Lesson 4: Cellular Nucleus PDF 2023/24

Cytology and Histology (Academic course 2023/24) Verónica Mª Molina Hernández LESSON 4: CELLULAR NUCLEUS I. INTRODUCTION The nucleus was the first intracellular structure to be discovered. Leeuwenhoek in 1700 observed a clear area in the centre of erythrocytes from salmon, which corresponded to t...

Cytology and Histology (Academic course 2023/24) Verónica Mª Molina Hernández LESSON 4: CELLULAR NUCLEUS I. INTRODUCTION The nucleus was the first intracellular structure to be discovered. Leeuwenhoek in 1700 observed a clear area in the centre of erythrocytes from salmon, which corresponded to the nucleus. But it was Robert Brown (1831) who defined it as a common component of eukaryotic cells viewing it as an opaque spot in the middle of the cell and named it the nucleus. Its study attracted the interest of numerous researchers due to several factors: its complex structure, its behaviour during cell division and its essential role in genetic transmission. It was precisely its location in the central part of the cell that gave it the name by which it is known today. Its name comes from the Latin nux (nut), because its position was reminiscent of that of a nut inside its shell. For the same reason, the Greek prefix "cario", Karyon (nut), is assigned to terms related to the kernel, core. The nucleus represents one of the main organoid of the eukaryotic cell and consists of a complex membranous structure containing the genetic information (deoxyribonucleic acid, DNA) forming part of the chromatin, which governs cell differentiation. II. NUCLEAR MORPHOLOGY Nucleus of most cells of different organs and tissues has certain morphological and functional characteristics that correspond to a morphofunctional state called the interphase nucleus. Interphase is the only cellular phase in which the nucleus appears as an easily discernible and well-demarcated compartment. As a physical entity, the nucleus is variable in shape, location, size and number in the cell. - The shape of the nucleus, in general, is adapted to the morphology of the cell in which it is found (Figure 1). Thus, for example, cubic cells normally have a rounded nucleus, while cells of endothelia or flat epithelia have a flattened nucleus and fibroblasts and muscle fibers have a spindle-shaped appearance. In contrast to this general rule, there are cells, such as neutrophils, which despite their more or less rounded shape have a nucleus with an irregular morphology. - In terms of location, the nucleus normally occupies a central position in the cell; however, this position changes depending on the functional characteristics of the cell. Thus, in secretory cells it is located in a basal position (opposite pole of the cell to the point where secretion is to take place), whereas in skeletal muscle it occupies lateral positions. - The size of the nucleus has a constant relationship with the size of the cytoplasm in each cell type, which in turn is related to its function, so that if the size of the cytoplasm increases, the nucleus will also increase. It is important to note that this 1 Cytology and Histology (Academic course 2023/24) Verónica Mª Molina Hernández relationship between the nucleus and the cytoplasm is broken in situations close to division, in which an increase in the size of the nucleus occurs without altering the size of the cytoplasm. - Finally, the number, most cells have only one nucleus, although there are exceptions to this rule. Thus, red blood cells or erythrocytes and platelets in mammals are structures that are considered cells but lack a nucleus and therefore cannot divide or carry out protein synthesis. In other cases, there are cells that contain more than one nucleus, such as hepatocytes, which can sometimes have two nuclei, or skeletal muscle cells, which are multinucleated. Figure 1. Different types of nuclei seen under the electron microscope. III. INTERFASE NUCLEUS The nucleus during interphase is surrounded by a membranous structure called nuclear envelope, which delimits a space known as nucleoplasm, karyoplasm or nuclear juice. This is nothing more than an aqueous solution composed mainly of enzymes involved in the metabolism of nucleic acids and containing two clearly identifiable substructures: chromatin and nucleolus. 1. Nuclear envelope The nuclear envelope is a boundary structure that separates the nuclear from the cytoplasmic content. Although the envelope is too thin to be seen under the light microscope, it is easily identifiable due to its intense basophilia, as it has one side with chromatin attached. The nuclear envelope is composed of a double membrane separated by a thin space called the perinuclear space. From a biochemical and ultrastructural point of view, the nuclear envelope has similar characteristics to the rough endoplasmic reticulum. In this respect, the outer membrane of the nuclear envelope has ribosomes attached to it. The relationship between the nuclear envelope and the rough endoplasmic reticulum is evident, mainly in cells after mitosis, as continuity between the two structures can be observed. The nuclear envelope consists of two membrane units, an inner nuclear membrane in contact with the nucleoplasm and composed of elastin-like proteins as well as carbohydrates, and an outer nuclear membrane, related to the cytoplasm, 2 Cytology and Histology (Academic course 2023/24) Verónica Mª Molina Hernández approximately 70-80Å thick, to which ribosomes are attached. The inner membrane is more rigid than the outer membrane, which is often scalloped. These membranes delimit a space of 150-300Å, although its width is not constant, as it depends on the functional state and the type of fixation to which the cell has been subjected, and in which we can appreciate a moderately electrodense material. In some places, this space communicates with the rough endoplasmic reticulum. Attached to the inner membrane, on the nucleoplasmic side, there is a continuous sheet of dense material about 2000Å thick, known as the dense inner lamina (fibrous lamina). This sheet has a hexagonal prism structure composed of fine filaments, which is why it has been called honeycomb layer (Figure 2). om am ps im dl pc Figure 2. Schematic diagram of the nuclear envelope and the pore complex. ps: perinuclear space; om: outer membrane; im: inner membrane; dl: dense inner layer; ca: chromatin attached; pc: pore complex. Continuity solutions are observed in both membranes, which are referred to as "pore complexes". Pore complexes interpose themselves in the nuclear envelope at more or less constant intervals, although they are destroyed and formed according to the functional needs of the cell. In a tangential section, the different structures that make up the pore complex can be seen: around both membranes of the nuclear envelope, where the pore opens, an electrodense material or annular material of a proteinaceous nature can be observed. Negative staining methods have shown that the ring or annular material actually has an octagonal shape with dimensions of about 600Å in the inner diameter and 780Å in the outer diameter, so that the pore may have different openings depending on its functionality and nuclear activity. The presence of a pore diaphragm is much debated, because while in some cells it seems to be clearly observed, in others it is completely absent. What has been demonstrated, however, is the presence of proteinaceous material towards the edges of the pore, which results in a real opening of about 500Å. Sometimes this annular protein material is so abundant that it occupies the lumen but is perforated and gives the appearance of a diaphragm (Figure 2). The nuclear envelope is not a static structure; on the contrary, its dynamism is due to its relationship with the cytoplasmic vacuolar system, to the direct correlation between the number of pores and nuclear activity, and to the relationship it often has with the nucleolus. But this functionality of the envelope is made more evident by its 3 Cytology and Histology (Academic course 2023/24) Verónica Mª Molina Hernández permeability to protein substances and ions (K+, Cl- and Na+) and the presence of enzymes (glucose-6-phosphatase, thiamine pyrophosphatase and acetylcholinesterase) in its structure. 2. Chromatin Chromatin is a complex substance consisting mainly of DNA and RNA together with proteins, histones and gene regulatory proteins. Due to its acidic nature, as it consists mainly of nucleic acids, chromatin stains with basic dyes (i.e. haematoxylin). In addition to this staining characteristic, chromatin can be identified by using special stains such as the Feulgen stain. The chromatin is partially extended and partially condensed in the interphase nucleus and therefore appears in some cases in coarse clumps and in others finely dispersed. The chromatin clumps, also called condensed chromatin or heterochromatin, are intensely basophilic and electrodense. Heterochromatin corresponds to the condensed and inactive chromatin portion and appears in most interphase nuclei, (1) either dispersed in the nucleoplasm, (2) in peripheral position forming dense clumps in close contact with the nuclear envelope or (3) associated with the nucleolus. Taking into account functional criteria, heterochromatin can also be classified into facultative heterochromatin, which is that portion of chromatin that, depending on the functional state of the cell, will be found in a condensed state or not, and constitutive heterochromatin, which is that portion of chromatin that will always be found in a condensed state. It has been shown that constitutive chromatin is not genetically inactive, as it contains euchromatic fragments intercalated from which the genes encoding rRNA (ribosomal) and tRNA (transfer RNA) are transcribed. Scattered chromatin, also called euchromatin, corresponds to extended chromatin and is related to portions of transcriptionally active DNA. The structure of euchromatin is too thin to be identified by light microscopy. Under the electron microscope, euchromatin has a less electrodense appearance than heterochromatin (Figure 3). A Figure 3. Nucleus with euchromatin (A) and heterochromatin (B). 4 B Cytology and Histology (Academic course 2023/24) Verónica Mª Molina Hernández 3. Nucleolus It is a rounded formation consisting of RNA and proteins. Due to its composition, it is a strongly basophilic structure, although it is negative with the Feulgen stain. The nucleolus consists of an electrodense material with a spongy appearance, which is not separated by a membrane. Depending on the appearance of the electrodense material that makes up the nucleolus, three parts can be distinguished in the nucleolus. The granular component (pars granulosa) occupies the largest area and is characterised by a granular appearance or morphology of an electrodense material. This part consists of ribosomal RNA (rRNA) composed of ribonucleoproteins and its function is the formation of ribosomes. The dense fibrillar component (pars fibrosa) consists of thin and tightly packed filaments corresponding to newly transcribed rRNA, and therefore not yet associated with proteins. Finally, and more difficult to identify, the fibrillar centre is described as an area consisting of a pale-staining central portion surrounded by an electrodense filamentous formation. The paler portion of the fibrillar centre are regions of activated chromatin corresponding to the nucleolar-organising region, which store the genetic information to regulate ribosome formation (Figure 4). Figure 4. Images of different nuclei - nucleoli - under the electron microscope. 5