TY - JOUR A1 - Thiry, Marc A1 - Scheer, Ulrich A1 - Goessens, Guy T1 - Immunoelectron microscopic study of nucleolar DNA during mitosis in Ehrlich tumour cells N2 - In order to investigate the DNA localization within Ehrlich tumor cell nucleoli during mitosis, two recent immunocytochemical methods using either an anti-DNA or an anti-bromodeoxyuridine (BrdU) monoclonal antibody have been applied. In both cases, the immunogold labeling has been performed on ultrathin sections of cells embedded either in Lowicryl K4M or in Epon, respectively. Identical results are observed with both immunocytochemical approaches. In the interphase nucleolus, besides the labeling of the perinucleolar chromatin shell and of its intranucleolar invaginations which penetrate into the nucleolar body and often terminate at the fibrillar centers, a few gold particles are also preferentially found towards the peripheral region of the fibrillar centers. In contrast, the dense fibrillar component and the granular component are never labeled. During mitosis, the fibrillar centers persist at the chromosomal nucleolus organizing regions (NOR's) and can be selectively stained by the silver method. However, these metaphase fibrillar centers are no longer decorated by the DNA- or BrdU antibodies. These results indicate that until the end of prophase, rRNA genes are present inside the fibrillar center material, disappear during metaphase and reappear in reconstituting nucleoli during telophase. Thus, fibrillar centers appear to represent structures sui generis, which are populated by rRNA genes only when the nucleolus is functionally active. In segregated nucleoli after actinomycin D treatment, the DNA labeling is exclusively restricted to the perinucleolar chromatin blocks. These findings also suggest that the DNA content of the fibrillar center material varies according to the rRNA transcription level of the cells. The results are discussed in the light of the present knowledge of the functional organization of the nucleolus. KW - Cytologie KW - Nucleolus KW - DNA KW - mitosis Y1 - 1988 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-40745 ER - TY - JOUR A1 - Rose, Kathleen M. A1 - Szopa, Jan A1 - Han, Fu-Sheng A1 - Cheng, Yung-Chi A1 - Richter, Arndt A1 - Scheer, Ulrich T1 - Association of DNA topoisomerase I and RNA polymerase I: A possible role for topoisomerase I in ribosomal gene transcription N2 - RNA polymerase I preparations purified from a rat hepatoma contained DNA topoisomerase activity. The DNA topoisomerase associated with the polymerase had an Mr of 110000, required Mg2+ but not ATP, and was recognized by anti-topoisomerase I antibodies. When added to RNA polymerase I preparations containing topoisomerase activity, anti-topoisomerase I antibodies were able to inhibit the DNA relaxing activity of the preparation as well as RNA synthesis in vitro. RNA polymerase II prepared by analogous procedures did not contain topoisomerase activity and was not recognized by the antibodies. The topoisomerase I: polymerase I complex was reversibly dissociated by column chromatography on Sephacryl S200 in the presence of 0.25 M (NH4hS04. Topoisomerase I was immunolocalized in the transcriptionally active ribosomal gene complex containing RNA polymerase I in situ. These data indicate that topoisomerase I and RNA polymerase I are tightly complexed both in vivo and in vitro, and suggest a role for DNA topoisomerase I in the transcription of ribosomal genes. Y1 - 1988 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-33901 ER - TY - JOUR A1 - Reimer, Georg A1 - Raska, Ivan A1 - Scheer, Ulrich A1 - Tan, Eng M. T1 - Immunolocalization of 7-2-ribonucleoprotein in the granular component of the nucleolus N2 - Certain autoimmune sera contain antibodies against a nucleolar ribonucleoprotein particle associated with 7-2-RNA (R. Reddy et al. (1983) J. Bioi. Chem . 258, 1383; C. Hashimoto and J. A. Steitz (1983) J. Bioi. Chem. 258, 1379). In this study, we showed by immunofluorescence microscopy that antibodies reactive with 7-2-ribonucleoprotein immunolocalized in the granular regions of actinomycin D and 5,6-dichloro-I-j3-D-ribofuranosylbenzimidazole (DRB)-segregated nucleoli from Vero cells. By electron microscopic immunocytochemistry, antigen-antibody complexes were located in the granular component of transcriptionally active nucleoli from rat liver hepatocytes and HeLa cells. Anti-7- 2-RNP antibodies from two autoimmune sera immunoprecipitated a major protein of Mr 40,000 from e5S] methionine-Iabeled HeLa cell extract. The immunolocalization data suggest that 7-2-ribonucleoprotein may be involved in stages of ribosome biogenesis which take place in the granular component of the nucleolus, i.e., assembly, maturation, and/or transport of preribosomes Y1 - 1988 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-33890 ER - TY - JOUR A1 - Benavente, Ricardo A1 - Schmidt-Zachmann, Marion S. A1 - Hügle-Dörr, B. A1 - Reimer, G. A1 - Rose, K. M. A1 - Scheer, Ulrich T1 - Identification and definition of nucleolus-related fibrillar bodies in micronucleated cells N2 - Small nucleolus-related bodies which occur in the nUcleoplasm of " micronuclei" lacking nucleolar organizers have been studied by immunofluorescence microscopy. These bodies stained specifically with three different antibodies directed against proteins that are normally associated with the dense fibrillar component of functional nucleoli, but not with antibodies specific for certain proteins of the granular component or the fibrillar centers. Our data show that, in the absence of rRNA genes, the various constituent proteins characteristic of the dense fibrillar component spontaneously assemble into spherical entities but that the subsequent fusion of these bodies into larger structures is prevented in these micronuclei. The similarity between these nucleolus-related bodies of micronuclei and the prenucleolar bodies characteristic of early stages of nucleologenesis during mitotic telophase is discussed. Y1 - 1988 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-39423 ER - TY - JOUR A1 - Thiry, Marc A1 - Scheer, Ulrich A1 - Goessens, Guy T1 - Localization of DNA within Ehrlich tumour cells nucleoli by immunoelectron microscopy N2 - The distribution of DNA in Ehrlich tumour cell nucleoli was investigated by means of an immunocytochemical approach , involving a monoclonal antibody directed against double- and single-stranded DNA. Immunolabelling was performed . either before or after the embedding process. The postembedding labelling method allows better ultrastructural preservation than the preembedding labelling method. In particular, the various nucleolar components are well preserved and identifiable. In the nucleolus, labelling is particularly concentrated over the perinucleolar chromatin and over its intranucleolar invaginations, which penetrate the nucleolar body and often terminate at the fibrillar centres. In addition, aggregates of gold particles are found in the fibrillar centres, preferentially towards the peripheral regions. By contrast, the dense fibrillar component is completely devoid of labelling. The results seem to indicate that DNA containing the rDNA genes is located in the fibrillar centres, with a preference for the peripheral regions. This finding suggests that transcription of the rDNA genes should occur within the confines of the fibrillar centre, probably close to the boundary region of the surrounding dense fibrillar component. The results are discussed in the light of present knowledge of the functional organization of the nucleolus. KW - nucleolus KW - DNA KW - monoclonal antibody Y1 - 1988 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-39327 ER - TY - JOUR A1 - Dabauvalle, Marie-Christine A1 - Schulz, Barbara A1 - Scheer, Ulrich A1 - Peters, Reiner T1 - Inhibition of nuclear accumulation of karyophilic proteins in living cells by microinjection of the lectin wheat germ agglutinin N2 - No abstract available Y1 - 1988 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-34288 ER - TY - JOUR A1 - Scheer, Ulrich A1 - Dabauvalle, Marie-Christine A1 - Merkert, Hilde A1 - Benavente, Ricardo T1 - The nuclear envelope and the organization of the pore complexes N2 - No abstract available Y1 - 1988 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-34275 ER - TY - JOUR A1 - Benavente, Ricardo A1 - Reimer, Georg A1 - Rose, Kathleen M. A1 - Hügle-Dörr, Barbara A1 - Scheer, Ulrich T1 - Nucleolar changes after microinjection of antibodies to RNA polymerase I into the nucleus of mammalian cells N2 - After microinjection of antibodies against RNA polymerase I into the nuclei of cultured rat kangaroo (PtKz) and rat (RVF-SMC) cells alterations in nucleolar structure and composition were observed. These were detected by electron microscopy and double-label immunofluorescence microscopy using antibodies to proteins representative of the three major components of the nucleolus. The microinjected antibodies produced a progressive loss of the material of the dense fibrillar component (DFC) from the nucleoli which, at 4 h after injection, were transformed into bodies with purely granular component (GC) structure with attached fibrillar centers (FCs). Concomitantly, numerous extranucleolar aggregates appeared in the nucleoplasm which morphologically resembled fragments of the DFC and contained a protein (fibrillarin) diagnostic for this nucleolar structure. These observations indicate that the topological distribution of the material constituting the DFC can be experimentally influenced in interphase cells, apparently by modulating the transcriptional activity of the rRNA genes. These effects are different from nucleolar lesions induced by inhibitory drugs such as actinomycin D-dependent "nucleolar segregation". The structural alterations induced by antibodies to RNA polymerase I resemble, however, the initial events of nucleolar disintegration during mitotic prophase. Y1 - 1988 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-40666 ER - TY - JOUR A1 - Reimer, Georg A1 - Raska, Ivan A1 - Tan, Eng M. A1 - Scheer, Ulrich T1 - Human autoantibodies: probes for nucleolus structure and function N2 - No abstract available Y1 - 1987 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-41410 ER - TY - JOUR A1 - Benavente, Ricardo A1 - Rose, Kathleen M. A1 - Reimer, Georg A1 - Hügle-Dörr, Barbara A1 - Scheer, Ulrich T1 - Inhibition of nucleolar reformation after microinjection of antibodies to RNA polymerase I into mitotic cells N2 - The formation of daughter nuclei and the reformation of nucleolar structures was studied after microinjection of antibodies to RNA polymerase I into dividing cultured cells (PtK2). The fate of several nucleolar proteins representing the three main structural subcomponents of the nucleolus was examined by immunofluorescence and electron microscopy. The results show that the RNA polymerase I antibodies do not interfere with normal mitotic progression or the early steps of nucleologenesis, i.e. , the aggregation of nucleolar material into prenucleolar bodies. However,they inhibit the telophasic coalescence of the prenucleolar bodies into the chromosomal nucleolar organizer regions, thus preventing the formation of new nucleoli. These prenucleolar bodies show a fibrillar organization that also compositionally resembles the dense fibrillar component of interphase nucleoli . We conclude that during normal nucleologenesis the dense fibrillar component forms from preformed entities around nucleolar organizer regions, and that this association seems to be dependent on the presence of an active form of RNA polymerase I. Y1 - 1987 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-33247 ER - TY - CHAP A1 - Scheer, Ulrich T1 - Contributions of electron microscopic spreading preparations ("Miller-spreads") to the analysis of chromosome structure N2 - No abstract available KW - Eukaryonten / Chromosom Y1 - 1987 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-39625 ER - TY - JOUR A1 - Scheer, Ulrich A1 - Raska, I. T1 - Immunocytochemical localization of RNA polymerase I in the fibrillar centers of nucleoli N2 - No abstract available Y1 - 1987 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-39618 ER - TY - JOUR A1 - Scheer, Ulrich T1 - Structure of lampbrush chromosome loops during different states of transcriptional activity as visualized in the presence of physiological salt concentrations N2 - Lampbrush chromosomes of amphibian oocytes were isolated in the presence of near-physiological salt concentrations, to preserve their native state, and studied by electron microscopy of ultrathin s~dions. The transcriptional state of the lampbrush chromosomes was experimentally modulated by incubating the oocytes for various time periods in medium containing actinomycin D. The observations show that the structure of the lateral loops changes rapidly in response to alterations in transcriptional activity. During decreasing transcriptional activity and reduced packing density of transcripts, the chromatin axis first condensed into nucleosomes and then into an approximately 30 nm thick higher order chromatin fiber. Packaging of the loop axis into supranucleosomal structures may contribute to the foreshortening and retraction of the loops observed during inhibition of transcription and in later stages of meiotic prophase. The increasing packing density of the DNA during the retraction process of the loops could also be visualized by immunofluorescence microscopy using antibodies to DNA. The dependence of the loop chromatin structure on transcriptional activity is discussed in relation to current views of mechanisms involved in gene activation. KW - lampbrush chromosomes KW - chromatin structure KW - electron microscopy KW - immunofluorescence microscopy KW - DNA antibodies Y1 - 1987 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-39304 ER - TY - JOUR A1 - Reimer, Georg A1 - Rose, Kathleen M. A1 - Scheer, Ulrich A1 - Tan, Eng M. T1 - Autoantibody to RNA polymerase I in scleroderma sera N2 - Autoantibodies to components of the nucleolus are a unique serological feature of patients with scleroderma. There are autoantibodies of several specificities; one type produces a speckled pattern of nucleolar staining in immunofluorescence. In actinomycin D and 5,6-dichloro-{j-D-ribofuranosylbenzimidazoletreated Vero cells, staining was restricted to the fibrillar and not the granular regions. By double immunofluorescence, specific rabbit anti-RNA polymerase I antibodies stained the same fibrillar structures in drug-segregated nucleoli as scleroderma sera. Scleroderma sera immunoprecipitated 13 polypeptides from (35S)methionine-labeled HeLa cell extract with molecular weights ranging from 210,000 to 14,000. Similar polypeptides were precipitated by rabbit anti-RNA polymerase I antibodies, and their common identities were confirmed in immunoabsorption experiments. Microinjection of purified IgG from a patient with speckled nucleolar staining effectively inhibited ribosomal RNA transcription. Autoantibodies to RNA polymerase I were restricted to certain patients with scleroderma and were not found in other autoimmune diseases. Y1 - 1987 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-34294 ER - TY - JOUR A1 - Scheer, Ulrich A1 - Messner, Karin A1 - Hazan, Rachel A1 - Raska, Ivan A1 - Hansmann, Paul A1 - Falk, Heinz A1 - Spiess, Eberhard A1 - Franke, Werner W. T1 - High sensitivity immunolocalization of double and single-stranded DNA by a monoclonal antibody N2 - A monoclonal antibody (AK 30-10) is described which specifically reacts with DNA both in double and single-stranded forms but not with other molecules and structures, including deoxyribonucleotides and RNAs. When used in immunocytochemical experiments on tissue sections and permeabilized cultured cells, this antibody detects DNA-containing structures, even when the DNA is present in very small amounts. Examples of high resolution detection include the DNA present in amplified extrachromosomal nucleoli, chromomeres of lampbrush chromosomes, mitochondria, chloroplasts and mycoplasmal particles. In immunoelectron microscopy using the immunogold technique, the DNA was localized in distinct substructures such as the "fibrillar centers" of nucleoli and certain stromal centers in chloroplasts. The antibody also reacts with DNA of chromatin of living cells, as shown by microinjection into cultured mitotic cells and into nuclei of amphibian oocytes. The potential value and the limitations of immunocytochemical DNA detection are discussed. KW - Cytologie KW - DNA antibodies KW - monoclonal antibodies KW - DNA immunolocalization KW - chromatin KW - mycoplasma tests Y1 - 1987 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-41063 ER - TY - JOUR A1 - Scheer, Ulrich A1 - Hansmann, Paul A1 - Falk, Heinz A1 - Sitte, Peter T1 - Ultrastructural localization of DNA in two Cryptomonas species by use of a monoclonal DNA-antibody N2 - Immunogold cytochemistry - DNA localization - Cryptomonas nucleomorph The distribution and subcellular localization of DNA in the unicellular alga Cryptomonas has been investigated electron-microscopically by indirect immunocytochemistry, using a monoclonal DNA antibody and a gold-Iabeled secondary antibody. This technique proved to be very sensitive and entirely specific. DNA could be demonstrated in four different compartments (nucleus, nucleomorph, plastid, and mitochondrion). Within the plastid, DNA is concentrated in stroma regions that are localized preferentially around the center of the organelle. The mitochondrion contains several isolated DNA-containing regions (nucleoids). Within the nucleus, most of the DNA is localized in the 'condensed' chromatin. DNA was also detectable in small areas of the nucleolus, whereas the interchromatin space of the nucleus appeared almost devoid of DNA. Within the nucleomorph, DNA is distributed inhomogeneously in the matrix. DNA could furthermore be detected in restricted areas of the 'fibrillogranular body' of the nucleomorph, resembling the situation encountered in the nucleol us. The presence of DNA and its characteristic distribution in the nucleomorph provide additional, strong evidence in favour of the interpretation of that organelle as the residual nucleus of a eukaryotic endosymbiont in Cryptomonas. KW - Cytologie Y1 - 1986 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-39746 ER - TY - JOUR A1 - Hadjiolova, Krassimira A1 - Rose, Kathleen M. A1 - Scheer, Ulrich T1 - Immunolocalization of nucleolar proteins after D-galactosamine-induced inhibition of transcription in rat hepatocytes N2 - No abstract available Y1 - 1986 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-33205 ER - TY - JOUR A1 - Reimer, Georg A1 - Scheer, Ulrich A1 - Peters, Jan-Michael A1 - Tan, Eng M. T1 - Immunolocalization and partial characterization of a nucleolar autoantigen (PM-Scl) associated with polymyositis / scleroderma overlap syndromes. N2 - Precipitating anti-PM-Sel antibodies are present in sera from patients with polymyositis. scleroderma. and polymyositis/scleroderma overlap syndromes. By indirect immunofluorescence microscopy. anti-PM-Scl antibodies stained the nucleolus in cells of different tissues and species. suggesting that the antigen is highly conserved. By electron microscopy, anti-PM-Scl antibodies reacted primarily with the granular component of the nuc1eolus. Drugs that inhibit rRNA synthesis had a marked effect on the expression of PM-Scl antigen. In actinomycin D-treated cells, immunofluorescence staining by anti-PM-Scl was signüicantly reduced with residual staining restricted to the granular regions of nuc1eoli. Treatment with 5,6-dichloro-beta-D- ribofuranosylbenzimidazole (DRB) also selectively reduced nuc1eolar staining. On a molecular level, anti-PM-Sel antibodies precipitated 11 polypeptides with molecular weights (Mr) ranging from 110,000 to 20,000. The Mr 80,000 and 20.000 polypeptides were phosphorylated. Evidence suggests that the PM-Scl antigen complex may be related to a prerlbosomal particle. Y1 - 1986 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-33191 ER - TY - JOUR A1 - Scheer, Ulrich T1 - Injection of antibodies into the nucleus of amphibian oocytes: an experimental means of interfering with gene expression in the living cell N2 - No abstract available Y1 - 1986 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-41182 ER - TY - JOUR A1 - Hügle, Barbara A1 - Scheer, Ulrich A1 - Franke, Werner W. T1 - Ribocharin: a nuclear M\(_r\) 40,000 protein specific to precursor particles of the large ribosomal subunit N2 - Using a monoclonal antibody (No-194) we have identified, in Xenopus laevis and other amphibia, an acidic protein of M, 40,000 (ribocharin) which is specifically associated with the granular component of the nucleolus and nucleoplasmic 65S particles. These particles contain the nuclear 28S rRNA and apparently represent the precursor to the large ribosomal subunit in nucleocytoplasmic transit. By immunoelectron microscopy ribocharin has been localized in the granular component of the nucleolus and in interchromatin granules. During mitosis ribocharin-containing particles are associated with surfaces of chromosomes and are recollected in the reconstituting nucleoli in late telophase. We suggest that ribocharin is a specific component of precursor particles of the large ribosomal subunit, which dissociates from the 65S particle before passage through the nuclear envelope, and is reutilized in ribosome biogenesis. Y1 - 1985 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-41169 ER -