Localization of chromatin in "persistent" nucleoli in okadaic acid treated HeLa cells.

In okadaic acid treated HeLa cells, the chromosomes sometimes condense without being accompanied by nuclear envelope breakdown. These cells show "persistent" nucleoli. Within these "persistent" nucleoli the intranucleolar chromatin condenses and can be observed in the region of the dense nucleolar component (DNC) of the nucleoli. Other nucleolar components, namely the fibrillar centre (FC) and the granular component (GC) remain unchanged. These observations strongly speak for the localization of nucleolar chromatin (ribosomal cistrons) within the dense nucleolar component of the interphase nucleolus.     

Effects of 2-mercapto-1-(beta-4-pyridethyl)-benzimidazole (MPB) on Ehrlich cell nucleoli: stereological analysis

The effects of MPB, a strong inhibitor of RNA synthesis, have been analysed at the ultrastructural level by means of stereological methods. After treatment, an increase in the nucleolar volume is observed. This enlargement is due to vacuolization of the nucleoli. However, the relative volumes of the nucleolar components are modified in various directions: the volume of the granular component decreases whereas the fibrillar centres increase in size. These results are discussed in terms of relations between morphology and function of the nucleolus.     

ULTRASTRUCTURE OF THE NUCLEOLUS DURING THE CHINESE HAMSTER CELL CYCLE

Changes in the structure of the nucleolus during the cell cycle of the Chinese hamster cell in vitro were studied. Quantitative electron microscopic techniques were used to establish the size and volume changes in nucleolar structures. In mitosis, nucleolar remnants, "persistent nucleoli," consisting predominantly of ribosome-like granular material, and a granular coating on the chromosomes were observed. Persistent nucleoli were also observed in some daughter nuclei as they were leaving telophase and entering G₁. During very early G₁, a dense, fibrous material characteristic of interphase nucleoli was noted in the nucleoplasm of the cells. As the cells progressed through G₁, a granular component appeared which was intimately associated with the fibrous material. By the middle of G₁, complete, mature nucleoli were present. The nucleolar volume enlarged by a factor of two from the beginning of G₁ to the middle of S primarily due to the accumulation of the granular component. During the G₂ period, there was a dissolution or breakdown of the nucleolus prior to the entry of the cells into mitosis. Correlations between the quantitative aspects of this study and biochemical and cytochemical data available in the literature suggest the following: nucleolar reformation following division results from the activation of the nucleolar organizer regions which transcribe for RNA first appearing in association with protein as a fibrous component (45S RNA) and then later as a granular component (28S and 32S RNA).     

Relative position of nucleolar chromatin and nucleolar components in ciliate <Emphasis Type="Italic">Didinium nasutum</Emphasis> somatic nuclei

According to our computer modeling data obtained earlier, nucleoli in interphase ciliates Didinium nasutum are complex netlike structures, in which the trabeculumor lamella-shaped fibrillar component is located on the periphery, and the granular component in the central part of the nucleolus. Chromatin bodies connected with nucleoli act as the nucleolar organizers in D. nasutum. In the present work, the arrangement of all chromatin bodies, which could correspond to nucleolar organizers by morphological criteria, is studied by means of a 3D-reconstruction. It is shown that all of these chromatin bodies are localized outside the nucleoli, on the fibrillar component’s periphery. Even those chromatin bodies which appeared to be completely surrounded by the fibrillar nucleolar component on single ultrathin sections are actually settled down in nucleolus cavities open to the nucleoplasm. This proves that the RNA processing in D. nasutum nucleoli is directed toward the center of nucleoli, where the granular component is located. The analysis of the nucleolar chromatin distribution made it possible to conclude that different parts of the complex interfase netlike nucleoli of D. nasutum have approximately the same activity.     

REFORMATION OF NUCLEOLI AFTER ETHIONE-INDUCED FRAGMENTATION IN THE ABSENCE OF SIGNIFICANT PROTEIN SYNTHESIS

The rat liver nucleolus, after fragmentation induced by ethionine treatment, has been found to undergo complete reformation by adenine in the presence of a dose of cycloheximide sufficient to cause inhibition of protein synthesis by 90–95%. In contrast, actinomycin D given along with adenine was followed by the appearance of a small compact mass containing only the fibrillar component with no evident granules. This structure resembled pseudonucleoli seen in the anucleolate mutant of Xenopus laevis or in certain early stages of amphibian oocytes. Actinomycin D administered 2 hr after adenine induced a segregation of the fibrillar and granular components of nucleoli similar to that induced in the normal nucleolus. The implications of these findings in relation to nucleolar organization are briefly discussed.     

Autoradiographic, ultrastructural and interferometric analyses of the temperature effect on the synthesis and migration of ribosomes in root meristem cells of Helianthus annuus L. relation to S phase initiation

Experimental model consisted in blocking cells in G1 phase by cold treatment (12 h, 10 degrees C); following 3 h of postincubation at 20 degrees C, cells initiated S phase. In the present studies it has been shown that 2 h postincubation at 20 degrees C of cold-treated young seedlings of Helianthus annuus L. results in transformation of inactive meristematic nucleoli, characterized by small sizes, reduced amount of dry mass and granular component and by the presence of few and large fibrillar centres into large active nucleoli displaying high dry mass and granular component contents, numerous and small fibrillar centres. After 3 h of postincubation at 20 degrees C, nucleoli lose their granular component, decrease in size and dry mass content. At this moment cytoplasm enriches in ribosomes and its dry mass increases. Maximum of nucleolar activity is preceded by an accumulation of proteins in nucleoli. It is concluded that an enhanced transport of ribosomes is one of the conditions of S phase initiation.     

Rearrangement of nucleoli in hepatocytes stimulated for proliferation and enhancement of their transcription function

The nucleoli of normally functioning guinea-pig hepatocytes that have a nucleolonemal (strand-like) organization differ from identical nucleoli of other cells. Their nucleolonema consists as a rule of a fibrillar component with 45S RNA and is poor in granulas that contain pre-rNA molecules of an intermediate size and 28S rRNA, a dense fibrillar component with nascent rRNPs in its composition was not revealed. In hepatocytes stimulated by a 2/3 liver resection rearrangements in nucleoli were found. This brought to a conclusion that rRNA metabolism undergoes some changes. In 2.5 and 5 hours after the resection the hepatocytes' nucleoli were characteristic of a greater thickness of strands and a smaller size of vacuoles, appearance of distinct zones of the dense fibrillar component and an increased amount of RNP-granules. All these observations taken together point out at an increased synthesis and processing of rRNA at early stages of the prereplicative period. In 9 hours the character of changes in nucleoli was different: the vacuoles were considerably widened, whereas the thickness of strands that consisted of a well-expressed dense fibrillar, fibrillar and granular components was lesser. Such rearrangement points out at an increased transport of preribosomes from the nucleolus, a high level of synthesis and processing of nascent RNP-product being maintained. The changes of nucleolar RNP-component were followed by appearance of greater blocks of perinucleolar condensed chromatin, which may be connected with "cutting-off" some tissue-specific genes and initiation of functioning of the mitotic operon genes.     

Nucleolar structure and synthetic activity during meiotic prophase and spermiogenesis in the rat

The ultrastructure of nucleoli was examined in developing rat spermatocytes and spermatids, with the help of serial sections. In addition, the radioautographic reaction of nucleoli as examined in rats sacrificed 1 hr after intratesticular injection of 3H(5')-uridine and taken as an index of the rate of synthesis of ribosomal RNA (rRNA). Primary spermatocytes from preleptotene to zygotene have small nucleoli typically composed of fibrillar centers, a fibrillar component, and a granular component, within which are narrow interstitial spaces. During early and mid-pachytene, nucleoli enlarge to about nine times their initial size, with the fibrillar and granular components forming an extensive network of cords--a nucleolonema--within which are wide interstitial spaces. Meanwhile, there appear structures identical to the granular component but distinct from nucleoli; they are referred to as extranucleolar granular elements. Finally, from late pachytene to the first maturation division, nucleoli undergo condensation, as shown by contraction of fibrillar centers into small clumps, while fibrillar and granular components condense and segregate from each other, with a gradual decrease in interstitial spaces. In secondary spermatocytes, nucleoli are compact and rather small, while in young spermatids they are also compact and even smaller. Nucleoli disappear in elongating spermatids. In 3H-uridine radioautographs, nucleolar label is weak in young primary spermatocytes, increases progressively during early pachytene, is strong by the end of mid pachytene, but gradually decreases during late pachytene up to the first maturation division. In secondary spermatocytes and spermatids, there is no significant nucleolar label. In conclusion, rRNA synthesis by nucleoli is low in young spermatocytes. During pachytene, while nucleoli enlarge and form a lacy nucleolonema, rRNA synthesis increases gradually to a high level by the end of mid pachytene. However, during the condensation and segregation of nucleolar components occurring from late pachytene onward, the synthesis gradually decreases and disappears. The small, compact spermatids arising from the second maturation division do not synthesize rRNA.     

Fine structure of nucleoli in micronucleated cells

The correlation between the number of nucleolus organizing regions (NOR) on metaphase chromosomes and the number of nucleoli was studied in normal and micronucleate cells. Many micronuclei, but not all, were able to form complete nucleoli with fibrillar and granular RNP components and fibrillar centers. Micronuclei which failed to form complete nucleoli often contained multiple electron-dense bodies of fibrillar material. These structures, which were much smaller than nucleoli, reacted with nucleolus-specific antibodies and the Ag-As method in the same way as complete nucleoli, but lacked fibrillar centers and granular RNP components. The data suggest that these nucleolus-like ‘blobs’ contain nucleolar material which, following mitosis, has been enclosed in micronuclei which do not contain nucleolus organizing chromosomes. No evidence was found for the activation of latent NORs not expressed in mononucleate cells.