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[The structure of the nucleolus during the inhibition of RNA-and protein synthesis (author's transl)].

In cells treated with antimetabolites to inhibit RNA- and protein synthesis, electron microscopic studies reveal structural alterations of the nucleolus. The morphological appearance of the nucleolus differs depending of the inhibitor used. If transcription is prevented, segregation of nucleolar components is observed. Inhibition of processing of newly synthesized RNA results in a degranulation and an increase in the amount of nucleolar fibrils. A disturbance of the release of nucleolar ribonucleoproteins into the cytoplasm leads to an enlargement and a hypergranulation of the nucleolus. On the other hand interruption of translation of mRNAs has no immediate effect on the appearance of the nucleolar structure. Only after longer treatment of the cells with the translation inhibitor the nucleolus shrinks and becomes degranulated. The use of inhibitors with clearly defined mechanisms of action in a morphological study should make it possible to interpret similar nucleolar alterations seen in cancer cells and virus-infected cells on a molecular biological basis.

Animals

Whole mount electron microscopy of the nucleolus in salivary gland cells of Drosophila melanogaster.

The nucleolus of Drosophila melanogaster salivary gland cells, examined by whole mount electron microscopy, consists of a fibrillar core region and a peripheral region containing both fibres and granules. These regions appear to correspond to the fibrillar and granular components, respectively, seen in thin sections. Most of the nucleoli were attached to the chromocenter region of the polytene chromosomes, containing the nucleolar organizer. Bundles of relatively straight chromatin fibres, 13 nm in diameter, extended from the chromocenter into the core region of the nucleolus, however it was not possible to trace the path of these chromatin fibres through the nucleolus since they were obscured within the mass of nucleolar fibres. The nucleolar fibres in both the core and peripheral regions were irregular and knobby, with a diameter of about 15 nm. In the core region, the fibres appeared to be of considerable length and were characteristically clustered together to form small interconnected masses. The fibres in the peripheral region were relatively short and some appeared to blend with amorphous, poorly-defined pools of material. Electron dense granules 15-20 nm in diameter were also associated with this amorphous substance. It is hypothesized that the formation and subsequent packaging of the 28s rRNA may be represented by a morphological transition of the peripheral fibres, via an amorphous pool-like intermediate stage, into the nucleolar granules. The results of this study indicate that whole mount electron microscopy may be a useful alternative to thin sectioning in high resolution studies of the nucleolus.

Animals

A quantitative autoradiographic study of nucleolus-associated RNA and DNA synthesis during the eclipse phase in Rous sarcoma virus-infected chicken fibroblasts.

Functional and morphologic differences between the sensitivity of nucleoli of Rous sarcoma virus-transformed cells and that of newly infected cells to the action of actinomycin D (AD) have been demonstrated by quantitative light and electron microscope autoradiography and utilized to investigate the function of the nucleolus in the early stages of infection. After a pulse exposure to low doses of AD, increased RNA synthesis is induced within 80 minutes in the fibrillar portion of the nucleolus by infection. A concomitant increase in the retention of tritiated AD in the nucleolus and a quantitative redistribution of intranuclear and cytoplasmic DNA label are interpreted as evidence for a virus-induced amplification of the binding sites of AD in nucleolar chromatin.

Animals

[Ultrastructural identification of the nucleolus organizer by the silver staining technic].

The nucleolus organizer regions can be selectively stained in metaphase chromosome preparations by the Goodpasture and Bloom's technique which was adapted to electron microscopy analysis of cells during interphase. Using this technique, a selective accumulation of silver grains was observed over nucleolus light areas. This selective accumulation allows the identification of the interphase fibrillar centers as the nucleolus organizer regions. Ultrastructural relationships between fibrillar centers and dense fibrillar component are discussed.

Cell Nucleolus

Different behaviour of Xbb+ and Xbb chromosomes in D. melanogaster with only one nucleolus organizer.

We have examined the rDNA content of male and female adult flies having only one nucleolus organizer (NO), using X chromosomes carrying wild or partially deleted bobbed loci (Xbb+/O, Xbb+/XNO- and Xbb/O, Xbb/XNO-). The results show that in Xbb+/O and Xbb+/XNO- flies, where only somatic gene compensation is supposed to occur, the rDNA increase, althought less pronounced than previously reported, is directly proportional to the number of rRNA genes initially present in the nucleolus organizer. In Xbb/O and in Xbb/Xbb/XNO- flies the rDNA increase is relatively much higher than that observed in flies carrying bb+ instead of bb. It is suggested that this may be due to rDNA premagnification and somatic gene compensation occurring simultaneously in the former flies.

Animals

Detection of nucleolus organizer regions in chromosomes of human, chimpanzee, gorilla, orangutan and gibbon.

Nucleolus organizer regions were detected by the Ag-AS silver method in fixed metaphase chromosomes from human and primates. In the human, silver was deposited in the secondary constriction of a maximum of five pairs of acrocentric chromosomes: 13, 14, 15, 21 and 22. The chimpanzee also had five pairs of acrocentric chromosomes stained, corresponding to human numbers 13, 14, 18, 21 and 22. A gibbon had a single pair of chromosomes with a secondary constriction, which corresponded to the nucleolus organizer region. In each case the Ag-AS method detected the sites which have been shown by in situ hybridization to contain the ribosomal RNA genes. An orangutan had eight pairs of acrocentric chromosomes stained with Ag-AS, probably corresponding to human numbers 13, 14, 15, 18, 21 and 22, plus two others. Two gorillas had silver stain over two pairs of small acrocentric chromosomes and at the telomere of one chromosome 1. The larger gorilla acrocentric chromosomes had no silver stain although they all had secondary constrictions and entered into satellite associations.

Animals

The effects of some carcinogenic nitroso compounds on the rat liver nucleolus.

The hepatocyte nucleoli of rats undergo a variety of specific and nonspecific alterations after the administration of 8 nitroso compounds and 5 corresponding non-nitroso compounds. After the oral administration of dimethylnitrosamine and diethylnitrosamine, and unusual segregation of the nucleolus was encountered in the cells with karyorrhexis. Most characteristic of the segregation are the scattered ribosome-like granules at the outermost layer of the altered nucleolus. Frequently microspherules with halos 150 nm in diameter are found in all rats treated with nitroso compounds, which have been reported to be carcinogenic in the organs other than the liver. With the exception of butylurea and butylamine no such specific microspherules were found in the nucleoli after the administration of corresponding non-nitroso compounds.

Animals

N-banding and nucleolus organisers in Asellus aquaticus (Crust. Isop.).

The N-banding technique was used to stain the nucleolus organiser of the karyotype of Asellus aquaticus (crust. Isop.). Observations were made on the morphological expression of nucleolus organisers as secondary constrictions and the presence of nucleoli in mitotic prophase. An attempt was made to correlate the various results and it seems likely that N-banding is not a reflection of NO activity.

Animals

The nature of the Ag-staining of nucleolus organizer regions. Electron- and light-microscopic studies on human cells in interphase, mitosis, and meiosis.

Electron micrographs reveal that the Ag-stainable substance is located on the outside of NOR's or around them but not in the chromosomes themselves. In association figures, the Ag-positive material lies between the acrocentric chromosomes. Light-microscopic studies show that the Ag stainability of the nucleolus in interphase is correlated with the function of the NOR, as seen from inactive and activated lymphocytes. Much more Ag-positive material is seen in prophase than in meta- and anaphase. It starts to increase again in late telophase. In male meiosis the NOR's remain Ag-positive until pachytene. First and second metaphase figures are negative. Experiments using RNase, TCA, and trypsin indicate that the Ag-stainable substance is an acidic protein. The precipitation of Ag granules in interphase nuclei seen in the electron microscope is greatest over the fibrillar component of the nucleolus. The most likely interpretation is that the Ag-stainable material is a component of ribonucleic protein accumulating around active NOR's. In mitosis some of this material remains at the NOR's. In first meiosis it is completely removed before diakinesis.

Animals

Nucleolus organizer activity and the origin of Robertsonian translocations.

Chromosomes with active nucleolus organizer regions (NOR's) were identified by combined Q-banding (in some cases), and silver staining in mouse cell lines. NOR-bearing chromosomes were overrepresented among the chromosomes involved in Robertsonian translocations in LM(TK-), A9, and RAG cell lines. Usually only one NOR-bearing chromosome was seen in any biarmed chromosome; relatively few contained two NOR-bearing chromosomes. Thus the nucleolus plays an important role, but nucleolar fusion is relatively unimportant, in the origin of Robertsonian translocations in the mouse.

Animals

Nucleolus organizers in Mus musculus subspecies and in the RAG mouse cell line.

Silver staining has been used to detect active nucleolus organizer regions (NOR's). By this criterion six mouse chromosomes, numbers 12, 15, 16, 17, 18 and 19, can have an NOR. The number and distribution of chromosomes with NOR's vary among inbred strains of Mus musculus musculus (C57BL/6J, BALB/cJ, C3H/HeJ and C3H/StCpr1BR) and in M. musculus molossinus. In a musculus X molassinus F1 hybrid, nucleolus organizers from each parent are silver stained.--Chromosomes which have NOR's in diploid cells also show them in tetraploid cells and in established cell lines. The BALB/cJ strain shows Ag-staining of NOR's on chromosomes 12, 15, 18 and occasionally 16. In the RAG cell line, which was derived from BALB/c, active NOR's are seen on 12, 15 and 18, even after these chromosomes have undergone structural rearrangements in the cell line. Some correlation exists between the amount of Ag-stain and the size of a secondary construction region, with a large amount of Ag-stain present on a chromosome which has a prominent secondary constriction. There is no correlation between the amount of Ag-stain and the presence or absence of C-band material.

Animals

Localisation of nucleolus-organizing regions in interphase cells.

A technique based on the use of silver solutions, which selectively stains the nucleolus-organising regions (NORs) in chromosomes, was applied to interphase Ehrlich tumour cells. The results indicate that nucleolar fibrillar centres correspond to the NORs.

Animals

Evidence for the inheritance of silver-stained nucleolus organizer regions.

The inheritance of nucleolus organizer regions (NORs) was investigated by examining the degree of silver-staining in individual acrocentric chromosomes in two successive generations. The study was undertaken in six Down's syndrome children and their respective parents. Quinacrine fluorescent polymorphisms were used to identify individual acrocentrics and to determine which of the child's acrocentrics were informative as to parental homologue of origin. Of the 66 acrocentrics in the six children, 31 were informative. The correlation between the degree of silver=staining in the child's chromosomes and the respective parental chromosomes of origin was highly significant (P less than 0.001), with a correlation coefficient of 0.90. The results suggest that the degree of Ag-AS staining is characteristic for a particular chromosome and that this characteristic is an inherited property.

Cell Nucleolus

Nucleolus organiser regions on mitotic and meiotic chromosomes from infertile males investigated using a specific silver stain.

Mitotic preparations from 30 subfertile males and meiotic preparations from 3 normal and 2 subfertile males were examined by means of the Ag-I technique of Bloom and Goodpasture (1976) to reveal nucleolus organiser regions (NORs). In the mitotic preparations, each subject was found to have a characteristic number of Ag-positive NORs per cell, within a range of 6--10. Analysis of satellite associations showed that the mean number of satellite associations per cell was related to the modal number of Ag-positive NORs for each subject. In the meiotic preparations, silver deposition was observed throughout meiotic prophase, but disappeared totally during diakinesis and metaphase II. It was seen again in early spermatids, and disappeared again as nuclear elongation took place. This pattern was observed in both normal and subfertile subjects, and may provide indirect evidence for the activation of rRNA genes during spermatogenesis.

Acrosome

Biochemical and cytogenetic studies on the nucleolus organizing regions (NOR) of man. II. A family with the 15/21 translocation.

The amount of ribosomal DNA (rDNA) was determined quantitatively by RNA-DNA hybridization in the genomes of a mother and her daughter, both with the karyotype 45,XX,t(15q21q). The saturation values found were 0.030% (mother), 0.023% (daughter), and 0.022% for the husband and father of the daughter. A detailed cytogenetic analysis of the short arms of the acrocentric chromosomes of these probands allowed the biochemical results to be interpreted in terms of the size of the individual set of nucleolus organizing regions (NORs) present in each proband. The correlation existing between the biochemical and the cytogenetic findings shows that the amount of rDNA in the human genome is not primarily a function of the number of acrocentric chromosomes, but depends on the individual combination of variant NORs occurring in the human genome.

Cell Nucleolus

Populational polymorphisms in silver staining of nucleolus organizer regions (NORs) in human acrocentric chromosomes.

The Ag stainability of the nucleolus organizer region (NOR) was studied in the acrocentric chromosomes identified by Q banding of cultured lymphocytes in 41 karyotypically normal persons (33 males and 8 females) originating from southeast Estonia. The data obtained are compared with those established earlier for a combined Vienna-Ulm population of 51 karyotypically normal persons (see Mikelsaar et al., 1977a). Significant differences between the two populations in the frequency and patterns of Ag-positive NORs were found. The following findings were most striking: the frequency of Ag-positive NORs in chromosome 14 and in the totals was significantly lower in the Estonian population than in the Vienna-Ulm population (P less than 0.01). The average modal number of Ag-positive NORs per individual was 7.8 in the Estonian population and 8.7 in the Vienna-Ulm sample (P less than 0.01). If the data of the two populations were combined the frequency of positive NORs was significantly (P less than 0.05) lower in chromosome 22 than in 13, 15, and 21, but not 14.

Cells, Cultured

Ag staining of the nucleolus organizer (NO) and its relationship to satellite association.

The frequency of involvement in satellite association and the frequency of selective staining of the secondary constrictions with silver solutions have been studied in five phenotypically normal individuals, all carriers of morphological variants of the nucleolus organizing region (NOR). The results show the preferential involvement of some morphological markers in satellite association, and also their preferential staining with Ag-I. It has also been shown that acrocentric chromosomes involved in satellite association are always stained by silver.

Cell Nucleolus

Silver stain reveals nucleolus organizer regions on a satellited Yq chromosome.

Chromosomes from a patient with a satellited Yq were stained with a silver procedure that differentially stains nucleolus organizer regions. The Yqs stained heavily in all cells examined, indicating the presence of ribosomal cistrons at this region. The Yqs also entered into satellite associations with the D and G group chromosomes at a frequency greater than would be expected through chance.

Azure Stains