Structural organization and functions of the nucleolus. Cell nucleus: functions and structure Formation of the nucleolus

Nucleolus– a spherical formation (1-5 microns in diameter), present in almost all living cells of eukaryotic organisms. In the nucleus one or several usually round bodies that strongly refract light are visible - this is the nucleolus, or nucleolus. The nucleolus perceives basic dyes well and is located among the chromatin. Nucleolar basophilia is determined by the fact that the nucleoli are rich in RNA. The nucleolus, the densest structure of the nucleus, is a derivative of the chromosome, one of its loci with the highest concentration and activity of RNA synthesis in interphase. The formation of nucleoli and their number are associated with the activity and number of certain chromosome sections - nucleolar organizers, which are located mostly in the zones of secondary constrictions; it is not an independent structure or organelle. In humans, such regions are found in the 13th, 14th, 15th, 21st and 22nd pairs of chromosomes.

The function of the nucleoli is the synthesis of rRNA and the formation of ribosomal subunits.

The nucleolus is heterogeneous in its structure: in a light microscope you can see its fine-fibrous organization. An electron microscope reveals two main components: granular and fibrillar. The diameter of the granules is about 15-20 nm, the thickness of the fibrils is 6-8 nm. Granules are the maturing subunits of ribosomes.

Granular component is localized in the peripheral part of the nucleolus and is a collection of ribosomal subunits.

Fibrillar component is localized in the central part of the nucleolus and represents ribonucleoprotein strands of ribosome precursors.

The ultrastructure of nucleoli depends on the activity of RNA synthesis: at a high level of rRNA synthesis, a large number of granules are detected in the nucleolus; when synthesis stops, the number of granules decreases, and the nucleoli turn into dense fibrillar bodies of a basophilic nature.

The participation of nucleoli in the synthesis of cytoplasmic proteins can be represented as follows:

Drawing? – SCHEME OF RIBOSOME SYNTHESIS IN EUKARYOTIC CELLS

Scheme of ribosome synthesis in eukaryotic cells.
1. Synthesis of mRNA for ribosomal proteins by RNA polymerase II. 2. Export of mRNA from the nucleus. 3. Recognition of mRNA by the ribosome and 4. synthesis of ribosomal proteins. 5. Synthesis of rRNA precursor (45S - precursor) by RNA polymerase I. 6. Synthesis of 5S rRNA by RNA polymerase III. 7. Assembly of a large ribonucleoprotein particle, including a 45S precursor, ribosomal proteins imported from the cytoplasm, as well as special nucleolar proteins and RNAs that take part in the maturation of ribosomal subparticles. 8. Attachment of 5S rRNA, cutting of the precursor and separation of the small ribosomal subunit. 9. Maturation of the large subparticle, release of nucleolar proteins and RNA. 10. Release of ribosomal subparticles from the nucleus. 11. Involving them in the broadcast.

Microphotographs of the nucleolus (according to electron microscopy)

Drawing? – Electron micrograph of a nucleus with nucleolus

1- Fibrillar component; 2- granular component; 3- perinucleolar heterochromatin; 4-karyoplasm; 5-nuclear membrane.

Drawing? – RNA in the cytoplasm and nucleoli of submandibular gland cells.

Coloring according to Brush, X400

1 – cytoplasm; 2 – nucleoli. Both of these structures are rich in RNA (mainly due to rRNA - free or as part of ribosomes) and therefore, when stained according to Brachet, they turn crimson.

Nucleolus (nucleolus)- an integral part of the cell nucleus, which is an optically dense body that strongly refracts light. In modern cytology (see), the nucleolus is recognized as the site of synthesis and accumulation of all ribosomal RNA (rRNA), except for 5S-RNA (see Ribosomes).

The nucleolus was first described in 1838-1839 by M. Schleiden in plant cells and by T. Schwann in animal cells.

The number of nucleoli, their size and shape vary depending on the type of cell. The most common nucleoli are spherical in shape. The nucleoli are capable of merging with each other, so the nucleus may contain either several small nucleoli, or one large one, or several nucleoli of different sizes. In cells with low levels of protein synthesis, the nucleoli are small or not visible. Activation of protein synthesis is associated with an increase in the total volume of nucleoli. In many cases, the total volume of nucleoli also correlates with the number of chromosome sets of the cell (see Chromosome set).

The nucleolus does not have a shell and is surrounded by a layer of condensed chromatin (see) - the so-called perinucleolar, or perinucleolar, heterochromatin. Using cytochemical methods, RNA and proteins, acidic and basic, are detected in the nucleoli. Nucleolar proteins include enzymes involved in the synthesis of ribosomal RNA. When staining preparations, the nucleoli are usually stained with a basic dye. In the eggs of some worms, mollusks and arthropods there are complex nucleoli (amphinucleols), consisting of two parts, one of which is stained with a basic dye, the other (the protein body) with an acidic dye. When rRNA synthesis ceases at the beginning of mitosis (see), the nucleoli disappear (with the exception of the nucleolus of some protozoa), and when rRNA synthesis is restored in the telophase of mitosis, they form again on chromosome sections (see), called nucleolar organizers. In human cells, nucleolar organizers are localized in the region of secondary constrictions of the short arms of chromosomes 13, 14, 15, 21 and 22. During active protein synthesis by the cell, nucleolar organizers are usually reduplicated, and their number reaches several hundred copies. In animal oocytes (for example, amphibians), such copies can break off from the chromosomes and form multiple marginal nucleoli of the oocytes.

The nucleolar organizers consist of repeated blocks of transcribed DNA sequences, including the 5.8S-RNA, 28S-RNA and 18S-rRNA genes, separated by two non-coding rRNA regions. Transcribed DNA sequences alternate with non-transcribed sequences (spacers). rRNA synthesis, or transcription (see), is carried out by a special enzyme - RNA polymerase I. Initially, giant 45S-RNA molecules are synthesized; during maturation (processing), all three types of rRNA are formed from these molecules with the help of special enzymes; this process occurs in several stages. Excess 45S-RNA regions that are not part of the rRNA decay in the nucleus, and mature rRNAs are transported into the cytoplasm, where the 5.8S-rRNA and 28S-rRNA molecules, together with the 5S-rRNA molecule synthesized in the nucleus outside the nucleolus and additional proteins, form a large unit ribosomes, and the 18S-rRNA molecule is part of its small subunit. According to modern concepts, pR NKs and their precursors are present in the nucleus at all stages of processing in the form of complexes with proteins - ribonucleoproteins. The attachment of proteins to the 45 S-RNA molecule occurs as it is synthesized, so that by the time synthesis is completed, the molecule is already a ribonucleoprotein.

The ultrastructure of the nucleolus reflects the successive stages of rRNA synthesis on the templates of the nucleolar organizers. On electron diffraction patterns, a fibrillar component (nucleolonema), a granular component and an amorphous matrix are distinguished in the nucleoli (Fig.). Nucleolonema is a filamentous structure 150-200 nm thick; it consists of granules with a diameter of about 15 nm and loosely arranged fibrils with a thickness of 4-8 nm. On sections of the nucleolonema, relatively light areas are visible - the so-called fibrillar centers. It is assumed that these centers are formed by non-transcribed regions of the DNA of the nucleolar organizers, which are in complex with argentophilic proteins. Fibrillar centers are surrounded by loops of transcribed DNA chains with 45S-RNA ribonucleoproteins synthesized on them. Apparently, the latter are revealed in electron diffraction patterns in the form of fibrils.

The granular component of the nucleolus contains ribonucleoprotein granules, which are various products of rRNA processing. Among them, it is sometimes possible to distinguish dark granules of the ribonucleoprotein precursor 28S-pRNA (32S-pRNA) and lighter grains containing mature 28S-pRNA. The amorphous matrix of the nucleolus is practically no different from the nuclear sap (see Cell nucleus).

Thus, the nucleolus is a dynamic, constantly renewed structure. This is the zone of the cell nucleus where rRNA is synthesized and matured and from where it is transported into the cytoplasm.

The pathways for the release of ribonucleoproteins from the nucleolus into the cytoplasm have not been sufficiently studied. It is believed that they pass through the porosomes of the nuclear membrane (see Cell Nucleus) or through areas of its local destruction. Connections between the nucleolus and the nuclear membrane in different types of cells occur both in the form of direct contacts and through channels formed as a result of invagination of the nuclear membrane. Through similar connections, the exchange of substances between the nucleoli and the cytoplasm also occurs.

In pathological processes, various changes in the nucleoli are noted. Thus, with malignancy of cells, an increase in the number and size of nucleoli is observed, with pronounced dystrophic processes in the cell - the so-called segregation of nucleoli. During segregation, a redistribution of granular and fibrillar components occurs. With pronounced segregation of the nucleoli, the nucleolonema may disappear, and dark and light zones are formed in the granular component - the so-called caps. These structural changes reflect disturbances in the synthesis, maturation, and intranucleolar transport of rRNA.

Bibliography: Zavarzin A. A. and Kharazova A. D. Fundamentals of general cytology, p. 183, D., 1982; Chentsov Yu. S. General cytology, M., 1984; Chentsov Yu. S. and Polyakov V. Yu, Ultrastructure of the cell nucleus, p. 50, M., 1974; V o u t e i 1 1 e M. a. D i-puy-Go in A. M. 3-dimensional analysis of the interphase nucleus, Biol. Cell, v. 45, p. 455, 1982; Busch H. a. Smetana K. The nucleolus, N.Y.-L., 1970; Hadjiolov A. A. The nucleolus and ribosome biogenesis, Wien - N. Y., 1985, bibliogr.

Nucleolus (nucleolus, plasmosome)- a dense formation detected in the interphase nuclei of eukaryotic cells, which is formed at certain chromosome loci (nucleolar organizer). The nucleolus is a derivative of the chromosome, one of its loci, actively functioning in interphase. A cell usually contains 1-2 cells, sometimes more than 2. The main function of cells is the synthesis of ribosomes; it contains factors involved in the transcription of ribosomal genes, processing of pre-rRNA, and assembly of preribosomal particles. Some cellular proteins are multifunctional and participate in a number of other processes in the cell, such as apoptosis, regulation of the cell cycle, etc.

The nucleolus is a highly organized structure within the nucleus. The nucleolus contains large DNA loops containing pRNA genes, which are transcribed by RNA polymerase I at an unusually high rate. These loops are called “nucleolar organizers”.

Unlike cytoplasmic organelles, the nucleolus does not have a membrane that surrounds its contents. It appears to be formed by immature ribosomal precursors specifically bound to each other in an unknown manner. (Fig. nucleolus) The size of the nucleolus reflects the degree of its functional activity, which varies widely in different cells and can change in an individual cell.

In the nucleolus, transcription of ribosomal genes, processing of rRNA precursors, and assembly of preribosomal particles from ribosomal proteins and rRNA occur. The mechanisms of nucleolus formation are not clear. According to one hypothesis, the nucleolus is considered as a nucleoprotein complex that spontaneously appears as a result of the association of regulatory protein-nucleic acid complexes that arise on repeating rDNA sequences during their transcription. Indeed, human rRNA genes are organized into 250 tandemly repeated sequences of 44 kb in length. each, which together with the proteins associated with them form the core of the nucleolus. It is filled with other components during rRNA processing and assembly of ribosomal subunits.

Morphologically, three main zones are distinguished in the nucleolus: a fibrillar center surrounded by dense fibrillar and granular regions.

In an electron micrograph of the nucleolus, these three discrete zones can be distinguished:

1) a weakly colored component containing DNA from the region of the nucleolar organizer of the chromosome,

2) a dense fibrillar component, consisting of many thin (5 nm) ribonucleoprotein fibrils, which are RNA transcripts and

3) granular component, which includes particles with a diameter of 15 nm, representing the most mature precursors of ribosomal particles.

Using specific antibodies and hybridization probes, it was established that rRNA genes, RNA polymerase I, transcription factor UBF and topoisomerase I are localized in the fibrillar center of the nucleolus. It is believed that the fibrillar center of the nucleolus is the site of assembly of regulatory nucleoprotein complexes necessary for the transcription of rRNA genes. The dense fibrillar component surrounding the center of the nucleolus is represented by growing chains of rRNA precursors and associated proteins involved in processing. In the granular region of the nucleolus, mature 28S and 18S rRNA, partially processed RNA, as well as products of assembly of ribosomal subparticles are found. Ribosome assembly intermediates are represented by particles with a diameter of 15-20 nm. The transfer of preribosomal subparticles to the cytoplasm is apparently ensured by specific proteins that move from the nucleolus to the nuclear envelope. Due to the hierarchy in the structural and functional organization of the nucleolus in the form of separate morphologically distinguishable compartments, it is often used as a model for the functional compartmentalization of mRNA synthesis, its processing and export into the cytoplasm.

The observed “highly ordered” spatial structure of the nucleolus may simply be a consequence of the functioning of a large number of rRNA genes organized in tandem repeats, which is accompanied by the accumulation of RNA polymerase I transcripts and their processing products in the vicinity of actively working genes. The structure of the nucleolus is dynamic, and its spatial location and structural features depend on the intranuclear localization and level of activity of the corresponding rRNA genes.

Even the yeast genome contains ~200 tandemly repeated rRNA genes. However, not all genes are functionally identical: only half of the rDNA sequences are transcribed, and only ~20% of the available replication origin regions are involved in their reproduction. The transfer of genes to the rDNA region is often accompanied by their repression, which is believed to be a consequence of the functioning of the mechanism of suppression of homologous recombination in genomic regions containing tandem repeats. Mutational disruption of this mechanism is accompanied by the formation of hundreds of extrachromosomal circular rDNAs, which are unevenly distributed between daughter cells during mitosis. The accumulation of extrachromosomal rDNA by maternal cells leads to a decrease in the ability of cells to divide. This phenomenon has been called "cellular aging". In addition, the nucleolus can regulate the entry of cells into meiosis, as well as the activity of Cdc 14 phosphatase, which controls the passage of the telophase of mitosis. Evidence was obtained that the repeating rDNA sequences of the nucleolus serve as the site of assembly of the regulatory protein complex RENT (regulator of nucleolar silencing and telophase exit), which includes a phosphatase and three other proteins that provide the regulatory functions of the nucleolus.

45S rRNA transcripts first form large complexes, binding to a large number of different proteins imported from the cytoplasm, where all cellular proteins are synthesized. Most of the 70 different polypeptide chains that form the ribosome, as well as 5S rRNA, are turned on at this stage.

Other molecules are also needed for the assembly process to proceed properly. For example, the nucleolus contains other RNA-binding proteins, as well as certain small ribonucleoprotein particles (including U3-snRNP) that are thought to catalyze ribosome assembly. These components remain in the nucleolus, and the ribosomal subunits are transported in finished form into the cytoplasm. A particularly prominent component of the nucleolus is nucleolin, a well-studied protein that is present in large quantities and appears to bind only to ribosomal RNA transcripts. Nucleolin is stained with silver in a special way. This staining characterizes the entire nucleolus as a whole.

During the processing of 45S-RNA, this giant ribonucleoprotein complex gradually loses some proteins and RNA sequences and is then specifically cleaved, forming independent precursors of the large and small ribosomal subunits.

30 minutes after the introduction of a radioactive label, the first mature small ribosomal subunits containing labeled 18S rRNA leave the nucleolus and appear in the cytoplasm.

The assembly of large ribosomal subunits containing 28S-RNA, 5.8S-RNA and 5S-RNA requires slightly more time (about 1 hour), therefore, much more unfinished large subunits accumulate in the nucleolus than small ones.

The final stages of ribosome maturation occur only after the release of ribosomal subunits from the nucleus into the cytoplasm. This achieves isolation of functioning ribosomes from immature nuclear transcripts.

There is evidence indicating the participation of the nucleolus in the regulation of the cell cycle.

Nucleolus is a derivative of the chromosome, one of its loci, actively functioning in interphase. The cell nucleolus is the site of formation of ribosomal RNA and ribosomes, on which the synthesis of polypeptide chains occurs. In prokaryotic cells, the formation of ribosomes is not associated with the isolation of a special locus in the form of a nucleolus, but, despite the absence of nucleoli in these cells, the process of ribosome synthesis itself is largely similar.

The nucleoli contain several types of proteins:

• acid phosphoproteins,
• main proteins of non-histone nature.

The concentration of RNA in the nucleolus can be 2–8 times higher than in the nucleus, and 1–3 times higher than in the cytoplasm. Nucleolar RNA is a precursor of cytoplasmic RNA. Since 70 to 90% of cytoplasmic RNA is ribosomal, then The nucleolus is the site of ribosomal RNA (rRNA) synthesis.

Nucleolar RNA

At the cistron of the ribosomal gene, a giant molecule is initially synthesized - a precursor with a sedimentation coefficient of 45 S (molecular weight 4.5 106), which is then split into two parts, giving rise to 18S and 28 S rRNA. In this case, about half of the initially synthesized molecule is destroyed. Heterogeneous ribonucleoprotein particles with different sedimentation coefficients from 40 S to 80 S and higher were isolated from the nucleoli, which are ribonucleoproteins - the precursors of ribosomal subunits. Starting from 45 S RNA, the protein associates with rRNA, and first heavy ribosome precursors are formed (about 80 S and 90 S), and then ribosomal subunits (60 S and 40 S).

Nucleolar DNA

The DNA content in the isolated nucleoli is 5–12% of the dry weight and 6–17% of the total DNA of the nucleus. The DNA of the nucleolar organizer is the same DNA on which the synthesis of nucleolar, i.e., ribosomal RNA, occurs. Based on the analysis of DNA saturation during hybridization with rRNA, it is concluded that cistrons responsible for rRNA synthesis are compactly located and, possibly, presented in the form of a polycistronic region that is part of the nucleolar organizer. In the nucleolus, numerous identical genes for rRNA synthesis are localized on the DNA of the secondary constriction. Synthesis proceeds through the formation of a huge precursor and its further transformation (maturation) into shorter RNA molecules for the large and small subunits of ribosomes.

Ultrastructure of nucleoli

A fibrous or reticular structure of the nucleoli is noted, enclosed in a more or less dense diffuse mass.

Fibrous part– nucleolonema, diffuse, homogeneous part– an amorphous substance, or an amorphous part. Both of these areas of the nucleolus are negative. In some cells, individual strands of nucleolemmes merge and the nucleoli can be completely homogeneous.

Main structural components of the nucleolus:

• dense granules with a diameter of about 150 A,
• thin fibrils with a thickness of 40 - 80 A.

In many cases, the fibrillar component is assembled into a dense central zone (core), devoid of granules, and the granules occupy the peripheral zone of the nucleolus. Between the granules in this zone, loosely arranged fibrils with a thickness of 40–80 A are always observed. In some cases, no additional structuring is observed in this granular zone. But often this zone is represented isolated filamentous structures about 1500 - 2000 A thick, consisting of granules and loosely arranged fibrils. The fibrillar part of the nucleolus is not always collected into a compact central zone.

The ultrastructure of the nucleoli depends on RNA synthesis activity: at a high level of rRNA synthesis, a large number of granules are detected in the nucleolus; when synthesis stops, the number of granules drops, the nucleoli turn into dense fibrillar bodies.