and escape of the animal from it
Step 7:Expansion of new cuticle, secretion of cement layer, tanning and sclerotisation of new exocuticle
Step 8:Secretion of new endocuticle which may continue through entire intermolt period. Endocuticle often has a layered structure: helicoidal chitin fibers are deposited at night and parallel fibers are deposited during the day

After formation of the cuticulin layer, the moulting fluid becomes activated and chemically "digests" the endocuticula of the old exoskeleton. Break-down products (amino acids and saccharides) pass through the cuticulin layer where they are recycled by the epidermal cells and secreted under the cuticulin layer as new procuticula (soft and wrinkled).

Generally, it is accepted that the transcuticular pore canals transport the liquid lipids and proteins to the new epicuticula where the wax layer is formed. But this can be questioned...
Apparently, these pore canals are the remnants of the epidermis cell microvilli (see Barra, 1973:(Figure 23): an oblique transversal section through the procuticula of Tomocerus minor clearly shows the actin filaments of the microvillar cytoskeleton in the so-called pore canals). After ecdysis, the epidermal cells retract their microvilli from the newly deposited procuticula. This retraction creates a vacuum in the left behind 'empty' pores of the procuticula. Due to the suction, as a result of the vacuum, the pores are filled with the liquid wax phase of the epicuticula. This proposal assumes that there is an alternative mechanism present that deposits the wax layer on top of the epicuticula (specialised glands?).

Even without a wax layer the Collembolan cuticula is relatively impermeable (Ghiradella & Radigan, 1974:305). The wax layer is not permeable to water, but it is to oxygen or atmospheric gasses in general. The very thin (optional) polyphenolic layer between the cuticulin layer and the wax layer is not permeable to gasses.

The epicuticula's main function is to reduce water loss. When young animals of Hypogastrura viatica, cultured submerged in water, are transfered from water to air, the cuticula becomes irreversibally hydrophobe and they cannot be resubmerged. From the age of three weeks after hatching, however, the animals do not survive the transfer to air (Mertens & al., 1983:576). This suggest that the wax layer is formed by the first instars only when it is required: when cultured submerged in water, water loss is not a problem and the wax layer has no function.
The waxed epicuticula of Pogonognathellus flavescens is not wetted by water at less than 2.5 atmosphere pressure (Ghiradella & Radigan, 1974:305).

When the new exoskeleton is ready, muscular contractions and intake of air cause the body to swell until the old exoskeleton splits open along lines of weakness, ecdysial sutures. The animal sheds its old exoskeleton (ecdysis) and continues to fully expand the new one. Over the next few hours, sclerites will harden and darken as quinone cross-linkages form within the exocuticle. This process, called sclerotisation or tanning, gives the exoskeleton its final texture and appearance.
The cement (shellac) layer is secreted by dermal glands after moulting.

An animal that is actively constructing new exoskeleton is said to be in a pharate condition. During the days or weeks of this process there may be very little evidence of change. Ecdysis, however, occurs quickly (in minutes to hours). A newly moulted springtail is soft and largely unpigmented (white or ivory). It is said to be in a teneral condition until the process of tanning is completed (usually a day or two).

Synthesis and secretion of the cuticular building components

Fat synthesis
Fat is one of the major components of the cuticula and also the most variable. The fat is composed primarily of triglycerides. The other lipids are monoacylglycerides, diacylglycerides and phospholipids.
Protein synthesis
Intracellular synthesis of proteins involves transcription of DNA to mRNA and the translation of the mRNA to protein using the amino acid - tRNA complexes, and translocation of the protein to the apical membrane. This involves membrane flow to the Golgi apparatus and the processing of proteins within the Golgi apparatus. Proteins are passed across the apical membrane into the lumen via exocytosis. Most of the protein is synthesized on the rough endoplasmatic reticulum because of the association of ribosomes (mRNA containing compounds) within the endoplasmatic reticulum.
Secretion
The fat and protein droplets are secreted in an aqueous mixture. Such a mixture is called a latex dispersion. The latex dispersion is secreted principally at the moment of moulting. The rate of latex dispersion secretion and, therefore, the level of the latex production depends on the: availability of precursors in the blood, availability of reabsorbed material in the moulting fluid, rate of blood flow in the hemocoel, uptake of precursors from blood by the epidermis, the rate at which the epidermis secretory cell transforms the blood precursors and recycled moulting fluid into latex components, and discharges them into the lumen between the old and new exoskeleton.
Secretion of fat
Fat is found primarily in the form of fat droplets. Small amounts of free fatty acids may also be found.
Within the epidermis secretory cell, the smallest droplets of fat are present near the basal membrane and the largest droplets appear near the apex. Fatty acids are esterified within the endoplasmatic reticulum and contribute to the small droplets as the droplets migrate to the apex of the cell. The increase in size of droplets may also result from the aggregation of separate droplets. Near the vicinity of the apical membrane, strong attractive forces (Landon - Van-der-Waals forces) cause the droplets to be enveloped by plasma membrane. At a certain point, the droplets become completely surrounded by plasma membrane and are released into the exuvial lumen.
Secretion of protein
Proteins are synthesized by the rough endoplasmatic reticulum and pass to the Golgi apparatus. Possibly the peptide chains pass through the lumen of the rough endoplasmatic reticulum directly into the Golgi apparatus, or the pinching off of endoplasmatic reticulum may form vesicles which either migrate to and merge with the Golgi apparatus or which become a Golgi apparatus directly. The lipoprotein micellae are formed inside the endoplasmatic reticulum. These vesicles migrate to the apical membrane where it fuses to the plasma membrane. Then through the process of exocytosis, the proteins are released into the lumen between old and new exoskeleton. At this point, the vesicles become part of the plasma membrane and serve to replace, at least in part, the plasma membranes lost during the formation and secretion of fat droplets.
Secretion of water
Moulting fluid is aqueous. Because moulting fluid is isotonic with blood, water transport across the apical membrane is governed by osmotic pressure associated with solutes secreted into moulting fluid. Fat and protein molecules are too large to exert osmotic pressure. Free ions exert high osmotic pressure and largely determine the movement of water into the lumen.
Reabsorption of secretory products
The epidermis cell is like most exocrine glands capable of reabsorbing secreted products.

To be completed.

Microvilli

Microvilli are apical cell unit membrane surface modifications with main purpose to increase the surface area of the cell.
"Microvilli are membrane tubes of very constant diameter, and only relatively small variations (45 nanometer-80 nanometer) among various species of arthropods and cephalopods have been found. This constancy and accuracy in tube structure implies that the microvillus membrane is a more or less crystalline cylinder built of tightly packed membrane 'bricks'." (Hamdorf, 1979)
The microvillar cytoskeleton is formed by numerous actin filaments that are anchored into the cytoplasm of the cell. Each actin filament is made by a helical winding of two actin polymers.

Spatial microvillar pattern
Wartenberg, H.
Microvilli in general are 800 to 1000 nm in length and are packed between 10 to 15 per 1000 nm. The close sphere packing of the microvillar tubes results in cross section basically in a hexogonal pattern.

Outline 1:
1. reduction of microvilli precedes secretion
2. cells secrete monomer cuticulin by reverse pinocytosis (merocrine exocytosis) or by apocrine secretion of the microvillar apices (see 1)?
3. due to polyphenolic tanning (=formation of hydrogen cross links between protein chains) the monomers self-assemble into monolayers or spherical micellae
4. micellae self-assemble into particle films onto the microvillar apices of the epidermal cells into a pattern that is superimposed on the spatial microvillar pattern: the origin of the hexagonal epicuticular texture.
While the new exocuticula and endocuticula is formed, the microvilli themselves gradually extend, and the exocuticular and endocuticular micellae are deposited into the intermicrovillar space: from the spatial microvillar pattern also the so-called transcuticular pore canals originate.

Outline 2:
1. tightly packed, stub-shaped microvilli
2. microvillar height, array, architecture
3. inter-microvillar spacing
4. glycocalyx or 'slime layer' = membrane carbohydrates that bind excreted products of cell metabolism

To be completed.

Glycocalyx

A cellular component which strictly speaking is neither an organelle nor an inclusion, but falls into the category of plasma membrane specializations. This is the glycocalyx or cell surface coat. All cells are covered on their outside surfaces by a saccharide material they secrete, which is bound to the outer leaflet of the plasma membrane.

The surfaces of cells are covered with glycoproteins, glycolipids, and proteoglycans, forming a dense layer called the glycocalyx. This covering provides a protective barrier for the cells and serves as a supporting matrix for secretions. In addition, the individual glycoconjugates bind to growth factors, enzymes, and adhesive proteins, and thereby participate in a wide variety of biological phenomena related to cell differentiation, proliferation, tissue formation and morphogenesis. To a large extent these interactions are determined by the structure and binding properties of the polysaccharide chains (glycans) that distinguish glycoconjugates from other types of macromolecules.
The study of the structure, function, and metabolism of glycoconjugates defines the field of glycobiology. Interest in this field is rapidly expanding, since it is clear that to understand macromolecular function requires an appreciation of glycosylation.
Proteoglycans.
These molecules are common to all multicellular organisms, and are found on the surface of cells, in secretory granules and in the extracellular matrix. Most cells produce several types of proteoglycans, usually containing chains belonging to the heparan sulfate or chondroitin sulfate families of glycosaminoglycans. The glycosaminoglycans bear a strong negative charge due to hexuronic acid epimers and sulfate groups. The arrangement of sulfate groups and the epimers promote specific protein-carbohydrate interactions, which affect fundamental properties of cells, such as adhesion and endocytosis.

Apinhasnit & al. (1988) showed that the conventional method of tissue preparation for TEM did not preserve the glycocalyx in its entity. They concluded that the glycocalyx consists of two parts, an inner continuous layer which is tightly bound to the apical plasma membrane and is always preserved, and an outer filamentous layer which is not always preserved by the conventional method. The glycocalyx is mainly a negatively charged glycoprotein matrix with a substantial amount of many types of sugar residues: such as a-D-mannose, a-D-glucose, N-acetyl-glucosamine, N-acetyl-neuraminic (sialic) acid and b-D-galactose. This negatively charged glycocalyx provides a receptor site for ...

The cell coat is a layer of carbohydrates on the surface of the cell membrane. It is made up of the oligosaccharide side-chains of the glycolipid and glycoprotein components of the membrane and may include oligosaccharides secreted by the cell.

Paustian (2000) attributes the following functions to the glycocalyx: attachment to environment; reservoir for certain nutrients: the glycocalyx will bind certain ions and molecules that can then be made available to the cell; depot for excreted products: excretions will accumulate in the glycocalyx, that binds them up.
The glycocalyx serves two important purposes. First, its oligosaccharide and polysaccharide constituents absorb water to form a slimy surface. This surface protects the membrane from mechanical and chemical damage and prevents the cell from sticking nonspecifically to substrates. Second, the glycocalyx functions as the cell's uniform. Particle-cell adhesion occurs by recognition of specific carbohydrate moieties in the glycocalyx. Proteins that recognize carbohydrate sequences are called lectins.

Brown & al (1981) found three predominant glycoproteins in the rat thymocyte plasma membrane. Two of these have carbohydrate compositions that are characteristic of structures N-glycosidically linked to protein. The other glycoprotein is very different, having about 20 O-glycosidically linked carbohydrate units per 100 amino acids.

The epidermal microvilli have projections of glycoprotein molecules which are termed the glycocalyx. The sugars from heavily glycosylated membrane proteins and from glycolipids in the plasma membrane form the glycocalyx, a thick carbohydrate layer that covers the cell surface. This glycocalyx has enzymatic properties, like saccharidase, alkaline phosphatase and aminopeptidase. These glycoprotein enzymes have a hydrophobic end imbedded in the lipid layer of the cell membrane and a hydrophilic end projecting into the exuvial lumen. This hydrophilic end contains the particle binding site. The glycocalyx enables absorption of water, minerals, amino acids and simple sugars from the exuvial space. The glycocalyx enables the cell to adhere particles to its surface.
Glycocalyx formation: The cell coat is a secretion product incorporated into the cell surface that undergoes continuous renewal. Glycocalyx glycoproteins are regularly replaced via biosynthesis in the ribosomes of the rough endoplasmic reticulum, followed by final assembly with the oligosaccharide moiety in the Golgi apparatus, that subsequently packs them into membrane bound secretion granules, that get integrated in the plasma membrane.
Biological membranes are asymmetric with respect to lipid composition and transmembrane protein orientation. Lipid synthesis and ribosomal transmembrane protein synthesis takes place on the cytosolic surface of the endoplasmic reticulum. Glycosylation of lipids and transmembrane proteins occurs in the lumen of the endoplasmic reticulum and in the lumen of the Golgi complex. Mature lipids and proteins are delivered to the plasma membrane by vesicles. Through the events of vesicle extrusion and fusion, the interior surface of the endoplasmic reticulum and Golgi complex membranes are topologically equivalent to the exterior surface of the cell.
The high density of charged hydrophilic oligosacharide side-chains of glycoproteins of the glycocalyx region retains a layer of immobile water. The plasma membrane exhibits a glycocalyx, consisting of carbohydrates anchored to the membrane bilayer. The morphological features of this glycocalyx differ in function of its constituents. A dense, continuous layer of glycolipids extends 10-20 nm from the phospholipid-water interface. A more fibrous glycocalyx is made of glycoproteins and/or proteoglycans typically measuring 5-8 nm thick and 100-200 nm long consisting of 10,000 atoms or more.

Plasma membranes are mechanically supported by the cell cortex on their cytosolic surface, and are protected by the glycocalyx on their exterior surface. The sole function of glycophorin, an abundant membrane protein in erythrocytes (red blood cells), is to participate in the glycocalyx and cortex structures.

Membrane carbohydrate.
Carbohydrate makes up 2 to 10% of the plasma membrane by weight. It is exclusively on the outer surface of the cell; the fuzz of carbohydrate is called the glycocalyx. Membrane carbohydrate comes in three basic forms.
Glycolipids are lipid molecules with covalently attached sugar groups: these are found only in the outer leaflet. More than 30 different kinds of glycolipids have been identified. The most complex glycolipids are gangliosides, which contain one or more sialic acid residues. In turn, more than 30 different gangliosides are known: their functions are unknown, but one acts as a cell surface receptor.
Glycoproteins are proteins with covalently attached short chains of sugar groups, called oligosaccharides. Some of these are integral to the membrane, others are adsorbed.
A final form of membrane carbohydrate is the proteoglycans, which are adsorbed to the membrane. Proteoglycans differ from glycoproteins in being dominated by carbohydrate (they are 90-95% carbohydrate by weight), and are therefore much larger, with long, unbranched side chains, called polysaccharides. Proteoglycans were formerly called mucoproteins. The carbohydrate part of the molecule is called a glycosaminoglycan (GAG), it was formerly called a mucopolysaccharide. The GAG and the protein portions of a proteoglycan are covalently linked.
Transmembrane Proteins
Transmembrane proteins, also called integral membrane proteins. Ion channels are integral membrane proteins that can be present in as few as 50 copies in an entire cell. The combination of complexity and rarity means that transmembrane proteins are difficult to study.
Glycophorin was the first membrane protein to be sequenced. It is a glycoprotein, with 131 amino acid residues in the protein portion and 16 oligosaccharide side chains carrying almost 100 sugar residues. Each red blood cell has 600,000 glycophorin molecules.

Although most of the carbohydrate is attached to intrinsic plasma membrane molecules, the glycocalyx usually also contains both glycoproteins and proteoglycans that have been secreted into the extracellular space and then adsorbed onto the cell surface. (Alberts & al., Molecular Biology of the Cell, 3rd ed., p.502)

As the oligosaccharides and polysaccharides in the glycocalyx adsorb water, they give the cell a slimy surface. This helps as lubrication during apolysis.

Knudson (1998) found that tumor cells have the capacity, when in the presence of binding proteoglycans, to assemble components into a pericellular matrix shell or coat. Analogous to a bacterial glycocalyx, this pericellular coat may serve to cocoon the malignant cells.

To be completed.

Self-Assembly

Van der Waals forces, or dispersion forces - which are correlated dipolar charge fluctuations -, are responsible for the attraction that exists between nonpolar molecules. The presence of polar groups or permanent dipoles in a molecule can according to mutual orientation, enhance or reduce this attraction. Molecules will minimize their potential energy by nearing each other until a balance is reached between attraction and repulsion - the latter resulting most commonly from overlap of the molecular outer electron shells. As one particular molecular configuration of an ensemble of molecules will have the lowest possible energy, under ideal conditions at absolute zero, the ensemble will assume that specific configuration. This defines a van der Waals crystal (one type of crystal along with ionic, covalent, and metallic crystals), and its formation process is called crystallisation. However, when there is a multitude of forces involved, such as attraction or chemical bonding to a substrate, or the ordering of larger, macromolecular bodies, such as DNA, proteins or micellae the more complicated process that results is referred to as self-assembly, and in the case of proteins: folding.
The lowest energy configuration is not assumed by every molecule. This is because perfect conformity comes at the price of loss of entropy, or conformational freedom: there are a vast multitude of orientations possible, and only one of them is the one of lowest energy. To study the order of matter is to study the balance between these two competing factors: energy and entropy. It is a balance that is played out in every crystal, metal, liquid, gel, interface, or membrane. [after Schwartz, 1998].

To be completed.

Micellae

"Rontgenographic tests and polarized light studies, as described by W. J. Schmidt (1930), have shown that chitin has a fibrous structure, and that the fibers are composed of elongate, submicroscopic, crystalline parts (micellae) which lie parallel with the fiber axes. The chitinous mass, furthermore, is penetrated by fine intermicellar spaces and therefore possesses submicroscopic pores. This porous character, Schmidt points out, accounts for the permeability of chitin to gases and liquids, as in the chitin-covered chemoreceptive sense organs, tracheae, absorptive surfaces of the alimentary canal, and discharging surfaces of glands." (Snodgrass, 1935).

Self-assembly of amphiphilic molecules.
Membranes:
- self-assembled monolayer of amphiphilic molecules.
- self-assembled bilayer of amphiphilic molecules.
Micellae:
- micella: self-assembled structure of amphiphilic molecules in a polar liquid.
- inverted micella: self-assembled structure of amphiphilic molecules in a non-polar liquid.

Lipid-protein complexes, lipoproteins have a micellar structure, with a spherical core of triacylglycerols and cholesterol esters having a coating of phospholipids, cholesterols and apolipoproteins.

To be completed.

Discussion

To be completed.

Conclusion

To be completed.

Acknowledgements

I would like to thank
Dr Helen T. Ghiradella for her permission to use the SEMs of the Tomocerus sp. cuticle;
Dr José G. Palacios-Vargas for his permission to use the SEMs of the Sminthurides sp. cuticle;
Dr Neil Plant for his permission to use the SEMs of the Entomobrya and Tomocerus sp. cuticle;
Dr David E. Walter for his permission to use the SEM of the Megalothorax sp. cuticle;
Drs Nicole Balaguer, Helen T. Ghiradella, Johan Mertens and José G. Palacios-Vargas for sharing papers on the subject;
Dr Kenneth Christiansen and Ides Büscher for their constructive comments.

To be completed.

Glossary of terms applied to the integument

Basement Membrane (BMb).
The inner noncellular membranous lining of any epithelial layer.
Body Wall. (BW).
The integument of the body, formed of the ectoderm, consisting of epidermis, cuticula, and basement membrane.
Chitin.
The chemical substance that forms the groundwork of the cuticula, but not necessarily the principal part of it.
Cuticula (Ct).
The outer noncellular layers of the body wall.
Ecdysis.
The shedding of the cuticula. (Moulting.)
Endocuticula (Enct).
The inner softer layer of the cuticula.
Epicuticula (Epct).
The nonchitinous external filmlike covering of the exocuticula. (Grenzlamelle.)
Epidermis (Epd).
The epithelium of the body wall. (Hypodermis.)
Exocuticula (Exct).
The outer chitinous layer of the cuticula, containing the sclerotic deposits of the cuticula when the latter are present.
Exoskeleton.
Collectively the external plates of the body wall.
Exuviae.
The cuticular parts discarded at a moult.
This word in its Latin usage has no singular form; "exuvium", sometimes used, is without grammatical standing.
Hypodermis.
See epidermis.
Moulting.
The periodic process of loosening and discarding the cuticula, accompanied by the formation of a new cuticula, and often by structural changes in the body wall and other organs.
Sclerotization.
The hardening of the body wall by the deposit of sclerotizing substances in the exocuticula.

History

References