Fine structural organization of a non-reticulate plant cell nucleus

We studied the fine structural organization of the meristematic nucleus in roots of Lycopesicon esculentum (tomato) using ultracytochemical and immunocytochemical approaches. The nucleus has a non-reticulate (i.e. low DNA content) structure whose supramolecular organization differs in some respects from that in reticulate nuclei, principally in the organization of the chromocentres associated with the nuclear envelope, with which centromeric structures appear to be associated. The main difference at the nucleolar level is found in the fibrillar centres, which have a low amount of DNA labelling and in which inclusions of condensed chromatin are present only very rarely. The distribution of nucleolar DNA amongst the nucleolar compartments is similar to that in reticulate nucleoli as demonstrated using an anti-DNA monoclonal antibody. Tomato nuclei have nucleolus-associated bodies or karyosomes, like other plant species with a low DNA content and non-reticulate nuclear organization. The nuclear ribonucleoprotein structures in the inter- and perichromatin regions, namely inter- and perichromatin fibrils and granules, show similar ultrastructural and cytochemical characteristics in both types of nuclei.Abbreviations NAC nucleolus associated chromatin - CES centromeric structures - NOR nucleolar organizing region - NAB nucleolus associated body - IG interchromatin granules - RNP ribonucleoprotein - Mab monoclonal antibody by M.F. Trendelenburg     

豌豆细胞核仁中rDNA的超微定位和排布构型

利用细胞化学DNA特异染色法——NAMA-Ur特异染色法对豌豆细胞核仁中rDNA的位置及其排布构型进行了原位观察。结果表明,核仁中的rDNA位于纤维中心(FC)以及FC与致密纤维组分的交界处,以环绕FC的形式排布。不同位置的rDNA成分都具有集缩和解集缩两种形态结构,核仁外的核仁伴随染色质经过核仁通道进入核仁,沿FC周边排列,与其中的DNA相连。     

豌豆细胞核仁中rDNA的超微定位和排布构型

利用细胞化学DNA特异染色法——NAMA-Ur特异染色法对豌豆细胞核仁中rDNA的位置及其排布构型进行了原位观察。结果表明,核仁中的rDNA位于纤维中心(FC)以及FC与致密纤维组分的交界处,以环绕FC的形式排布。不同位置的rDNA成分都具有集缩和解集缩两种形态结构,核仁外的核仁伴随染色质经过核仁通道进入核仁,沿FC周边排列,与其中的DNA相连。     

Cytochemical variations in the nucleolus during spermiogenesis in man and monkey

Summary The fine structure, nature and fate of the components of the nucleolus were studied in young (steps 1, 2), intermediate (steps 3, 4, 5) and mature spermatids (steps 6, 7, 8) of man and monkey, by use of several cytochemical techniques (alcoholic PTA; sodium tungstate; EDTA; HAPTA; nuclease-gold complexes; NOR silver staining). As controls, comparative ultrastructural and cytochemical observations of the nucleolus in spermatids and Sertoli cells were made in the same sections of seminiferous tubules. In the young spermatids of the two species studied, the nucleolar masses exhibited identical features. Segregation of the nucleolar components took place in the nuclei of step 1 spermatids. No typical fibrillar center was observed. In spermatids at steps 1 and 2, the nucleolar masses appeared to be made up of two fibrillar components of equal density, one spherule-shaped, the other forming cords, both surrounded by clusters of 15–20 nm-diameter granules. Alcoholic PTA and sodium tungstate yielded a selective positive contrast of the two fibrillar components whereas EDTA and RNase-gold reacted with the peripheral granular material. Treatment with RNase-gold and DNase-gold complexes resulted in preferential labeling at the periphery of the fibrillar components. After NOR silver staining, numerous small silver grains were localized over the fibrillar cords, suggesting the persistence of specific acidic non-histone proteins. On the contrary, the spherule was never stained. In intermediate spermatids, when the nucleolar components were dissociated, scattered clusters of granules stained by EDTA and HAPTA remained in the entire nucleoplasm. Nucleolar disintegration was accompanied by dispersion of argyrophilic material. In mature spermatids granular material revealed by PTA and silver staining methods was found in the nuclear pockets bounded by the redundant nuclear envelope.     

Ultrastructural cytochemistry of the nucleolus in rat oocytes at the end of the folliculogenesis

Various ultrastructural changes occur during follicular growth in the rat oocyte nucleolus. The nucleolus, which has a reticulated fibrillogranular structure at the primordial and primary follicle stages, becomes entirely compact and is made up of a conspicuous and homogeneous mass at the antral follicle stage. In order to define the nature and the functions of this homogeneous mass, cytochemical methods allowing detection of nucleic acids, proteins and lipids were performed at the light microscopic and ultrastructural levels. The results obtained suggest that this nucleolar mass is probably composed of acid proteins which are not silver stained. This proteinaceous mass could be a special kind of nucleolar secretion providing material for meiotic resumption in the oocyte. Cytochemical researches now in progress should supply new information concerning the exact nature and the role of the nucleolar compact mass, which is the essential nucleolar component at the antral follicle stage and which really plays a role in the nucleolus in the first stages of embryogenesis.     

Ultrastructural cytochemistry of the nucleolus in rat oocytes at the end of the folliculogenesis

Summary Various ultrastructural changes occur during follicular growth in the rat oocyte nucleolus. The nucleolus, which has a reticulated fibrillogranular structure at the primordial and primary follicle stages, becomes entirely compact and is made up of a conspicuous and homogeneous mass at the antral follicle stage. In order to define the nature and the functions of this homogeneous mass, cytochemical methods allowing detection of nucleic acids, proteins and lipids were performed at the light microscopic and ultrastructural levels. The results obtained suggest that this nucleolar mass is probably composed of acid proteins which are not silver stained. This proteinaceous mass could be a special kind of nucleolar secretion providing material for meiotic resumption in the oocyte. Cytochemical researches now in progress should supply new information concerning the exact nature and the role of the nucleolar compact mass, which is the essential nucleolar component at the antral follicle stage and which really plays a role in the nucleolus in the first stages of embryogenesis.     

Oldies but goldies: searching for Christmas trees within the nucleolar architecture

The nucleolus is the prominent nuclear organelle in which the biogenesis of ribosomal RNA and ribosomes takes place. Understanding of the molecular processes in the nucleolus is rapidly expanding; however, opinions and results on the precise localization of active ribosomal genes - in either of two nucleolar subcompartments, fibrillar centers and dense fibrillar components - are still divided. This review discusses the difficulties in studying the nucleolar structure using microscopy, and provides an overview of the published data, critically examining their relevance to the controversy. Additionally, evidence showing that the dense fibrillar components encompass the Christmas tree structures is discussed and ways to reconcile the controversy are proposed.     

The slender lobes gene, identified by retarded mushroom body development, is required for proper nucleolar organization in Drosophila

The nucleolus dynamically alters its shape through the assembly and disassembly of a variety of nucleolar components in proliferating cells. While the nucleolus is known to function in vital cellular events, little is known about how its components are correctly assembled. Through the analysis of a Drosophila mutant that exhibits a reduced number of mushroom body (MB) neurons in the brain, we reveal that the slender lobes (sle) gene encodes a novel nuclear protein that affects nucleolar organization during development. In sle mutant neuroblasts, the nucleolus was packed more tightly, forming a dense sphere, and the nucleolar proteins fibrillarin and Nop60B were abnormally distributed in the interphase nucleolus. Moreover, another nucleolar marker, Aj1 antigen, was localized to the center of the nucleolus in a manner complementary to the Nop60B distribution, and also formed a large aggregate in the cytoplasm. While developmental defects were limited to a few tissues in sle mutants, including MBs and nurse cells, the altered organization of the nucleolar components were evident in most developing tissues. Therefore, we conclude that Sle is a general factor of nuclear architecture in Drosophila that is required for the correct organization of the nucleolus during development.     

Detection of sugar-binding sites in the fibrillar and the granular components of the nucleolus: An experimental study in cultured mammalian cells

The intranucleolar distribution of sugar-binding sites (i.e., lectin-like molecules) was analyzed in segregated nucleoli of actinomycin D-treated HeLa cells. The detection of sugar-binding sites was performed by incubation either of permeabilized nuclei in the presence of fluorescein-labeled neoglycoproteins or of ultrathin sections cut through in situ-fixed nuclei in the presence of gold-labeled neoglycoproteins. In the former case, the fluorescent nucleolar components were identified by comparison with the nucleolar components of similarly treated cells observed in electron microscopy. For the first time, this study reveals the presence of sugar-binding sites in both the fibrillar and the granular components of the nucleolus. In view of the data already reported on the biochemical composition of the nucleolus, some of our results led us to conclude that the nucleolar sugar-binding sites are lectin-like proteins. These proteins could be associated with preribosomes since the nucleolus is the site of both synthesis and stockage of ribosomal precursors. Some results from this study, however, show that the possibility of a relationship between some lectins and a structural component cannot be excluded.     

Nucleoli undergo structural and molecular modifications during hibernation

The nucleolus is a very dynamic structure able rapidly to adapt its activity to the cellular metabolic state. An interesting physiological model characterized by drastic modifications of cellular metabolism is represented by hibernating animals. In the present study we investigated the hepatocyte nuclei of euthermic and hibernating edible dormice (Glis glis) with the aim of revealing, by means of ultrastructural and immunocytochemical analyses, possible modifications of nucleolar components during hibernation. Our observations demonstrate that, in deep hibernation, nucleoli undergo structural and molecular modifications: (a) they show numerous nucleoplasmic invaginations and clumps of dense fibrillar component extend from the nucleolar surface; (b) they are frequently in contact with coiled bodies and fibro-granular material, two nuclear bodies usually occurring in the nucleoplasm; (c) the dense fibrillar component contains significant amounts of small nuclear ribonucleoproteins, splicing factors usually distributed in the nucleoplasm. Taken together, these results suggest that during hibernation complex relationships are established between the nucleolus and nucleoplasm, probably related to functional activities peculiar to this physiological phase. However, since no evident nucleolar modification was found in early hibernating dormice, it seems likely that the particular structural and molecular arrangement of nucleoli establishes progressively during hibernation, becoming evident only in the deepest phase, and then disappears upon arousal.     

Subnucleolar distribution and organization of Vicia faba L. rDNA in situ

The distribution and organization of nucleolar DNA in Vicia faba L. was analyzed by specific cytochemical staining using NAMA-Ur. The results showed that nucleolar DNA was distributed in the FCs and at the FC/DFC junctions. Statistical analysis showed that the rRNA genes occupied about one-third of the total dense fibrillar component region. The rDNA was condensed in some regions and uncondensed in others. Nucleolus-associated chromatin extended from outside the nucleolus to the periphery of the FCs via nucleolar channels, suggesting a possible origin for nucleolar DNA.     

The nucleolus: a model for the organization of nuclear functions

Nucleoli are the prominent contrasted structures of the cell nucleus. In the nucleolus, ribosomal RNAs (rRNAs) are synthesized, processed and assembled with ribosomal proteins. The size and organization of the nucleolus are directly related to ribosome production. The organization of the nucleolus reveals the functional compartmentation of the nucleolar machineries that depends on nucleolar activity. When this activity is blocked, disrupted or impossible, the nucleolar proteins have the capacity to interact independently of the processing activity. In addition, nucleoli are dynamic structures in which nucleolar proteins rapidly associate and dissociate with nucleolar components in continuous exchanges with the nucleoplasm. At the time of nucleolar assembly, the processing machineries are recruited in a regulated manner in time and space, controlled by different kinases and form intermediate structures, the prenucleolar bodies. The participation of stable pre-rRNAs in nucleolar assembly was demonstrated after mitosis and during development but this is an intriguing observation since the role of these pre-rRNAs is presently unknown. A brief report on the nucleolus and diseases is proposed as well as of nucleolar functions different from ribosome biogenesis.Robert Feulgen Lecture presented at the 48th Symposium of the Society for Histochemistry in Stresa, Lake Maggiore, Italy, 7–10 September 2006.     

Electron Microscopic Studies on the Silver-stained Nucleolar Cycle of Physarum polycephalum

The dynamic changes of nucleolar ultrastructure in the cell cycle of Physarum polycephalum Schw. were studied by an en bloc silver-staining method. The results showed that the nucleolus was large in size and situated in the center of the nucleus in late G2-phase, and the fibrillar centers, dense fibrillar components and granular components could be observed in the nucleolus. During prophase, the nucleolus moved towards the periphery of the nucleus and in late prophase disintegrated near the nuclear envelope. In metaphase, the disintegrated nucleolar components were dispersed in masses and located at the periphery of the chromosomal region of the nucleus. No specifically silver-stained area and argentophilic protein sheath were observed on the chromosomes, but there were some big dispersed silver particles within the chromosomes. During telophase the nucleolar components moved towards the two poles along with the chromosomes and co-existed with the decondensing chromatin in daughter nuclei. The nucleolar components then gradually converged with one another and separated from the chromatin. A big nucleolus was formed in the nucleus about 120 min after the completion of mitosis.