Nucleolus

The nucleoli were first discovered by the Italian naturalist Felice Fontana in 1774. The first reliable descriptions of the nucleolus were independently performed by Rudolf Wagner (1835) and Gabriel Gustav Valentine (1836 and 1839). In 1898, published a monumental monograph on the nucleolus. His work contained 346 handwritten drawings of nuclei and nucleoli from various biological objects. In the 1930s, several researchers ( S. G. Navashin , , Barbara McClintock ) showed that the nucleoli appear on special sections of chromosomes called the nucleolar organizers ^[2] . In the 1940s, RNA was detected in the nucleoli, which explained the affinity of the nucleoli for alkaline dyes due to the acidic nature of RNA ^[3] .

For a long time, the functions of the nucleolus were not clear; up to the 1950s, it was believed that the nucleolus substance is a kind of reserve that is used and disappears during cell division ^[4] . In the 1960s, the results of a series of fundamental experiments were published that showed that the nucleoli are places of ribosome biogenesis. In 1969, Oscar Miller and Barbara Beatty using an electron microscope for the first time visualized working ribosomal genes ^{[5] [6]} . In subsequent years, the main directions in the study of nucleoli were the study of their structure, the process of assembly of ribosomes, and the determination of various structural components of ribosomes. At the turn of the century, data began to appear on new functions of the nucleolus, in no way related to ribosome biogenesis ^[7] .

The nucleolus is the most noticeable visible structure of the nucleus found in all eukaryotic organisms. For a long time it was believed that the only organism devoid of nucleoli is the diplonade Giardia lamblia , but recently a very small nucleolus has also been described in it ^[8] . The nucleolus is the most dense structure of a eukaryotic cell. The density of the nucleolus is due to the high protein content (up to 70-80% of dry weight). In addition to protein, the nucleolus contains 5-14% RNA and 2-12% DNA ^{[4] [9]} .

Using the electron microscopy method in this non-membrane organelle, three structural components were identified that correspond to different stages of ribosome biogenesis: the fibrillar center (FC), the dense fibrillar component (PFC), and the granular component (HA). Fibrillar centers are formed by fibrils with a diameter of about 5 nm ; they are partially surrounded by densely packed fibrils forming a dense fibrillar component. The granular component consists of granules with a diameter of 15-20 nm. In the nuclei of human fibroblasts , the dense fibrillar component accounts for 15% of the volume of the nucleolus, and the granular component accounts for 75%. In the nucleoli of higher plants, the proportion of PFC is significantly higher. In the nucleoli of the yeast Saccharomyces cerevisiae , only PFC and HA are found. It has been suggested that the evolution of PCs, PFKs and HAs began with a two-part system in which the components of PCs and HFCs were mixed ^[10] .

In yeast S. cerevisiae, the nucleolus intensely contacts the nuclear membrane . In higher eukaryotes, the nucleoli are located on or adjacent to the nuclear membrane. In HeLa cells, the protrusion of the nuclear membrane forms several nucleolar channels that are in direct contact with the nucleolus located in the center of the nucleus. The functions of these channels are unknown. Nuclei also interact with the nuclear lamin , and laminates are involved in the organization of chromatin. It was shown that supports the functional plasticity of the nucleoli and is involved in the structural reorganization of the nucleus and nucleoli after mitosis ^[11] .

Chromatin is found in several areas of the nucleolus. chromatin is located on the periphery of the nucleolus, as well as in the granular component, and non-condensed chromatin is in the dense fibrillar component and fibrillar centers. It is assumed that DNA filaments found in the fibrillar centers correspond to rDNA (nucleolar organizers) ^[12] . The nucleolus inserts specific proteins (such as the Nop90 scleroderma antigen in the region of the nucleolar organizers) into chromatin in the region of the nucleolus border to separate itself from the rest of the nucleus; therefore, the nucleolus is often surrounded by heterochromatin ^[13] . Chromatin domains associated with the nucleolus were found in plants ( Eng. Nucleolus-associated chromatine domains, NADs ) ^{[14] [15] [16]} .

At the periphery of the nucleolus, there is a perinucleolar compartment , a dynamic structure containing a large number of , as well as ^[17] .

Any nucleolus is formed around special DNA sequences - nucleolar organizers. The nucleolar organizers are rDNA genes assembled in rows from tandem repeats and separated by spacers . The nucleolar organizers and FCs consist of tightly associated fibrils from 6 to 10 nm thick, both contain RNA polymerase I and are characterized by a unique feature - the ability to stain with silver salts (argyrophilicity) ^[18] . Using an electron microscope, rRNA genes are seen as forming “fir-tree” type structures in which lateral curved strands are pre-rRNA transcripts, and the granules sitting in the branch are RNA polymerase I molecules ^[19] .

In humans, approximately 400 copies of repeating units of 43 kb rRNA genes (kilobases) are located on all acrocentric chromosomes (chromosomes 13 , 14 , 15 , 21 and 22 ). However, not all nucleolus DNA is represented by nucleolar organizers: for example, plants also have pseudogenes , non-coding repeating sequences , tRNA genes, and genes transcribed by RNA polymerase II ^[15] .

Fibrillar centers

Fibrillar centers are characterized by the presence of rDNA (nucleolar organizers), subunits of , I, and transcription factor UBTF . At their core, fibrillar centers are tightly packed tandem repeats of inactive rDNA and intergenic spacers. In many cell types, only some rDNA genes are transcriptionally active, despite the fact that the rest are also in the nucleolus ^[20] . RDNA transcription does not occur inside, but on the periphery of the FC. In the nucleoli of cells of different lines, a diverse number of FCs of different sizes is found, and the number of fibrillar centers is inversely dependent on their size. The nucleoli of differentiated human lymphocytes contain a single fibrillar center. If the cell cycle is activated in a lymphocyte, the formation of ribosomes starts in it, and the only fibrillar center dissolves, since the transcription of the rDNA genes contained in it begins and a dense fibrillar component is formed. Thus, the fibrillar centers contain inactive components of the rDNA transcription apparatus ^[21] .

Dense fibrillar component

The dense fibrillar component consists of fibrils of lower electron density than fibrillar centers ^[22] . In the dense fibrillar component, newly synthesized rRNA transcripts (pre-rRNA 45S ^[23] ) are detected; in addition, the early stages of rRNA processing occur in it. Here are localized proteins involved in the early stages of rRNA processing, such as fibrillarin and Nopp140, as well as ribonucleoprotein complexes containing small nucleolar RNAs (snoRNA ^[24] from the English small nucleolar ). Fibrillarin, functioning as , serves as a good marker for PFC ^[25] .

Granular component

The granular component is typically located on the periphery of the nucleolus, although in some cases the fibrillar and granular components are evenly distributed in the nucleolus. In the latter case, fibrillar-granular components often form filamentous structures — nucleonomes , or nucleolar filaments about 100-200 nm thick and distinguishable even under a light microscope (with special contrast). In nucleonomes, in addition to granules with a thickness of 15 nm, there are many thin fibrils that can form condensations ^[26] . The granules forming the granular component most likely correspond to the immature 60S ribosomal subunits. In compact nucleoli, granules are densely packed, and in branched nucleoli they form a network. In GC, and are processed, as well as the assembly of large ribosomal subunits (60S). Proteins such as nucleophosmin , , Nop52, , and the PM-Scl 100 subunit of the exosome complex ^[27] can serve as markers of HA ^.

Number and size of nucleoli

The number of nucleoli in the cell nucleus is determined by its stage of development or differentiation , and their size depends on the level of ribosome synthesis . In this case, the maximum number of nucleoli in the cell is determined by the number of nucleolar organizers, in addition, nucleoli are always larger in large polyploid nuclei. Thus, the Spur frog has two chromosomes carrying nucleolar organizers, and, therefore, usually 1-2 nucleoli ^[28] .

In dividing cells that actively synthesize ribosomes, the size of the nucleoli varies from 0.5 to 7 microns in diameter. In most cancer cells, the nucleolus is larger than in normal cells of the tissue and organ from where they originated. In the case of cells of an aggressive form of breast cancer , an increase in the size of the nucleolus by 30% is observed as the tumor develops. In differentiated cells, the formation of ribosomes decreases or ceases altogether (for example, in red blood cells and lymphocytes ), and the size of their nucleoli decreases to 0.1-0.3 microns ^[9] .

Nuclei of lower eukaryotes

The nuclei of the lower eukaryotes differ in structure from the well-studied mammalian nucleoli. For example, there are no distinguishable subcompartments in the nuclei of the mucus of , and the nucleolus is connected to the internal nuclear membrane. The assembly of ribosomes can occur throughout the nucleolus. rDNA is located not in the center of the nucleolus, as in most other organisms, but along the periphery. In addition, the rDNAs of this organism are not located on the chromosomes, but constitute a linear extrachromosomal DNA fragment, 20% of which encode rRNA ^[29] . The extrachromosomal arrangement of rRNA genes is also characteristic of such low eukaryotes as Tetrahymena pyriformis and yeast ^[30] . In the unicellular parasite Leishmania major , the do not line up in tandem rows, as in the rest of eukaryotes; instead, only 11 copies of this gene are scattered between various other genes transcribed by RNA polymerase III ^[31] . In Aspergillus mold, the nucleolus contacts the nuclear membrane, as in yeast, but has a convex shape, as in Dictyostelium ^[32] . In the crustacean Daphnia, the nucleolus is round and is located next to the nuclear membrane, but does not contact it. In the fungus, the structure of the nucleolus is similar to that in Daphnia ^[33] .

Structural Types

The severity of HA and PFC, as well as other structural features, allow us to distinguish several structural types of nucleoli: reticular (nucleonemic), compact, ring-shaped, residual (resting) and segregated ^[34] .

The reticular nucleoli are inherent in most cells, both animal and plant. Such nucleoli have a nucleolonemic structure, PFC and HA are well developed, but FCs are often poorly expressed due to active transcription ^[34] .

The compact type of the nucleolus differs from the reticular one by a lower severity of the nucleonemic structure and a higher frequency of occurrence of FC. Compact nucleoli are found in actively dividing cells, for example, plant meristem cells and tissue culture cells . Apparently, the compact and reticular types can transform into each other ^[34] .

The ring-shaped type is found in animal cells. Nuclei of this type in a light microscope look like a ring with an optically bright central zone, which is a circular center surrounded by fibrils and granules. Typical ring-shaped nucleoli are found in cells with a low level of transcription, such as lymphocytes and endotheliocytes ^[35] .

Residual nucleoli are inherent in cells that have completely lost the ability to synthesize rRNA: , differentiated enterocytes , cells of the skin epithelium and others. Often they are difficult to distinguish in a light microscope due to the small size and circumference of condensed chromatin. Sometimes they can be activated and take an active reticular or compact form ^[36] .

The segregated type of nucleoli occurs in cells in which rRNA synthesis is terminated by antibiotics , for example, actinomycin D and amphotericin , and other chemicals, or DNA and protein synthesis is damaged by mitomycin , puromycin, and many carcinogens . Different components of the nucleolus are separated from each other, but the volume of the nucleolus is progressively decreasing ^[36] .

During the cell cycle, the disassembly of the nucleoli occurs in prophase , and their assembly begins in telophase . Protein complexes and ribonucleoproteins left over from nucleoli disassembled in prophase are used to assemble nucleoli in daughter cells ^[37] .

Nucleolysis begins at an early prophase, and its final step is to stop the transcription of rDNA. Earlier, before the RNA polymerase I stopped working and the nuclear envelope was destroyed, the nucleolus was left with RNA processing proteins and snoRNP ^[24] and fixed on the surface of chromosomes, which were retained by an unknown method. The key role in the regulation of nucleolus disassembly, apparently, is played by the complex of ( English Cyclin-dependent kinase 1, CDK1 ) and ^[38] .

The assembly of the nucleoli begins with the activation of transcription in the area of ​​the nucleolar organizers. However, transcription activation alone is not enough; nucleolus assembly also depends on processing proteins and snoRNPs. In the telophase, they gather close to the chromosomes, forming point clusters, known as prenucleolar bodies. During the G1 phase of the cell cycle, the protein moves from the prenucleolar bodies to the nucleolar organizers, which results in the gradual assembly of the nucleolar compartments: fibrillar centers, dense fibrillar component and granular component ^[39] .

It has been shown that Alu- containing transcripts of RNA polymerase II, called AluRNAs, are important regulators of the assembly of nucleoli in response to cell stimuli and during the cell cycle ^[40] .

In some groups of lower eukaryotes, the behavior of the nucleoli during mitosis differs from that described above; in particular, in euglena and it can persist throughout mitosis ^[41] .

The development of methods for the isolation of nucleoli allowed for ten years ^{[ when? ]} expand the list of known nucleolar proteins from 100 to 6000. Proteomic analysis allowed the identification of more than 200 plant proteins and more than 6000 human proteins that are shared with the nucleoli. It has been shown that about 90% of nucleolus proteins in budding yeast have homologs among human nucleolar proteins. Thus, the proteome of the nucleolus remained very conservative throughout evolution . Nucleolus proteins are classified into functional groups, and only 30% of the nucleolar proteins are associated with the formation of ribosomal subunits ^[42] .

For a number of permanent nucleolar proteins, the presence of a nucleolar localization signal (NLS ) has been shown . However, many resident nucleolar proteins have no nucleolar localization signal ^[43] . It is assumed that the nucleolar localization signal is needed to retain the protein in the nucleolus rather than to deliver it ^[44] . Some sources mention nucleotide retention signals ( NoRS ), although no difference between NoLS and NoRS has been shown ^[45] . It is hypothesized that many nucleolar proteins constantly move along the nucleus and can either enter the nucleolus or leave it. However, their movement in the nucleolus slows down, possibly due to numerous interactions with other nucleolar proteins, as well as with their targets; due to the effect of slowing down, such proteins are most numerous in the nucleolus. Nucleolus nucleic acids attract structural proteins that recruit other nucleolar molecules . Among these recruiting nucleolar proteins are UBTF, fibrillarin, nucleolin and nucleophosmin. Disorders in the UBTF gene cause the formation of defective nucleoli ^[46] . Some proteins are attracted to the nucleolus only under certain circumstances, for example, in the case of DNA damage, and mitosis ^[47] .

Many proteins characteristic of the nucleolus are also localized in another nuclear body , known as the Kakhal body , so there seems to be a close relationship between these bodies ^{[48] [49]} . It has been shown that a particularly close physical relationship between Cajal bodies and nucleoli is observed under conditions of transcription suppression ^[50] . The participation of Cahal bodies in the formation of nucleoli during the development of mouse oocytes was demonstrated ^[51] .

A key function of the nucleolus is the formation of ribosome subunits in eukaryotic cells ^[20] . However, many nucleolar proteins perform completely different functions — for example, they participate in the response to cell stress ^[52] and interact with viral proteins ^[53] . In the nucleolus, also assembled ^[54] .

Ribosome Education

The formation of ribosomes begins with transcription of rDNA genes by RNA polymerase I. It is rRNA synthesis that determines the ability of a cell to grow and proliferate , and almost all cell pathways affecting them directly regulate rRNA synthesis. In mammals, rRNA gene clusters are repeating units of intergenic spacers of about 30 kb in length and pre-rRNA coding regions of about 14 kb. In higher vertebrates, the rRNA gene encodes a precursor transcript, which is co- or post-transcriptionally modified with the participation of small nucleolar RNAs, so that ultimately one molecule of 18 S, 5.8 S and 28 S rRNA is formed from it, which make up the “framework” ribosomes ^[55] . To initiate transcription mediated by RNA polymerase I, a number of specific transcription factors are needed, such as UBTF and the promoter selectivity factor, designated SL1 in humans and TIF-IB in mice. UBTF is a numerous nucleolar DNA-binding protein that activates transcription by RNA polymerase I and serves as a marker of fibrillar centers ^[56] .

As the first RNA polymerase molecule passes through one transcriptional unit of rRNA genes, the next RNA polymerase sits on the released site and synthesizes a new RNA. The final product is 45S pre-rRNA. As synthesis progresses, pre-rRNA is enveloped in ribosomal proteins that enter the nucleus from the cytoplasm . It is rRNA transcription products that formed the PFC zone around the FC. After the separation of 45S rRNA, it cleaves into smaller molecules that give rise to the 40S and 60S ribosomal subunits. Small subunits are synthesized in the nucleolus in about 30 minutes, and the synthesis of large subunits takes about an hour. The immature 60S subunit combines in the nucleolus with the third (in addition to 28S and 5.8S) rRNA molecule - 5S rRNA. Newly formed subunits exit the nucleus into the cytoplasm through the nuclear pores. The complete 80S ribosome is formed after the small subunit binds to mRNA and then to the large subunit ^[57] .

RNA polymerase I mediated transcription elongation is promoted by proteins capable of chromatin remodeling such as nucleofosmin (B23), nucleolin and . The mechanisms of termination of transcription by RNA polymerase I remained conservative during evolution. Terminator elements are recognized by DNA-binding proteins; however, they recognize specific sequences that contract with RNA polymerase I and begin transcription termination. In mice, 10 terminator elements, called Sal boxes, are clustered several hundred base pairs below the site encoding pre-rRNA and flanked by long pyrimidine tracts. A similar terminator element, designated T ₀ , is located immediately prior to the rDNA promoter. It was shown that the nucleolar protein TTF-I binds to Sal boxes and stops the elongating RNA polymerase I ^[58] . T ₀ is a necessary promoter element ^[59] . It was also shown that transcription by RNA polymerase I is promoted by nuclear forms of actin and myosin ^[60] . In addition, it is under the regulation of various growth factors ^[61] , and may also vary depending on the conditions in which the cell is located, such as nutrient supply ^[62] . It is regulated by various oncogenes and tumor suppressor genes ^[63] . The nucleolar proteins can participate in the regulation of transcription in the nucleolus by interacting with topoisomerases (for example, the nucleolar protein interacts with topoisomerase I) ^[64] .

There are many non-coding RNAs in the nucleolus called small nucleolar RNAs (snoRNAs). They are divided into several classes depending on the presence of certain conservative motives , and the most numerous are two classes containing the motives H / ACA-boxing and C / D-boxing, respectively. snoRNAs bind to various proteins and form small nucleolar ribonucleoproteins (snoRNPs), which play an important role in the processing and maturation of rRNA ^{[65] [66]} . Most snoRNPs catalyze nucleotide modifications, but some snoRNPs are involved in cutting the precursor transcript (pre-rRNA) ^[67] . snoRNPs are delivered to the nucleoli by special chaperones known as Nopp140 and treacle ^[68] .

The formation of ribosomes is one of the most energy-intensive processes occurring in a eukaryotic cell, and it is strongly associated with the cell cycle and cell proliferation. It has been shown that activation of ribosome formation causes rapid cell growth and division. Many protein factors that regulate the formation of ribosomes are also directly involved in some stages of the cell cycle in both yeast and mammals. Mechanisms that control the biosynthesis of ribosomes also work during the G1 phase of the cell cycle and communicate with proteins that make the transition from the G1 phase to the S phase , as a result, the cell may enter division or not divide depending on the intensity of the process of formation of ribosomes ^[69] .

Stress Response

It was shown that the nucleolus plays a key role in the regulation of the p53 –Mdm2 loop. p53 and are mainly nucleoplasmic proteins, with p53 being the most important protein responsible for the cell’s response to stress (DNA damage, activation of the oncogen , abnormalities in the ribosomes), and Mdm2 serves as its negative regulator. Tumor suppressor protein is a nucleolar protein that inhibits the functioning of Mdm2, inhibiting its activity as an E3 ubiquitin ligase or isolating it in the nucleolus, as a result of which p53 protein is stabilized and activated. In addition, ARF is a key regulator of cell aging ^[70] . Another nucleolar protein known as nucleophosmin (B23) is involved in response to cellular stress. It can isolate ARF in the nucleolus and, depending on the situation, B23 acts as an oncogen or tumor suppressor gene. In addition, nucleolin and B23 can be involved in the repair of damaged DNA ^[71] . p53 can also cause a cellular response in the form of a cell cycle arrest in response to disturbances in ribosome biogenesis in the nucleolus ^[72] .

The nucleolus may be involved in a response to stress of a different kind. For example, in conditions of hypotension, the nucleolus turns into numerous small nucleoli, which, when the cells are transferred to normal conditions, merge with each other, forming nucleoli. Interestingly, one of the most important nucleolar proteins, nucleophosmin, does not accumulate in the nuclei, but circulates between the nucleoli and the nucleoplasm ^[73] .

Signal recognition particle assembly

Signal recognition particles (SRPs ) are ubiquitous cytoplasmic ribonucleoprotein complexes that deliver some ribosomes to the rough endoplasmic reticulum (EPR) for further cotranslational translocation into the EPR of synthesized membrane and secreted proteins. First, the SRP recognizes the signal peptide of the growing secreted or membrane channel as it leaves the ribosome. Further, SRP temporarily stops protein synthesis and delivers the protein synthesized ribosome to the cytoplasmic side of the EPR, and further protein synthesis occurs simultaneously with its translocation into the EPR ^[74] . When the RNA, which is part of SRP, was introduced into the nucleus of a mammalian cell, it very quickly appeared in the nucleolus. After some time, the level of fluorescence in the nucleolus fell, but increased in some places of the cytoplasm ^[75] . Localization of SRP RNA cannot be linked to one of the three nucleolus domains: the localization region passed through the entire nucleolus ^[76] . It has been shown that in the nucleolus, the final stages occur with the interest of SRP RNA and assembly of SRP itself ^[77] .

Other features

For the work of activated macrophages, cathepsin cysteine proteases play an important role. In endosomes and lysosomes, they play a crucial role in the formation of an acquired immune response (antigen processing and presentation ), as well as an innate immune response (activation of Toll-like receptors ). It has recently been shown that these cysteine ​​proteases and their inhibitors also perform certain functions in the nucleus and nucleolus. For example, upon macrophage activation, and the Spia3g inhibitor are localized in the nucleolus ^[78] .

It has been shown that some nucleolar proteins in plants can be involved in maintaining the nucleolar chromatin and telomere structure ^[15] .

In yeast, mRNAs are sent to the nucleoli if nucleocytoplasmic transport, rRNA biogenesis, or mRNA processing are impaired ^[79] .

In an inactive form, when the transcription of rRNA genes is reduced, the nucleolar organizer is represented by one large fibrillar center. Ribosomal DNA at this moment is condensed (i.e. compactly laid). When nucleolus activation begins, rDNA decondensation occurs, and it begins at the periphery of the fibrillar center. As a result, RNP transcripts are formed, which, when ripe, form granules - the precursors of ribosomes, which occupy the periphery of the activated nucleolus. As transcription is further enhanced, a single fibrillar center breaks up into a series of smaller ones that are connected to each other by completely decompacted (i.e., unfolded) rDNA regions. The more intense the transcription, the more small FCs in the nucleolus are connected with each other and surrounded by PFCs. If rDNA activation is complete, then all the FCs are decondensed, and the active rDNA is in PFC. When the nucleolus is inactivated, the FCs form again and merge with each other, they increase in size, and the proportion of PFC decreases. When the nucleolus is completely inactivated, it is represented by only one large FC (up to 4-5 microns) of a spherical shape, surrounded by a layer of condensed chromatin. The inactivated nucleolus is structurally close to the nucleolar organizers of mitotic chromosomes. Such transformations are called activation and deactivation of the nucleolus, respectively ^[80] .

Although there are many copies of ribosomal DNA genes in cells, not all of them are used to form rRNA. It was shown that active and silent rDNA genes are characterized by various epigenetic labels. So, in silent rDNA genes, methylation was detected, which is characteristic of heterochromatin and transcriptionally inactive genes, while active genes do not have such a label. RDNA hypomethylation has also been observed in some types of cancer, in particular lung cancer and hepatocellular carcinoma . However, there is evidence of a positive role for CpG methylation in the synthesis and processing of rRNA ^[81] .

It was shown that silencing patterns of rRNA genes caused by DNA methylation are transmitted from the original cell to daughter cells during cell division ^[82] . Silence of rDNA genes can be associated not only with DNA modification, but also with special marks on histones . It has been established that active rDNA genes acquire heterochromatin labels in response to changes in cell energy status and differentiation, and these labels may not be related to CpG methylation ^[83] . A number of silent rDNA genes are located in the extra-nucleolar space and are often associated with perinucleolar heterochromatin (while active rDNA genes are located inside the nucleolus in the fibrillar component), for example, centromeric heterochromatin. The silent status and heterochromatin state of these genes are believed to be associated with limited access to the nucleolus of recombination proteins. They can also contribute to the structure of the nucleolus and nucleus ^[84] . Finally, there is evidence that in the cells of the female body, the nucleolus is associated with an inactivated X chromosome ^[85] .

Mature mammalian oocytes, as well as blastomeres of very early stages of crushing of the mammalian embryo, contain inactive atypical nucleoli, which differ significantly from the nucleoli of mature cells and contain only a dense fibrillar component ^[86] . They are called nucleolar precursor bodies (NPBs ), and it is believed that they serve as storage sites for molecules, from which, as the nucleus develops, material is taken to assemble typical active nucleoli. The number and distribution of Taurus inclusions in pronuclei of a human embryo can serve as a simple non-invasive indicator of further embryonic development ^[87] . When oocytes enter meiosis, the dense fibrillar component of the nucleoli disintegrates and enters the cytoplasm, and the nucleolus becomes invisible until fertilization and formation of pronuclei ^[86] .

In 2003, it was shown that precursor bodies could be removed from a fully matured pig oocyte using microsurgery methods. This method of removing the nucleolus is called enucleolation , and the extracted nucleolus is called nucleoloplast . In this case, the bodies are secreted with a certain amount of oocyte cytoplasm covered with a . Oocytes survive this operation and can achieve metaphase II division; thus, the nucleolus does not play a significant role in the maturation of mammalian oocytes. This seems unusual because in yeast cells and somatic cells, the nucleolus plays an important role in the transition from metaphase to anaphase . Nevertheless, it was shown that the nucleolus is nevertheless necessary for the regulation of the onset of maturation of mammalian oocytes ^[88] . The possibility of nucleolus transplantation in mammalian oocytes has been shown ^[89] .

The nucleolus is involved in the development of many infectious and non-infectious diseases in humans. The role of nucleolus in the development of various groups of diseases is discussed below.

Viral diseases

Viruses from various groups ( DNA-containing , , retroviruses ) encode proteins that are localized in the nucleolus during infection. Such viruses include, for example, herpes simplex virus ^[90] , cytomegaloviruses ^[91] , flaviviruses ^[92] , ^[93] and HIV ^[94] . Some of these viruses, such as poxviruses , replicate in the cytoplasm, and herpes viruses and adenoviruses replicate in the nucleolus. The initial stages of HIV-1 replication proceed in the cytoplasm, and its further activity occurs in the nucleolus ^[53] . Some subviral agents can also be localized in the nucleolus, for example, hepatitis delta virus , satellites and viroids ^[95] . Viruses can interact with the nucleolus to take control of the functioning of the host cell and attract nucleolar proteins to facilitate their own replication and regulation of viral gene expression ^[96] . For example, in the nucleolus, two regulatory proteins of HIV gene expression are localized - and ^[97] . Studying the ways in which viruses interact with the nucleolus may help develop new antiviral therapy ^[98] .

Ribosomopathies

Several hereditary human diseases - such as , , Tricher-Collins and — arise from disturbances in the protein factors necessary for the formation of ribosomes. These diseases are characterized by noticeable pleiotropy , manifest themselves in disruption of the work of many types of tissues, and the severity and list of symptoms are different in different patients. These diseases are characterized by the following symptoms in various combinations: hematopoiesis disorders, developmental abnormalities and cancer susceptibility. Such diseases, which are based on changes in genes encoding the constituents of mature ribosomes and / or factors involved in the formation of ribosomes, are called ^[99] .

Cancer

Since the rate of ribosome synthesis determines the ability of cells to proliferate, abnormalities in the synthesis of ribosomes in the nucleolus often lead to the development of cancerous tumors. For example, abnormalities in the number, size, and morphology of nucleoli are often associated with the onset and further development of breast cancer ^[100] . The marker of breast cancer is argyrophilic (ie, stained with silver salts) nucleolar organizers ^[101] ; in addition, the nucleolus can modulate the work of the estrogen receptor , which plays a key role in the development of this type of cancer ^[102] . However, the nucleolus may also have an inhibitory effect on the tumor; the nucleolus contains such tumor suppressor genes as BRCA1 , ARF, p53, retinoblastoma protein (RB) and PTEN , as well as HOTS ^[103] . The table below lists the nucleolar proteins involved in the development of various cancerous tumors ^[104] .

Other diseases

According to one hypothesis, the nucleolus is involved in the development of autoimmune diseases ^[13] . It is also possible that this organelle may be involved in the development of Parkinson's disease ^[105] . Tau protein , which has recently been associated with the development of Alzheimer's disease , can also be localized here ^[106] . The nucleolus can also play an important role in the development of the eye and neurodegenerative diseases of the retina ^[107] . The nucleolar protein nucleostemin may serve as a marker of osteoarthritis ^[108] .

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