Source: Cell and Tissue Biology: A Textbook of Histology (Leon Weiss, ed). Sixth Edition, Urban & Schwarzenberg

Chapter 1 The Cell: The Structure of the Cell (pg 18-20)

Biological Membranes

Membranes are metabolically active sheets that enclose the cell as the plasma membrane and occur within the metazoan cell as nuclear membranes, endoplasmic reticulum, Golgi membranes, and as the membranes enclosing lysosomes, pinocytotic and phagocytic vacuoles. Membranes thus bound the cell and compartmentalize its elements. Many of the functions of the cell, such as secretion of protein, synthesis of fat, drug detoxification, phagocytosis, respiration, and active transport depend on membranes.

The plasma membrane is the outer limit of the living cell and its face to the environment. It controls which substances enter the cell, providing it with selective permeability. The plasma membrane contains many and diverse molecules in its surface, which confer the capacity to interact with other cells and the extracellular environment. The fluidity of the membrane is determined by the ratio of cholesterol to phospholipid. Its permeability is also dependent on its lipid content. The membrane contains enzymatic pumps that control the levels of Na+,K+, Ca++, and other ions both in the cell and its environment. It contains the enzyme adenosine triphosphatase (ATPase), which breaks down adenosine triphosphate (ATP) to the diphosphate (ADP), thereby providing energy for active transport (pumping), endocytosis, and other energy-costing membrane functions.

Some of the molecules that extend from the surface of the plasma membrane are receptors capable of selectively linking with substances outside the cell, including receptors on other cells. Many essential cell functions are receptor-mediated, such as conduction, phagocytosis, antibody production, antigen recognition, hormone-induced activities, cell homing and cell sorting. Some receptors are shared by many cell types, such as insulin receptors needed in carbohydrate metabolism. Other receptors are quite restricted to cell type, such as the erythropoietin receptors on erythroblasts, needed to capture the hormone erythropoietin, which drives the proliferation and differentiation of red cell precursors. A cell type may show a succession of receptors as it differentiates, well exemplified by lymphocytes.

Cells contain an array of molecules on their surface, distinctive to cell type, encoded by the major histocompatibility complex (MHC) of genes, the "supergene" on chromosome 6 in human beings, that governs many cellular interactions, including immune-related actions. Cells, particularly those that are metabolically active, bristle with surface molecules, an expression of the extraordinary importance of these molecules in regulating cell function.

Plasma membranes, like other membranes, are complex and diverse. They can be isolated by cell disruption followed by differential centrifugation and studied by x-ray diffraction, freeze-fracture-etch, and microchemistry. The erythrocyte plasma membrane has been extensively studied because large amounts can be easily prepared. As is the case with most membranes, it is preponderantly protein (50-60% of dry weight), indicating high metabolic activity and structural stability. A notable exception is the myelin-rich membrane of myelinated nerves, the myelin serving as insulator.

By TEM of sectioned tissue, the plasma membrane is approximately 75 Angstroms in thickness with a range of about 60 to 90 Angstroms. As with most intracellular membranes, it is seen as a trilaminar structure, termed the unit membrane, with outer darker lines approximately 20 Angstroms wide and an inner lighter line, approximately 35 Angstroms wide (Fig. 1-12).

A valuable technique in revealing structural heterogeneity in membranes is freeze-fracture-etch. By this method membranes are typically split into outer and inner leaflets, the split tending to occur in the central lucent zone (Figs. 1-11 and 1-14).

As a result of this split there are four surfaces. The original surface facing to the exterior is the E face and the original surface facing to the interior, or protoplasm, of the cell is the P face. The fracture face on the exterior leaflet is the EF (Exterior Fracture) face while the fracture face on the interior leaflet is the PF (Protoplasmic Fracture) face. Particles may be seen on the split surfaces (Fig. l-13).

The number, size, and pattern of these particles differ from place to place in a given membrane and from membrane to membrane.

A number of models for the organization of the plasma membrane have been put forth. That of Singer and Nicolson has received wide support (Fig. 1-14).

Like other models, it postulates a lipid bilayer consisting primarily of phospholipid molecules oriented with their hydrophilic ends directed both to the outside and to the inside surfaces. Being hydrophilic, the surfaces of the membrane interact with watery environments. The phospholipid constituents of the outside and inside surfaces of the membrane, moreover, are somewhat different from one another. Within the membrane, however, lie the long-chain nonpolar hydrocarbon portions of the fatty acid constituents of the phospholipid bilayer. The internum of the membrane is, therefore, fatty and hydrophobic. Cholesterol molecules are dispersed throughout the membrane. Lying in the phospholipid bilayer like 'icebergs in a lipid sea" are the proteins. They are most likely amphipathic with their hydrophobic ends lying within the membrane among the hydrophobic fatty acids and hydrophilic pole protruding from the outside or inside hydrophilic surface of the membrane. Certain intrinsic proteins are longer than the width of the membrane and therefore cross it, protruding from both inside and outside surfaces. These proteins are presumably hydrophilic at the ends and hydrophobic in the center. There are places in membranes, such as in the synaptosome of nerve and junctional complexes, which contain concentrated protein molecules linked to one another. Proteins may be fixed in the membrane or may be rather loosely attached and move about in the plane of the membrane. Such movement can be demonstrated by staining certain receptors in the plasma membrane of mouse cells with a fluorescent marker of one color and those of human cells with a fluorescent marker of another color. The plasma membranes, and thereby the cells, are then fused by the action of sendai virus. At first the labeled receptor substances remain apart, but within 10 min. they appear completely intermixed. The mixing occurs at physiological temperature: but is inhibited at 4 degrees C. This temperature-dependence suggests simple diffusion as the basis of mixing.

In addition to proteins intrinsic or integral to the membrane, there are peripheral proteins, the extrinsic proteins, that are linked to the membrane. The contractile protein actin lies directly beneath the plasma membrane of the microvilli which is moved when actin contracts. Spectrin is a linear structural protein that forms a bridgework beneath the plasma membrane of erythrocytes and inserts into the underside of the membrane through a protein ankrins. Spectrin both strengthens the plasma membrane, protecting it against the shearing forces of the circulation, and anchors many of those intrinsic membrane proteins that extend into the subjacent cytoplasm. Spectrin is in the class of structural filaments known as intermediate filaments. Carbohydrates are regularly attached to the outside of the plasma membrane. Among them are sialic acid and other glyco- or mucoproteins. The carbohydrate-rich coat may be so heavy as to be visible as a fuzzy layer called the glycocalyx and can be selectively stained by ruthenium red or lanthanum. If sufficiently thick, it can be visible by light microscopy when stained by the periodic acid Schiff procedure. The sarcolemma of muscle and basal laminae are sites of large-scale accumulation of proteoglycans extrinsic to the plasma membrane.

INTERNATIONAL REVIEW OF CYTOLOGY, SUPPLEMENT 17 (1987, p275)

RONALD B. LUFTIG

The concept of membranes as we know it today, ‘viz.’ that all membranes have an underlying bilayer composed of phospholipids, was originally proposed by Danielli and Davson in 1935. ...

The bilayer concept as proposed by Davson and Danielli was initially based ...(in part)...on the experimental observations of Gorter and Grendel (1925) who measured extracted lipid from erythrocyte membranes and showed that enough lipid was present to construct a membrane about two lipid molecules thick. Thermodynamic constraints imposed on the constituent phospholipids to remain in a bilayer lead to the following early rules concerning their configuration: (1) polar head groups were to face out on to the aqueous phase, where protein molecules were located, and (2) hydrophobic tails were buried within the bilayer.

In addition to the classification of membrane proteins by their relative avidity to lipid in the "fluid mosaic" membrane model, there is also a hypothesized fluidity of the proteins consonant with fluidity of the underlying phospholipid bilayer.... The rates at which certain integral membrane proteins, e.g., glycophorin in the erythrocyte, move laterally within the bilayer are dependent not only on the percentage of unsaturated fatty acids, but also on the concentration of planar neutral lipids such as cholesterol. Specifically, if cholesterol is added to membranes, this tends to decrease the fluidity of proteins in the bilayer.

"UNIT MEMBRANE" STRUCTURE

The bilayer concept of membrane structure (Danielli and Davson, 1935) ... has proven over the years to be consistent with the trilaminar image of membranes which arose from electron microscopic observations (Robertson, 1957), as well as X-ray diffraction analysis of myelin (Schmitt et al., 1935: Bear et al., 1941; Finean and Burge, 1962).

It has now become clear from a large number of studies using a variety of techniques, e.g., freeze-fracture electron microscopy, thin-section electron microscopy with improved fixation, nuclear magnetic resonance, and immunological labeling of membranes with specific probes, that integral proteins can be localized within membranes. Freeze-fracture electron micrographs of all these systems show intramembrane particles of varying sizes, which presumably correspond to different integral or embedded proteins. An interesting oddity is the camel erythrocyte membrane where the protein-to-lipid ratio is very high, e.g., 3:1 (Eitan et al., 1976), and correspondingly one finds a vastly increased number of clustered intramembranous particles relative to the human erythrocyte ghost.

A trilamellar image can be observed for the ghost membranes only when they are fixed with osmium tetroxide (OsO4). This image appears to be an artifact caused by removal of membrane proteins due to OsO4, fixation, since freeze-fracture electron microscopic studies of erythrocyte ghosts, after exposure of the ghosts to varying concentrations (0.025-1.0%) of OsO4, (McMillan and Luftig, 1975; Luftig et al., 1977), show that the erythrocyte ghost intramembrane particles become increasingly soluble after such OsO4 fixation. (Figs .1 and 2).

The erythrocyte ghost intramembrane particles correlate with... the major transmembrane protein that is looped through the membrane several (3) times to form an anion channel (Steck, 1978; Jennings et al., 1984).