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COMMENTARY The nucleolus today

DANIELE HERNANDEZ-VERDUN

Institut Jacques Monod, 2 place Jussieu, 75251 Paris Cedex 05, France

Introduction antibodies that reduce the amount of free polymerase should also stop transcription. The nucleolus, the most prominent feature of the inter- In addition to the studies devoted to the interpretation of phase nucleus, is involved in biogenesis (Warner, nucleolar structures, others have attempted to draw up an 1990). During evolution, the nucleolus first appeared at inventory of the nucleolar , in order to investigate the time when compartmentation of the nucleolar complexity and to understand nucleolar func- cells was taking place. I would like in this review to tions. These investigations recently attracted much atten- provide information showing that the nucleolus is a tion (Sollner-Webb and Mougey, 1991; Warner, 1990) and particular nuclear territory in respect of the compartmen- in this commentary, I would like to focus particularly on tation of nuclear functions. the information available to date about nucleolar polarity, In eukaryotic cells, the nucleolus is the site at which nucleolar-specific proteins, nucleolar cycle, nucleolar ribosomal gene transcription takes place and the machin- targeting and small nucleolar . ery necessary for the production of the ribosomal subunits is assembled (Hadjiolov, 1985). At this site, the 18 S, 28 S and 5.8 S RNAs are synthesized and assembled with proteins such as ribosomal SI (Htigle et al. 1985a) Nucleolar polarity and 5 S RNA. The finding that chromosomes occupy a specific territory The nucleolus is not a stable . Its structure, in the nuclear volume, defined as the chromosome domain size and organization depend on . The (Hilliker and Appels, 1989), was recently demonstrated. different steps in this biogenesis correspond to nucleolar This demonstration was done by chromosome 'painting1, domains that can be identified by their morphology. There using specific probes corresponding to a single chromo- are three basic nucleolar domains, the fibrillar centers some, and in situ hybridization to locate specific chromo- (FCs), the dense fibrillar component (DFC) and the somes during interphase (Pinkel et al. 1989). It is therefore granular component (GC) (see nucleolar nomenclature not surprising that ribosomal gene distribution follows the reviewed by Jordan, 1984). They are found in all except a general rule of higher-order nuclear structure. However, very few nucleoli. Traditionally, the FCs are considered to the nucleolus organizer regions (NORs) are greatly be the storage sites of non-transcribed ribosomal genes, involved in nuclear polarity (Manuelidis and Borden, the DFC is the site of transcription of these genes and the 1988). Besides constituting a traditional cytological land- GC is the site of maturation and storage of the ribosomal mark, there is also evidence for polarity between different subunits (Goessens, 1984; Hernandez-Verdun, 1986; Som- NORs, and between NORs and the nuclear envelope. To merville, 1986). However, the actual sites of transcription underline the importance of nucleolar polarity, only these are still extremely controversial. Some authors believe two points will be discussed. Nevertheless, there is also they are located in the FCs (Scheer and Benavente, 1990); evidence for a specific arrangement of the in others at the border between the FCs and DFC (Derenzini Sertoli cells that reflects the activation or inactivation of et al. 1990; Thiry et al. 1991); and others again in the DFC the ribosomal genes (Haaf et al. 1990) as well as a specific only (Hartung et al. 1990; Wachtler et al. 1989). At present, association between the nucleolus and the of if we take into account all the results obtained so far, the chromosomes 1 and 9 in neurons (Manuelidis and Borden, most reasonable interpretation is to propose that the 1988). ribosomal transcription units are compacted in the DFC, as recently concluded by Jordan in an extensive review In the metaphasic plate, the NOR-bearing chromosomes (Jordan, 1991). The main results that argue for the are closer to each other than would be expected from a localization of this transcription in the FCs are based on random chromosomal distribution. This observation indi- the labelling of the FCs by anti-polymerase I antibodies cates that there is certainly a polarized arrangement of the (Scheer and Benavente, 1990). However, the presence of NOR-bearing chromosomes in the interphasic nuclei. RNA polymerase I in the FCs does not prove that they Clearly, the NOR association seems to reveal the nucleolar constitute the site of RNA transcription. The polymerases fusion that takes place in species that generally possess detected can only be the free form, and the engaged form several pairs of NORs. After this fusion, or in species cannot be accessible during transcription. As there is an without nucleolar fusion, the positions of the nucleoli equilibrium between these two forms of RNA polymerase I (Marilley and Gassend-Bonnet, 1989; Sentenac, 1985), Key words: nucleolus, interphase, nucleus. Journal of Cell Science 99, 465-471 (1991) Printed in Great Britain © The Company of Biologists Limited 1991 465 remain fairly stable, even in rotating nuclei (Bard et al. end up in the mature ribosome and their role in ribosomal 1985). The effect of the respective positions of the different biogenesis is only conjectural for the great majority. They nucleoli might explain the nucleolar dominance in have been suggested as having roles in the transcription, competition for activation molecules, as several authors maturation, packaging and transport of ribosomal par- have recently proposed (Appels, 1989; Hilliker and Appels, ticles (Reeder, 1990). 1989). As revealed by in situ hybridization of rDNA The number of proteins located in the nucleoli is very sequences, certain ribosomal sites are not associated with large. Twenty years ago, 97 different nucleolar protein a nucleolus (Manuelidis, 1985; Wachtler et al. 1986). It is spots had already been detected by two-dimensional gel therefore likely that some non-transcribed rDNA regions electrophoresis (Orrick et al. 1973). As this detection was 3 are located separately or segregated from the rest of the limited to the major nucleolar proteins below 120 x 10 MT, NORs. Thus, in non-stimulated lymphocytes, the only we may assume that several hundred proteins are remaining nucleolar structure possesses a nuclear marker specifically confined within the nucleolar territory. The of transcriptional activity (see below), although in a small notion of nucleolar territory does not exclude the presence amount, but the other NOR sites lack the special proteins of nuclear proteins, which have also been found to be associated with transcription (Manuelidis, 1985; Wachtler associated with other genes, and can accumulate or be et al. 1986). This specific distribution implies precise located in the nucleoli. The best example of accumulation targeting of the proteins associated with the NORs that that is not in itself specific for ribosomal gene function is are or have been expressed. that of DNA topoisomerase I, which is found at the site of Very frequently, in higher eukaryotic cells, the nucleoli ribosomal transcription in the nucleoli. Such accumu- are located at or near the nuclear envelope (reviewed by lation was to be expected in view of the fact that the Bourgeois and Hubert, 1988), or in nuclei they are nucleoli contain the highest concentration of highly active juxtaposed to the inner nuclear membrane (reviewed by genes in the nucleus (Reeder, 1990). There are also Clark et al. 1990). This position seems related to the numerous proteins that in the nuclei are targeted to the presence of specific skeletal structures at the site of the nucleoli (see below under Nucleolar targeting) and are not nucleolar attachment to the envelope. This nucleolar present in other nuclear territories. skeleton either adheres directly to the or is attached to it by a pedicle as visualized in spread lamina RNA polymerase I preparations (Bureau et al. 1986). At present, little This is localized in the nucleoli and is specific for ribosomal information is available about the characterization of the gene transcription of large ribosomal RNAs. This polym- nucleolar skeleton, especially its specificity compared to erase is a large molecule with complex subunit structures that of the (reviewed by Bouteille et al. composed of at least six polypeptides (Sentenac, 1985). Its 1983) and its action on the structure of the nucleolar two large components, which are highly conserved domain. A filamentous complex enriched in a protein of (Rowland and Glass, 1990), are involved in the basic 3 145 x 10 Mr, was identified as a specific nucleolar skeleton polymerization reaction, whereas the small subunita seem in amplified Xenopus laevis nucleoli (Franke et al. 1981). to have accessory roles. It has been proposed that one of 3 Moreover, the 180 x 10 MT nucleolar protein that contrib- these roles could be nuclear localization or enzyme utes to the general structure of the DFC (Schmidt- assembly (Rowland and Glass, 1990). In practice, the Zachmann et al. 1984) might be one of the nucleolar active RNA polymerase I is required for the formation of skeletal proteins playing a role in nucleolar architecture. the nucleolus as its major component, as demonstrated in In nuclei with centrally located nucleoli, there is a yeast (Hirano et al. 1989). Moreover, re-formation of the folding of the nuclear envelope called the nucleolar canal, active nucleoli was inhibited by injection of antibodies which is in direct contact with the nucleoli (Bourgeois et directed against RNA polymerase I in mitotic cells al. 1982). This canal was shown to be a nuclear envelope (Benavente et al. 1987). However, RNA polymerase I specialization that depends on the presence of active NORs activity is dependent on the presence of nucleolar (G^raud et al. 1989). We observed PtKl micronucleated transcription factors such as UBF and SL1 (Jantzen et al. cells in which the chromosomes are segregated in different 1990) and on a growth-dependent transcription initiation micronuclei. The cells were serially sectioned and three- factor (TIF-IA) (Schnapp et al. 1990). So far, nothing is dimensional reconstitutions of electron micrographs were known about the kinetics of the association of these factors made. The micronuclei possessing ribosomal genes but not with rDNA and the initial targeting of the DNA binding the other micronuclei have a nucleolar canal. Therefore, proteins. RNA polymerase I is still present in the NORs the nucleolar canal is controlled by the NOR-bearing during but there is no transcription at this stage. chromosomes, since each of these micronuclei only We therefore propose as a working hypothesis that the contains one chromosome (Ge'raud et al. 1989). starting of the rDNA transcription taking place at the It is possible that the nuclear canal, and the close beginning of is induced either by other exogen- relationship between the nucleoli and the nuclear envel- ous proteins or by modifications of the proteins already in ope, constitute nuclear envelope specializations that favor place. nuclear—cytoplasmic exchanges. Nucleolin Nucleolin, also called C23 or 100k, is a phosphorylated Nucleolar-specific proteins protein present in large amounts in nucleoli with active ribosomal biogenesis (Caizergues-Ferrer et al. 1987; A nucleolar-specific protein can be defined as a protein Lapeyre et al. 1987). This protein exists in all that is specifically located in nucleoli and is involved in (Srivastava et al. 1990) and its molecular weight varies 3 ribosomal biogenesis. A hypothesis was recently formu- between 92 and 105xl0 Mr, depending on the species. lated that such proteins may reside or be engaged in a Nucleolin comprises three distinct domains: an acidic shuttle process between the nucleoli and the amino-terminal region, four RNA-binding domains and a (Borer et al. 1989). The nucleolar-specific proteins do not glycine-rich carboxyl terminus (Caizergues-Ferrer et al.

466 D. Hernandez-Verdun 1989). It is thought to participate in the early processes of cellular proteins remain unstained (Goodpasture and ribosome biogenesis, such as the regulation of the Bloom, 1975). These proteins are also found during transcription by RNA polymerase I, the modulation of the interphase in the nucleoli and their presence is necessary conformation in the nucleolus (Erard et al. for ribosomal gene transcription (Miller et al. 1976). They 1988), and binding to nascent rRNA (Caizergues-Ferrer et have also been used as markers of 'active' NORs. In al. 1989; Srivastava et al. 1989), but its function is not electron microscopy, they have been found in the FC and clear. In the nucleoli, nucleolin is found in the DFC and DFC but never in the GC (Hernandez-Verdun et al. 1980). GC (Biggiogera et al. 1989; Escande-Ge>aud et al. 1985; On ribosomal transcriptional units spread on grids, the Noaillac-Depeyre et al. 1989). It was recently shown to Ag-NOR proteins have been localized in the transcribed shuttle between the nucleoli and the cytoplasm (Borer et part of the units and displayed a linear distribution, al. 1989). Therefore, this protein seems able to follow the indicating preferential localization on the DNP axis chain of reactions ensuring ribosome biogenesis, but is not (Angelier et al. 1982). Initially, interest in this set of part of the final product. proteins was aroused by their ability to localize particular chromosomal sites via an easy reaction. Then it was Nucleolar protein B23 reported that the variability of their staining intensity 3 revealed the degree of nucleolar activity, which was B23 (Mr, 37X10 ; pi, 5.1) (Michalik etal. 1981), also called numatrin (Zhang et al. 1989) or No38 (Schmidt-Zachmann presumably transcriptional activity. This explains the et al. 1987), is a major RNA-associated phosphoprotein recent rush on the detection of Ag-NOR proteins in that is considered to be one of the factors responsible for cells (362 papers in the last 3 years), since their amounts preribosomal particle assembly (Schmidt-Zachmann et al. might indicate the level of cell activity. However, although 1987). This protein, identified as a member of the Ag-NOR protein detection is selective, widely used and nucleoplasmin family (Schmidt-Zachmann et al. 1987), has probably a good marker of nucleolar activity, there is at been found to be widely distributed in higher eukaryotes present no clear way of identifying these proteins. On the basis of the molecular weights of the bands revealed on with the same apparent molecular weight of 3 3 gels by Ag-NOR staining (chiefly 104 and 3'7xlO Mr, and 37-38xlO Afr. It binds cooperatively, with high affinity 3 for single-stranded nucleic acids, and exhibits RNA helix- also 190, 135, 78 and 29xlO Mr) (Buys and Osinga, 1984; Lischwe et al. 1979; Pfeifle et al. 19866; Williams et al. destabilizing activity. These features may be related to its 3 1982), it has been proposed that nucleolin (100xl0 Mr) role in ribosome assembly. Protein B23 is mostly located in 3 the GC (Schmidt-Zachmann et al. 1987) and is associated and protein B23 (37xlO Afr) are the major Ag-NOR with the most mature nucleolar preribosomal RNP proteins. However, neither has been detected in FC, which is stained by silver, but both are present in the GC, (Dumbar et al. 1989). It was found to migrate out of the 3 nucleoli when RNA synthesis decreased during serum although it is never silver stained. The 190xl0 Afr band starvation (Chan et al. 1985). This protein, which appears has been proposed to be the large subunit of polymerase I. to be involved in the later stages of ribosome assembly, We have no indication of the identities of the other bands also seems, like nucleolin, to shuttle between the nucleoli (Masson et al. 1990). A yeast nucleolar protein, SSB-1, was and the cytoplasm (Borer et al. 1989). The protein recently found to be one of the silver-binding nucleolar originally described as ribocharin (Hiigle et al. 1985) proteins, and is strongly associated with snRlO, an snRNA turned out to be an isoelectric variant of B23 as mentioned involved in pre-rRNA processing in yeast (Clark et al. in the review by Warner (1990). Ribocharin was described 1990). Identification of the silver-stained proteins might as a protein involved in the transport of large ribosomal permit further characterization, either of these proteins or subunits (Hiigle et al. 19856). Therefore, protein B23 of the modifications they undergo by association with appears to be a shuttle protein playing a role in transport ribosomal transcription. from the nucleolus to the cytoplasm. Other nucleolar-specific proteins Autoimmune sera directed against the nucleoli (reviewed Fibrillarin was first described in Physarum polycephalum; by Reimer et al. 1987, and Tan, 1989) contain antibodies it is a nucleolar protein located in the DFC (Ochs et al. that recognize RNA polymerase I, fibrillarin and protein 1985). It is a basic protein (pi 8.5) with a molecular weight B23 (Kindas-Miigge, 1989), as well as many other 3 between 34 and 36xlO Mr. Autoantibodies against fibril- nucleolar proteins. Recently, these sera were systemati- larin have been found in human patients with scleroderma cally used to identify and locate new nucleolar proteins. and can also be induced in mice by mercuric chloride The finding that it was possible to conserve some of these treatment (Reuter et al. 1989). Fibrillarin has been during evolution indicates that they might participate in conserved from yeast (designated NOP1) (Henriquez et al. some fundamental function. This function may be restric- 3 1990; Schimmang et al. 1989) to the human nucleolus and ted to one nucleolar component such as the 116xlO Mr appears essential for cell growth, since the cells are not protein specific for the DFC (Masson et al. 1990). Another 3 viable in the absence of the corresponding genes (Schim- protein, the NOR-90xl0 Mr protein, seemed specific for mang et al. 1989). It became one of the 'favourite' nucleolar the NORs, even during mitosis (Rodriguez-Sanchez et al. proteins when it was demonstrated to be associated with 1987). An association with the preribosomal particle 3 U3 small nucleolar RNA (snRNA) (Lapeyre et al. 1990; fraction was proposed for the 35, 37, 69 and 92-93x 10 Mr Lischwe et al. 1985) and subsequently with U8 and U13 polypeptides recognized by four different sera (Pfeifle et al. snRNAs (Tyc and Steitz, 1989; for review, see Tollervey 1986a). Such an association was demonstrated for the 3 and Hurt, 1990). 'anti-To' antibodies, which recognize a 40xl0 Mr protein forming a complex with 7-2 nucleolar RNA (Reddy et al. 1983). In contrast, PM/Scl proteins, a complex of 11 The Ag-NOR proteins 3 3 These are a set of proteins specifically located in the NORs proteins ranging from 110xl0 Mr to 20xl0 Mr charac- during mitosis, and have been identified by their ability to teristic of polymyositis/scleroderma, failed to immuno- reduce silver under acidic conditions in which most other precipitate RNA (Reimer et al. 1987). Anti-To and PM/Scl

The nucleolus today 467 antibodies were localized by electron microscopy in the GC Cell-cycle-regulated nucleolar proteins only. Another autoantibody found in scleroderma, called The disorganization of the nucleolar territories and 3 ScBr, recognized the 94xlO Afr protein that was distrib- subsequent dispersion of the nucleolar proteins are among uted in both the DFC and the GC (Hernandez-Verdun et al. the first events in mitosis. This dispersion is not simply 1988). due to the ribosomal transcription switch-off, because cell- Although this list of nucleolar-specific proteins is not cycle-dependent modifications of the nucleolar proteins exhaustive, it indicates the power of immunological also take place at that time. It has been demonstrated that detection to characterize new nucleolar proteins and two major nucleolar proteins, nucleolin and protein B23, possibly to investigate their role. Note that there are still are highly phosphorylated during mitosis. Results indi- few monoclonal antibodies against nucleolar proteins cated that they are both substrates of p34cdc2 kinase (Peter (Kistler et al. 1984; Schmidt-Zachmann et al. 1984), except et al. 1990). In the same way as for histone HI, the for the 'favourite' nucleolar proteins (RNA polymerase I, consensus phosphorylation sequence is a repeated motif nucleolin, fibrillarin and protein B23) and the cell-cycle- (TPXKK) (Belenguer et al. 1990). This phosphorylation related nucleolar proteins (Chatterjee etal. 1987; Gerdes et has been suggested to control the mitotic changes in al. 1984; Verheijen et al. 1989; Waseem and Lane, 1990). nucleolar structures and activity. If correct, this would mean that the nucleolar dispersion would be controlled by the same mechanisms as chromosome condensation, spindle formation and nuclear envelope breakdown (Coch- The nucleolar cell cycle rane et al. 1990). Note that during interphase the activity Some nucleolar proteins appear to be cell-cycle-dependent of nucleolin also seems to be regulated by phosphorylation, but at a serine not a threonine site, and by a kinase other because they are only found in cycling cells or because cdc2 they are regulated by the cell cycle. The proteins in the than p34 (Belenguer et al. 1990). first category can be considered to act as nucleolar During mitosis, the nucleolar-specific proteins are markers of the cell cycle by their presence or level of located in different places. Some remain in association accumulation in the nucleolus. However, other aspects of with the NORs, whereas others are scattered in the the situation are not clear. For example, the presence of cytoplasm or distributed around each chromosome (Som- some of the proteins that are found in highly proliferating merville, 1986). RNA polymerase I remains bound to the cancer cells and are assumed to be proliferation-associated NORs, as well as nucleolin, Ag-NOR proteins and DNA nuclear antigens might be related to cell transformation, topoisomerase 1. This list, which is not exhaustive needed for ribosomal synthesis or for a rapid cell cycle (for (Courvalin et al. 1986), indicates that the basic com- review see Chatterjee et al. 1987). There are also similar ponents of ribosomal transcription remain together during possibilities with regard to the Ag-NOR proteins, since one mitosis, even in the absence of ribosomal transcription. group of investigators found evidence indicating that This argues in favor of fast activation of the ribosomal these proteins affect the timing of the cell cycle (Trer6 et al. genes at the end of telophase, because only the addition of 1989). one transcription factor and/or a dephosphorylation process can induce the switch-on of these genes. Nucleolar cell cycle markers The first nucleolar cell cycle marker was recognized by the Ki-67 monoclonal mouse antibody (Gerdes et al. 1984). Nucleolar targeting However, although it is very specific and routinely used, we do not yet know which epitopes are recognized. It is, The specific compartmentalization of nucleolar processes however, the standard marker for the characterization of in the nucleus demonstrates the existence of some sort of cycling cells (Verheijen et al. 1989). cellular machinery that generates this nuclear and Among these nucleolar markers, we found the nucleolar nucleolar organization. The localization of the nucleolar form of the proliferating cell nuclear antigen (PCNA) proteins in specific regions of the nucleolus indicates that (Waseem and Lane, 1990), which is different from the these proteins must contain signals that determine their nuclear form. Nuclear PCNA is an auxiliary protein of final nucleolar destination. The first amino acid sequence polymerase, and is known to be associated with DNA that targets a protein to the nucleolus, i.e. a nucleolar replication. When the nucleolar DNA replicates, that is, targeting signal (NOS), was found in the rex protein of the during a short period of the S phase, the nuclear form of human T-cell leukemia virus type 1 (HTLV1) (as reviewed PCNA is detected in the nucleoli. We do not know why a by Hatanaka, 1990). This NOS is a highly basic sequence nucleolar PCNA variant is present in the nucleoli of 19 amino acids that is able to target non-nuclear throughout the cell cycle (Waseem and Lane, 1990) but it proteins to the nucleoli. In addition, the NOS contains a does underline the particular nature of the functions in the nuclear signal that permits crossing of the nuclear nucleolus compared to the rest of the nucleus. envelope. Similar nucleolar targeting has been found in The characterization of proliferation-associated nu- other viral proteins, including the HTV-1 tat and rev cleolar proteins has been in progress for a long time in proteins (Cochrane et al. 1990; Siomi et al. 1990). Busch's group (reviewed by Busch et al. 1987). In addition Furthermore, this location in the nucleolus seems import- 3 3 ant for rev and rex functions (Cochrane et al. 1990), and to the 145xl0 Mr and 125xlO Mr proteins and the 3 some authors suggested that this specific location is 86-70xl0 Mr complex, they recently described pl20 (Jhiang et al. 1990) and p40 proteins (Chatterjee et al. involved in the export of unspliced viral mRNA (Nosaka et 1987) as nucleolar markers of cell proliferation. However, al. 1989). the function of these markers is not clear, since when the Another approach recently showed that a nuclear epitope region of pl20 was inhibited by antibody injec- localization signal, the SV40 T antigen signal, was able to tions, both DNA and RNA syntheses were inhibited bind specifically to the nucleolar protein pl40 (Meier and (Valdez et al. 1990). Blobel, 1990). This probably illustrates the potential of the

468 D. Hernandez-Verdun nucleolar-specific protein shuttle, which might be involved Although, the precise roles of these nucleolar snRNPs in in nucleus-cytoplasm exchanges. ribosome biogenesis are not yet known (Luhrmann, 1990), The nucleolar proteins tend to gather around the it is clear that their functions differ from those of ribosomal genes at the end of mitosis. As these proteins nucleoplasmic snRNPs, which are involved in the splicing are bound in a sequential order, we propose that they of the pre-mRNAs. reach their respective targets in response to a specific signal. The acidic groups of major nucleolar proteins, such as nucleolin, B23 and pl20, might be involved in this Conclusions process. Only one ribosomal gene is able to induce nucleolus formation by protein aggregation, but although There was no nucleolus before the appearance of eukary- ribosomal transcription is essential for this purpose it is otes. Its differentiation seems to be linked to the not necessarily a primary determinant of nucleolus regulation of ribosomal biogenesis, the transport of the formation (Karpen et al. 1988). To investigate the latter's products and also to other undefined functions, potential of nucleolar protein aggregation, we followed the especially in relation to the viral proteins. The hypothesis distribution and localization of nucleolar proteins in that the nucleolar territory might constitute an alterna- micronucleated cells. In the micronuclei that only con- tive route for the traffic of certain non-nucleolar molecules tained one chromosome, active ribosomal genes and is attractive and is in line with the hypothesis formulated nucleoli were only found in micronuclei containing NOR- by Harris's group (Sidebottom and Harris, 1969), who bearing chromosomes (Labidi et al. 1990). In the others, proposed that the nucleoli are involved in the transport of nucleolar proteins corresponding to the different nucleolar certain messenger RNAs. domains still accumulated, despite the absence of ribo- The nucleolus is the site of specific functions that somal genes (Hernandez-Verdun et al. 1991). These implicate specific RNA polymerase, specific snRNAa and proteins tend to aggregate in fibrillar structures, but these specific proteins. It is an organelle dependent on the cell structures are distributed at random in the nuclear cycle and the site of intense exchanges between the volume. Therefore, ribosomal gene transcription seems nucleus and cytoplasm involving specific shuttled proteins necessary in this experimental model, as it is during the and specific relationship with the nuclear envelope. In cell cycle, in order to coordinate the correct positioning of conclusion, the nucleolus constitutes a very original prepackaged nucleolar proteins. This raises the question nuclear territory that reflects the compartmentation of of what initial events are necessary for the switch-on of nuclear functions. This can in part answer the question of ribosomal transcription. why there is a nucleolus in the eukaryotes.

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