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Biomedical subjects

H Busch

Publications and source records attributed to H Busch.

At least 145 records · Page 8Linked to original sources
PubMed1984

Molecular lesions in cancer.

Newer methods of identifying biochemical events associated with cancer include recombinant DNA technology, monoclonal antibodies and improved analysis of nuclear and other cell functions to determine specific events which occur commonly in cancer cells. 'Onc-gene' products offer potential opportunities for new approaches to cancer treatment and the hope of inducing differentiation of cancer cells toward their normal counterparts. Studies on antigens which react with monoclonal antibodies offer the opportunity for 'epitope attack' which may be effected by improved drugs or by design of totally new drugs to bind to specific reactive sites. The complexity and pleiomorphism of cancer do not permit predictions as to whether these approaches will be more effective than the empirical approach to cancer treatment.

Animals↗
PubMed1984

Interactions of U1 RNP with heterogeneous nuclear RNA in rat Novikoff hepatoma nuclei.

The nature of the association of U1 RNA with rapidly sedimenting RNP structures in rat hepatoma nuclei was investigated. The effects of salt and proteinase K treatment on the stability of this 'bound' form of U1 RNA were studied using sucrose density gradient analyses. Quantitation of the amount of U1 RNA remaining associated with large structures after treatment was used to assess the relative contribution of protein-protein (and protein-RNA) versus RNA-RNA interactions. Forty-eight percent of the total nuclear U1 RNA released by sonication was found in a 'bound' form when the sonicate was centrifuged through gradients containing 50 mM NaCl. Fifty percent of this 'bound' U1 RNA remained associated with rapidly sedimenting RNPs when the NaCl concentration was raised to 500 mM. To assess the contribution of protein independent interactions, large RNPs were completely deproteinized and their RNA moieties were then recentrifuged on gradients. By this analysis, 27% of the U1 RNA originally 'bound' to hnRNPs was associated with rapidly sedimenting (greater than 30 S) RNA (at 50 mM NaCl) suggesting their association by RNA-RNA hydrogen bonds. When the concentration of NaCl was 500 mM, 31% of the U1 RNA was associated with large RNA. Hence, approximately 30% of the U1 RNA molecules originally 'bound' (or about 15% of the total nuclear U1 RNA) were found to be associated by RNA-RNA hydrogen bonds while the remainder of the binding of U1 RNP to hnRNP was by protein-protein and/or protein-RNA interactions.

Animals↗
PubMed1984

Purification of p53/55 kinase from nuclear ribonucleoproteins of Namalwa cells.

A nuclear p53/55 protein kinase has been isolated from nuclear ribonucleoprotein particles from human tumor cells. The enzyme was purified approximately 2200-fold cell nuclei by sequential ribonuclease digestion of the RNP particles, DEAE cellulose and phosphocellulose chromatography. The kinase which was cAMP independent, catalyzed the phosphorylation of rabbit muscle glycogen synthase in the amino terminal domain, and conversion of the I to D form. The D synthase had a phosphorylation stoichiometry of 8 moles 32P per mole of synthase subunit with maximal specificity for ATP as phosphate donor; its Km was 30 microM. An antinucleolar antibody inhibited enzyme activity by 80%. Substrates for most other kinases were inactive. The kinase was essentially unaffected by the Walsh inhibitor, EGTA, regulatory subunits of protein kinase, calmodulin, trifluoperazine or heparin. Its activity was lost at 1 mM polyamine, but was enhanced 3-fold by MnCl2 and 4- to 9-fold by deoxymononucleotides. The nuclei of HeLa cells contained 64% of the total kinase of which 64% of the total kinase of which 11% were in nucleoli; the specific activity of the nucleolar kinase was twice that of the nuclear supernatant and four times that of the cytoplasmic kinase. These results indicate that nucleolar ribonucleoprotein particles of human tumor cells contain a cAMP-independent protein kinase which is similar to glycogen synthase kinase.

Burkitt Lymphoma↗
PubMed1984

Silver staining, immunofluorescence, and immunoelectron microscopic localization of nucleolar phosphoproteins B23 and C23.

Nucleolar organizer region (NOR)-specific silver staining and immunolocalization of nucleolar phosphoproteins B23 and C23 were compared in Novikoff hepatoma ascites cells. Silver staining and protein C23 immunostaining were both localized in the fibrillar shell surrounding the fibrillar center and in the fibrillar center. During mitosis, silver staining and protein C23 were localized at the NORs. Therefore, protein C23 and the silver-staining protein both seem to be associated with rDNA-containing structures (Mirre and Stahl 1981). A comparison of toluidine blue staining specific for RNA and B23 immunostaining demonstrated that protein B23 was associated with RNA-containing regions of the nucleolus and was absent from the fibrillar centers. Localization of these proteins and their functions are discussed in relation to the organization of the nucleolus.

Animals↗
PubMed1984

Onc genes and other new targets for cancer chemotherapy.

Recent advances in molecular biology have raised the hope that understanding of human cancer might progress rapidly and that improvements in therapy might result (Bishop 1983a, b; Busch 1962; Busch 1976; Duesberg 1983). With the development of gene cloning, DNA sequence analysis and improved hybridization methods, it became possible to evaluate whether cancer results from alteration in gene dosage, point or multiple mutation of genes, translocations, deletions, insertions, inversions, cis or trans altered promoters, amplification, and a variety of other genetic factors, including enhancer elements that alter rates of readouts of particular mRNA species. "Onc genes" are under intensive study because they offer manageable probes for evaluation of these various possibilities and also because the study of their cellular analogs may further understanding of the molecular biology of normal fetal and malignant cells. Despite the excessive enthusiasm of some proponents of this field and the negativism of its critics (Bishop 1983 a, b; Duesberg 1983), it is clear that analytical tools and new information will be of value in further studies on experimental cancer, regardless of whether cellular oncogenes (c-onc genes) have anything to do with human cancer or not. In the meantime, studies on enzymes, proteins and epitopes involved in growth processes, have opened new avenues for inhibition of human cancer by quantitative reduction of biosynthetic reactions.

Animals↗
PubMed1984

Immunofluorescence studies on proteins B23 and C23 in nucleoli of human lymphocytes.

Nucleoli of normal and leukemic lymphocytes were studied by cytochemical and immunofluorescence methods to provide more information on the nucleolar presence and distribution of proteins B23 and C23. Annular nucleoli of human lymphocytes represent a very convenient subject for such studies, since they consist of one centrally located large fibrillar center surrounded by RNP components. In such nucleoli, protein C23 was present mainly in the central nucleolar region and protein B23 was found mostly in the periphery. The nucleolar area immunostained for protein B23 was usually larger than that stained for protein C23. The distribution of protein C23 appeared to be similar to that of intensely stained nucleolar argyrophilic components. No substantial differences were found between the distribution of proteins B23 and C23 in nucleoli of normal and leukemic lymphocytes. In lymphocytes of patients treated with chemotherapy, the immunofluorescence was diminished for protein B23 and particularly so for protein C23.

Cell Nucleolus↗
PubMed1984

Further evidence that phosphoprotein C23 (110 kD/pI 5.1) is the nucleolar silver staining protein.

Previously we demonstrated a similar distribution between nucleolar organizing region-(NOR)-specific silver staining and localization of nucleolar phosphoprotein C23 (MW 110 kD/pI 5.1) [1, 2]. We now report that under fixation conditions which allow for antibody binding and subsequent silver staining, monoclonal antibody against protein C23 blocks NOR silver staining as well as silver staining in interphase nucleoli. Monoclonal antibody against nucleolar phosphoprotein B23 (MW 37 kD/pI 5.1) did not block silver staining in either NORs or interphase nucleoli. These, along with earlier observations, provide evidence that nucleolar phosphoprotein C23 is the major silver staining protein of the nucleolus and that it is directly or indirectly associated with rDNA.

Animals↗
PubMed1984

Further studies on satellite nucleoli in rat and mouse hepatocytes.

To provide more information on satellite nucleoli, these nuclear structures were studied by means of cytochemical and immunofluorescence procedures in rat and mouse hepatocytes without and following experimental inhibition of the RNA synthesis. The immuno-staining specific for nucleoli or B23 as well as C23 proteins demonstrated that satellite nucleoli and characteristic nucleoli exhibit the same fluorescence. The number of satellite nucleoli decreased after inhibition of nucleolar RNA synthesis in a similar way to the number of silver-stained granules (SSGs) of characteristic nucleoli. Inhibition of RNA synthesis also reduced the number of hepatocytes containing satellite nucleoli. Thus, satellite nucleoli seem to be real nucleoli from single NORs which did not fuse in the formation of a characteristic nucleolus.

Animals↗
PubMed1983

Isolation and partial characterization of dinoflagellate U1-U6 small RNAs homologous to rat U small nuclear RNAs.

The dinoflagellates are a group of diverse eukaryotic algae possessing a number of unique cellular properties. Evidence is presented for the presence of six capped small nuclear RNAs in these dinoflagellates. By several criteria such as the (a) presence of trimethylguanosine cap structure in U1 to U5 RNAs, (b) sequence homology between rat and dinoflagellate U2, U5 and U6 RNAs, (c) presence of other post-transcriptional modifications such as sugar and base modifications, and (d) association of Sm antigen with five of these six RNAs, the six RNAs of dinoflagellates appear to be similar to the well characterized U1 to U6 RNAs found in higher eukaryotes. This is the first demonstration of antigenic small nuclear RNA-containing particles in any unicellular organism. These results suggest that the U1 to U6 RNAs and the associated Sm antigen evolved at a very early stage of eukaryotic evolution. With respect to U small nuclear RNAs and their associated proteins, the dinoflagellates appear to exhibit eukaryotic characteristics.

Animals↗
PubMed1983

Identification of a La protein binding site in a RNA polymerase III transcript (4.5 I RNA).

Anti-La antibodies are frequently found in patients with autoimmune diseases; the antigen was reported to be a 50,000-Da protein (Rinke, J., and Steitz, J. A. (1982) Cell 29, 149-159). Because this protein was associated with many nascent RNA polymerase III transcripts, it was suggested to be an RNA polymerase III transcription factor. The present study was designed to analyze 4.5 I ribonucleoprotein, an RNA polymerase III transcript which contains the La antigen. It was found that the 3'-end 20-30-nucleotide portion was the most protected portion of 4.5 I RNA when 4.5 I ribonucleoprotein was digested with T1 RNase. When U2 RNA (an RNA polymerase II transcript) and 4.5 I RNA were incubated with the S-100 fraction of Novikoff hepatoma cells, the 4.5 I RNA bound La antigen but the U2 RNA did not. When partial and complete T1 RNase digestion fragments of 4.5 I RNA were incubated with the S-100 fraction, the 3'-end fragments bound preferentially to the La antigen. However, the fragments of 4.5 I RNA bound less efficiently to La antigen than whole 4.5 I RNA. These results indicate that the 3'-end of 4.5 I RNA is the La antigen binding site in this molecule and suggest that the overall conformation of RNA aids in the binding of La antigen.

Animals↗
PubMed1983

Phosphorylation of purified Novikoff hepatoma topoisomerase I.

The purified Novikoff hepatoma nuclear phosphoprotein with a molecular weight of 110 kdalton and pI 8.4, was found to be a type I topoisomerase. When isolated from 32P-labeled Novikoff ascites cells or incubated in vitro with protein kinase, phosphoserine was found to be its major phosphorylated amino acid. The enzymatic activity of topoisomerase I was altered by changes in phosphorylation. Its activity was increased by protein kinase and it was decreased by alkaline phosphatase.

Alkaline Phosphatase↗
PubMed1983

Detection of a nucleolar 7-2 ribonucleoprotein and a cytoplasmic 8-2 ribonucleoprotein with autoantibodies from patients with scleroderma.

In studies on antinucleolar antibodies in sera from 24 patients with scleroderma, an autoimmune disease, one serum, designated "anti-To", contained antibodies against a nucleolar 7-2 ribonucleoprotein and a novel cytoplasmic 8-2 ribonucleoprotein. The 7-2 and 8-2 RNAs are distinct RNAs with a pppG terminus. They are partially conserved between rat and human species and are present in distinct ribonucleoprotein particles. Eight sera contained antibodies that precipitated particles containing nucleolar U3 RNA; these antibodies appear to be directed against preribosomal particles containing U3 ribonucleoprotein, rather than the U3 ribonucleoprotein particles alone. All these ribonucleoproteins required proteins for antigenicity. These antibodies will be of use in studies on the structure and function of these novel small ribonucleoproteins.

Animals↗
PubMed1983

The nucleotide sequence of 8 S RNA bound to preribosomal RNA of Novikoff hepatoma. The 5'-end of 8 S RNA is 5.8 S RNA.

8 S RNA of Novikoff hepatoma was characterized by fingerprinting, sequencing gels, and by hybridization to rat ribosomal DNA clones. The data obtained show that 8 S RNA is 273 or 274 nucleotides long; ribosomal 5.8 S RNA is its 5'-terminal 156 nucleotides. All the post-transcriptional modifications found in 5.8 S rRNA were also found in 8 S RNA; no other modifications were found. The 3'-terminal 118 nucleotides were consistent with the adjoining internal transcribed spacer sequence in rDNA (Subrahmanyam, C. S., Cassidy, B., Busch, H., and Rothblum, L. (1982) Nucleic Acids Res. 10, 3667-3680). Based on its nucleolar localization, the finding that all the 8 S RNA is hydrogen-bonded to preribosomal RNA and its consistency in sequence to the cloned rat ribosomal DNA sequence, it appears that 8 S RNA is a relatively stable intermediate in the formation of 5.8 S rRNA from 45 S pre-rRNA. This stable intermediate RNA may be a useful substrate for studies on rRNA processing and for studies on eukaryotic rRNA-processing enzyme(s).

Animals↗
PubMed1983

Proliferating cell nuclear antigen (PCNA) and human malignant tumor nucleolar antigens (HMTNA) in nucleoli of human hematological malignancies.

Lymphoma (Lymphocytic non-Hodgkin's malignant lymphoma) and leukemic (chronic lymphocytic, acute and chronic myeloid, myelomonocytic leukemia) cells were studied by indirect immunofluorescence to evaluate the presence of proliferating cell nuclear antigen (PCNA) and human malignant tumor nuclear antigen (HMTNA) in their nucleoli. Most cells in lymph node smears of lymphocytic non-Hodgkin's malignant lymphoma (NHML) developed a bright nucleolar fluorescence with HMTNA antibodies. PCNA was detected in nucleoli of a limited number of cells which apparently represent the proliferating cell population in these lymphomas. Similarly, in the bone marrow smears of patients with chronic lymphocytic leukemia most cells possessed a nucleolar fluorescence for HMTNA and PCNA was present in nucleoli of a limited number of cells. In the bone marrow smears of patients with myeloid or myelomonocytic leukemias most blastic or monocytoid cells also developed a bright nucleolar fluorescence with HMTNA antibodies and PCNA was present only in a small percentage of these cells. Leukemic cells with PCNA in their nucleoli like thekhuntigen might represent a proliferating cell population in late G1-early S phase.

Antigens, Neoplasm↗
PubMed1983

Identification and purification of Namalwa nuclear RNP antigen 52/5.3.

Using affinity purified rabbit antibodies to HeLa nucleoli and the Western blotting techniques, an antigen with an approximate molecular weight of 52,000 and pI of 5.3 was found in Namalwa cells (a Burkitt lymphoma), but not in normal liver cells. This antigen was purified from Namalwa RNP by column chromatography on Sephacryl S-200, hydroxylapatite and one-dimensional SDS gel electrophoresis. A liver protein with the same molecular weight and pI value was purified from RNP fraction by one-dimensional SDS gel electrophoresis. Both proteins had similar amino-acid compositions. The tryptic map of 125I-labeled protein 52/5.3 contained approximately nine major spots; spot 9 was present in the Namalwa protein but not in the liver protein. The similarity of the structures of these proteins and their differences in antigenicity are noteworthy and require further structural and functional analysis.

Amino Acids↗
PubMed1983

Localization of nucleolar phosphoproteins B23 and C23 during mitosis.

Nucleolar phosphoproteins B23 and C23 were simultaneously localized in unsynchronized male rat-kangaroo PtK2 cells during mitosis using a mouse monoclonal antibody against protein B23 and a rabbit antibody against protein C23. The distribution of proteins B23 and C23 during mitosis was compared with the distribution of the silver staining protein. During interphase, proteins B23 and C23 were both localized to the nucleolus. As the nucleolus disappeared in prophase, the distribution of protein B23 became nucleoplasmic, whereas most of protein C23 remained associated with the disappearing nucleolus. Throughout metaphase and anaphase protein B23 was found associated with the chromosomes, whereas protein C23 seemed to disappear. When the nucleolus reformed during telophase, protein C23 appeared first in 'prenucleolar bodies' and then in the nucleolus, whereas protein B23 did not appear in the nucleolus until late telophase or early G1 phase. Silver staining during mitosis closely paralleled the distribution of protein C23, supporting previous conclusions that protein C23 is a silver staining nucleolus organizer region (NOR) protein [19, 20].

Animals↗
PubMed1983

Elevated levels of a 17-kilodalton nuclear protein in carcinomas.

The presence of a 17-kilodalton nuclear protein with pI variants of 6.7-6.9 in solid human malignant tumors is reported. This protein was more prominent in the carcinomas sampled compared to the sarcomas studied. Small amounts were detectable in placenta, but not in adult human liver, suggesting that the protein may be the product of a gene involved in normal differentiation.

Adenocarcinoma↗