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Biomedical subjects

H Busch

Publications and source records attributed to H Busch.

At least 181 records · Page 10Linked to original sources
PubMed1983

Ultrastructural and purification studies on human tumor nucleolar antigens and nucleolar particles.

The presence of common nucleolar antigens in a broad array of human malignant tumors has led to several lines of investigations. In addition to studies on an increasing number of benign and malignant neoplasms with a variety of antibodies designed to statistically evaluate the presence of nucleolar antigens, purification procedures and chemical analyses are being used to characterize the specific antigens. The localization of the nucleolar antigens in HeLa cells was studied by the postembedding immunoelectron microscopic procedure employing rabbit antibodies to nucleoli or nuclear Tris extracts of these cells. The products of the peroxidase-antiperoxidase complexes visualized by the reaction with diaminobenzidine in nucleoli were mainly found in the nucleolonemas which contain the dense nucleolar RNP components. When these nucleoli became compact after treatment of HeLa cells with adriamycin, the distribution of the immunoreactive products was altered along with distribution of the dense nucleolar components. The human nucleolar antigens were mainly localized to nucleolar regions containing the nucleolar RNP components. Improved purification of the antigens made it possible to provide a satisfactory amino acid analysis of one pI 6.3 antigen. Interestingly, some of the nucleolar antigen was found in miniparticle undescribed until now.

Amino Acids↗
PubMed1983

A 38,000-dalton antigen found in Namalwa cells induced by Newcastle disease virus.

An antigen has been isolated from Namalwa cells, a Burkitt lymphoma line, that was induced by Newcastle disease virus (NDV) for interferon production. The antigen was extracted by 3 M NaCl from ribonucleoprotein particles (RNP), obtained from the nuclear 0.01 M Tris extract, and was purified by hydroxylapatite chromatography, phosphocellulose chromatography, and preparative sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Its molecular weight was 38 kilodalton (kDa) as determined by SDS-PAGE. The tryptic peptide map of 125I-labeled antigen contained seven major peptides. The antigen was not found in HeLa cells, normal human liver or in Namalwa cells that had not been induced by the virus. This result suggests that this antigen was produced in Namalwa cells as a result of induction by the NDV virus.

Animals↗
PubMed1983

Indirect immunofluorescence studies of proliferating cell nuclear antigen in nucleoli of human tumor and normal tissues.

A proliferating cell nuclear antigen (PCNA) was identified with autoantibodies from a patient with systemic lupus erythematosus. Specific antibodies were purified by affinity chromatography in which Novikoff hepatoma nucleolar proteins were conjugated to Sepharose-4B. The purified anti-PCNA antibodies produced bright nucleolar fluorescence in tumor cells as shown by indirect immunofluorescence. PCNA was found in nucleoli of human cell lines, HeLa, Hep-2, and Namalwa, and a solid human renal and a prostate carcinoma. Both strong and weak nucleolar fluorescence areas were found in the renal and prostate carcinoma indicating that there are varying degrees of proliferation among tumor cells. Two human colon carcinoma cell lines, omega (an aggressive, fast-growing clone of human colon carcinoma cell line HCT 116) and CBS [a slow-growing human colon carcinoma cell line (group 3)], with different growth rates were compared. The fast-growing colon carcinoma cells, omega, exhibited a higher percentage of nucleolar fluorescence (28.5%) than that of the slow growing colon cells (13.6%). By enzyme-linked immunosorbent assays, the omega cell extract had a higher PCNA antigen content (2.8-fold) than that of the CBS cell extract which, in turn, was higher than that of human liver extract. PCNA was also found in a human fetal lung fibroblast cell line (IMR-90). Very weak or negative nucleolar fluorescence was observed in several normal human tissues including liver, kidney, prostate, and cheek cells. Nucleolar fluorescence was also observed in rat Novikoff hepatoma cells. Although normal rat livers do not have PCNA nucleolar fluorescence, nuclear and nucleolar fluorescence were observed at 18 hr after partial hepatectomy.

Antigens↗
PubMed1983

Characterization of nucleolar antigens of normal rat liver, regenerating rat liver, and Novikoff ascites hepatoma cells following in vitro translation of polyadenylic acid-containing messenger RNA.

Nucleolar antigens of normal rat liver, regenerating liver, and Novikoff ascites hepatoma cells were transplanted in vitro from polyadenylic acid-containing messenger RNAs isolated from the respective tissues and immunoprecipitated with specific antinucleolar antibodies and Protein A. By two-dimensional gel electrophoresis of the translation products, five antigens were detected in normal rat liver. The antigens detected in 18-hr regenerating rat liver were the same as those of normal rat liver when immunoprecipitated with the anti-liver nucleolar antibodies. In the Novikoff tumor, 11 antigens were detected with anti-Novikoff nucleolar antibodies. Of these, two were not found in either normal or regenerating liver. Four major antigens were detectable in both Novikoff hepatoma and regenerating liver messenger RNA translation products with anti-Novikoff nucleolar antibodies. Two antigens were found in normal and regenerating liver that were not found in Novikoff hepatoma. In agreement with previous results, these immunoprecipitation analyses provide further evidence that some nucleolar antigens are present in Novikoff hepatoma that are not found in either normal or regenerating rat liver.

Animals↗
PubMed1982

Nucleolar specific acidic phosphoprotein C23 is highly methylated.

Protein C23 (Mr = 110,000; pI, 5.5) is the major phosphoprotein in the nucleolus of Novikoff hepatoma cells and comprises 9.5% of the total nucleolar protein. In addition to being highly phosphorylated (1.2 mol % of phosphoserine), it is also highly methylated. Protein C23 contains 1.3 mol % of NG,NG-dimethylarginine and a trace of NG-monomethylarginine.

Amino Acids↗
PubMed1982

Nucleotide sequence of the region between the 18S rRNA sequence and the 28S rRNA sequence of rat ribosomal DNA.

The DNA sequence of the intragenic region of the rat 45S ribosomal RNA precursor was determined. This sequence contains 2282 nucleotides and extends from the conserved EcoR I site near the 3' terminus of 18S rRNA to 69 nucleotides downstream of the 5' terminus of 28S rRNA. The sequences corresponding to 18S and 5.8S rRNA were identified by comparison with previously published data. The 5' terminus of rat 28S rRNA was identified by S1 nuclease protection and reverse transcriptase elongation assays. The internal transcribed spacers were found to be 1066 and 765 nucleotides long and had little homology with those of Xenopus and yeast. Regions of sequence homology between rat and Xenopus were found at the junctions of the internal transcribed spacers with 18S, 5.8S and 28S rRNA. These homologies suggest that these sequences may function as recognition sites for the processing of the ribosomal precursor RNA.

Animals↗
PubMed1982

Nucleotide sequence of 7 S RNA. Homology to Alu DNA and La 4.5 S RNA.

7 S RNA, a component of normal higher eukaryotic cells and several oncornaviruses, was shown to be conserved in evolution (Erikson, E., Erikson, R. L., Henry, B., and Pace, N. R. (1973) Virology 53, 40-46). Recently, 7 S RNA was shown to be partially complementary to Alu family DNA sequences (Weiner, A. (1980) Cell 22, 209-218). In the present study the nucleotide sequence of Novikoff hepatoma 7 S RNA was determined to be: (formula, see text) Comparison of 7 S RNA, Alu and B1 family DNA, and La 4.5 S RNA sequences for homologies showed that 1) one-third of 7 S RNA, mainly the 5'-end, was homologous to Alu and B1 family sequences; 2) one 300-nucleotide long Alu family sequence contained two binding sites for 7 S RNA; and 3) the 5'-ends of 7 S RNA and La 4.5 S RNA also had extensive (60%) homologies. A model for the secondary structure of 7 S RNA based on maximal base pairing and preferential nuclease cleavage sites is also presented.

Animals↗
PubMed1982

Donut-shaped "miniparticles" in nuclei of human and rat cells.

Donut-shaped "miniparticles" were extracted from nuclei of various types of human and rat cells. Electron-microscopic investigation showed these particles were predominantly in sucrose density gradient fractions that had an approximate sedimentation coefficient of 21S. These particles were 113 +/- 8A in diameter and had an electron dense center of 29 +/- 6A. They appeared to be composed of 8 subunits. Quantitative analysis of the number of these particles by electron-micrographic field counting showed nuclei of tumor samples had a larger amount of the particles than the cytosol. However, normal cell cytosol had a larger number of particles than the nuclei. A group of proteins in the 25,000-33,000 molecular weight range was shown to be the main protein component by two dimensional gel electrophoresis.

Animals↗
PubMed1982

Isolation and partial characterization of a nuclear antigen (68/6.3) from the Namalwa cell line (a Burkitt lymphoma).

A nuclear antigen was purified from the 0.01 M Tris-HCl/pH8 extract of nuclei of the Burkitt tumor Namalwa cell line to electrophoretic homogeneity by DEAE cellulose chromatography, affinity chromatography, and preparative isoelectric focusing. The yield of antigens was 0.02% of the nuclear 0.01 M Tris-HCl/pH8 extract. On two-dimensional gel electrophoresis, the major antigen separated into two adjacent protein spots with molecular weights of 68,000 and an approximate pI of 6.3 (68/6.3 A and 68/6.3 B). A minor antigen had a molecular weight of 61,000 and pI of 6.0 (61/6.0). Fourteen 125I-labeled peptides were obtained from the tryptic digest of the major antigen (68/6.3 A and 68/6.3 B). The amino-acid composition analysis of the purified antigens indicated that the amino acids in the highest content were glycine (15%), glutamic acid (11.6%), and serine (9%); the ratio of acidic to basic amino acids was 1.95. In studies on nucleolytic activity, the purified antigen produced a single-stranded and then a double-stranded cleavage of PM 2 and pBR 322 DNA. This antigen is the first purified nuclear antigen that reacts with the HeLa-specific nucleolar antibodies.

Amino Acids↗
PubMed1981

Ubiquitin - protein conjugates.

The data available at present indicates there are three distinct functions of ubiquitin, two of which are related to protein conjugation. The first of these has been extensively studied by our laboratory and others interested in nucleosomes and changes in chromatin states. The ubiquitin-histone (Ub-2A, Ub-2B) conjugation reaction now appears to be a very dynamic process. In the deconjugation (lyase) reaction, both the histone 2A and the ubiquitin are left intact and in a form which makes possible ready reconjugation. Accordingly, this may be a mechanism for "moment-to-moment" Control of the genome. The second function in which ubiquitin is conjugated involves proteolytic activity. This activity is correlated with protein turnover. In this process, the ubiquitin-protein conjugate apparently serves as a "signal" for the protease cleavage of the protein. The released ubiquitin is also intact and is probably available for reconjugation. In the third function, ubiquitin was suggested to serve as a "hormone". The studies thus far have been carried out primarily on induction of T- and B-lymphocytes, reduction or delay of Coombs' positivity and reduction of spleen weight. The precise physiological role of this reported function is still unclear, particularly because the ubiquitin used was probably not the physiologically active form.

Amino Acid Sequence↗
PubMed1981

Primary and secondary structure of U2 snRNA.

With the improved rapid sequencing techniques, the earlier sequence of U2 RNA of Novikoff hepatoma (Shibata et al, J. Biol. Chem. 250, 3909-3920, 1975) was reanalyzed and modified. The improved sequence of U2 RNA is 188 (or 189) nucleotides long and is in register with a characterized U2 RNA pseudogene (Denison et al, PNAS 78, 810-814, 1981) except for an 11 nucleotide sequence (nucleotides 147-157) which is absent from the pseudogene. From these results, a secondary structure of U2 RNA is proposed which is supported by the preferred cleavage sites with T1-RNase, RNase A and S1 nuclease. Isolated U2 RNA was cleaved by T1-RNase preferentially at positions 64 and 164, whereas U2 RNA in U2-snRNP was cleaved only at position 64, indicating that position 164 is protected in U2-snRNP. As with U1 RNA (Epstein et al, PNAS 78, 1562-1566, 1981) the 5'-end of isolated U2 RNA was not preferentially cleaved by T1-RNase.

Animals↗