Methods in Cell Wall Cytochemistry

K V Krishnamurthy

• Publisher: CRC Press
• Year: 2020
• ISBN:
  9781000144154
• Edition: 1
• Topic: Biological Sciences
• Subtopic: Botany
• Index: Biological Sciences

Various methodologies designed to study cell walls are compiled in this book.
Methods in Cell Wall Cytochemistry covers the use of modern dyes, fluorescent chemicals, lectins, and antibody technology (immunocytochemisty.)
Cell wall morphology and chemical composition is covered as well as light and fluorescent cytochemistry; transmission electron microscopic cytochemistry; lectin cytochemistry; and, special emphasis on immunocytochemistry.
Addressing an emerging area of research and technology, this book will appeal to plant pathologists, cell biologists, as well as workers interested in stress response and those employing cell walls for biotechnological research.

Chapter Cell Wall — Morphology and Chemical Composition

1

Contents

1.1 Morphology

1.2 Chemical Composition

1.2.1 Carbohydrates

1.2.1.1 Cellulose

1.2.1.2 Callose and Other 1, 3 β-Glucans

1.2.1.3 Chitin

1.2.1.4 Pectins

1.2.1.5 Hemicelluloses

1.2.1.6 Alginic Acid

1.2.1.7 Sulphated Polysaccharides

1.2.2 Glycoproteins

1.2.2.1 Structural Proteins

1.2.2.2 Enzymes

1.2.3 Lipids and Related Substances

1.2.3.1 Cutin

1.2.3.2 Suberins

1.2.3.3 Waxes

1.2.3.4 Sporopollenin

1.2.3.5 Pollenkitt

1.2.3.6 Tryphine

1.2.4 Lignins and Other Phenolic Substances

1.2.4.1 Lignins

1.2.4.2 Phenolic Acids and Amides

1.2.5 Mineral Substances

1.2.5.1 Silica

1.2.5.2 Calcium

1.2.5.3 Boron

1.1 Morphology

The cell wall is unique to plants and envelops the protoplast. Its presence or absence serves as a criterion to find out whether a given taxon is a plant or an animal. This characteristic is more reliable than the presence or absence of chlorophyll, as plant cells can change into a heterotrophic way of life by losing chlorophyll. As Frey-Wyssling and Mühlethaler¹ had remarked, the plant cell walls historically played a special role in so far as they gave rise to the origin of the name “cell” itself. When Robert Hooke,² in 1667, discovered cells in bottle cork, he observed only cell walls which formed compartmented structures similar to the ones seen in a honeycomb. Had the cell not been discovered through its cell walls in a plant structure, but in an animal such as in a protozoan, it would scarcely have been called “cell”: the discipline “cytology” would not have been named that way either.

By virtue of its strategic position and structural organization, the cell wall or the extracellular matrix is meant to give shape or rigidity as well as protection and support to cells and tissues. It is also involved in the dimensional changes of the cells as these changes inevitably lead to changes in wall structure. The interaction between cell shape, cell growth rate, wall structure, and wall mechanics is influenced in large part by changes in biochemical processes inside the cell and vice versa. The other functions of the cell wall include control of intercellular transport, protection against other organisms, recognition and cell signalling, and storage of food reserves.³ It is at the heart of many key events in the life of a cell and the organism of which the cell is a part. In other words, “if the cell wall has something to do with the cell, it is equally true that the cell has something to do with the cell wall”.⁴

The cell wall is a layered structure formed just outside the plasma membrane. In multicellular plants, the cell walls of adjacent cells are separated, or rather cemented, by an amorphous intercellular layer called the middle lamella. Very often the cell walls are classified as primary and secondary. Such a classification is often beset with problems such as, for example, the exact definitions for primary and secondary walls. The primary wall is defined by some people on the basis of its thinness, relatively undifferentiated nature, and as the one that occurs around the protoplast of the cell at the time of cell enlargement.⁵ Walls of collenchymatous cells and of some algae defy this definition and may undergo thickness during the process of cell enlargement. Also, a distinction cannot be made with certainty in some higher plants as there are often transition lamellae. Kerr and Bailey⁶ had distinguished primary and secondary walls by the mechanism of wall growth, with primary walls showing intussusception and secondary walls apposition. Others found that primary walls showed a dispersed texture of microfibrils while secondary walls showed a parallel structure.¹ But later on it was found that these distinctions were also not tenable as both types of structures may be found in both walls. A chemical difference between the two was introduced by researchers working in higher plants. Primary walls lack lignin, cutin, suberin, and wax, whereas secondary walls contain one or more of these in addition to components of the primary wall. In algae and other thallophytes, such a distinction cannot be made and one can speak of only growing and nongrowing cell walls.⁵ However, in view of their establishment in botanical literature, the terms primary and secondary walls are regularly used in describing the cell walls of land plants.

The secondary wall of land plants is laid in certain cell types after the cessation of cell growth. The deposition often involves considerable increase in the thickness of the wall. By nature, the secondary wall does not permit the cell possessing it to enlarge further but it provides considerable strength and mechanical properties to the concerned cell. In cell wall research, it is the secondary wall that has received extensive scientific and technological attention because it is responsible for the special characteristics of raw materials, wood, textile fibers, paper fibers, cellulose, straw, and cork. In the xylary elements of gymnosperms and angiosperms, very often the secondary wall is demarcated into three layers, S₁, S₂, and S₃. ⁶ S₁ is towards the primary wall and S₃ is towards the cell lumen. Invariably S₂ is the thickest. Under the polarizing microscope, the cell wall of a cell with secondary deposition shows the following characteristics: The middle lamella is isotropic, the primary wall is weakly birefringent, S₁ and S₃ are very bright, and S₂ is dark. These differences are primarily due to the differences in the orientation of the microfibrils which make up the different layers of the cell wall. It is not necessary that all these layers should be developed by a cell. S₃ often may be absent or is replaced (often along with S₂) by a gelatinous layer as in tension wood fibers.

The three-ply structure of the secondary wall need not be present in all cell types showing secondary wall desposition. In laticifers, the secondary wall shows many submicroscopic lamellae.⁷ The same holds good for cotton hairs, where there are as many as 25 lamellae in the secondary walls. In the epidermis of many plants, the secondary wall is deposited in an altogether different fashion with a set of different chemicals. The same can be said of suberised cells of the cork.

1.2 Chemical Composition

The plant cell walls contain several groups of chemical substances. These have been variously classified. Frey-Wyssling and Mühlethaler¹ classified them on the basis of their behavior towards chemicals used in extraction analysis. They recognized the following: (1) Frame substances, which are responsible for the mechanical properties of the plant cell wall. These resist hot water and dilute alkali and acid treatments, e.g., cellulose, chitin, mannans, etc. (2) Ground or matrix substances, which are extracted by hot water and dilute alkali or acids or both. These include the pectins and hemicelluloses (such as xylans and arabinans). (3) Incrusting substances, which are substances left over when frame substances are destroyed. These include lignins, phenols, and minerals such as silica. (4) Adcrusting substances, which accumulate on the cell wall surface. These include callose, mucilaginous substances of some aquatic taxa, waxes, suberin, and cutin. A slightly modified classification, recognizing only the microfibrillar phase and matrix phase, has been followed by many.⁸ The former phase is distinguished from the latter by its high degree of crystallinity and its relatively homogenous chemical composition. Cellulose, mannans, and chitins individually make up the microfibrillar phase while all other chemicals constitute the matrix phase. Differential extraction procedures are convenient but not entirely selective in knowing about all chemical substances of the cell wall and the classification of cell wall components is best based on structure rather than on isolation procedures employed.⁹ Therefore, cell wall substances are often classified on the basis of their chemical structure into carbohydrates, proteins, phenol-derived compounds, lipid-derived compounds, minerals, etc. This classification has been followed here for the sake of simplicity.

1.2.1 Carbohydrates

Carbohyrates are polyhydroxyketones or polyhydroxyaldehydes or compounds that can be hydrolyzed to these substances using dilute mineral acids. They all share some chemical properties ...