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R K Busch

Publications and source records attributed to R K Busch.

65 records · Page 4Linked to original sources
PubMed1976

A fetal protein in chromatin of Novikoff hepatoma and Walker 256 carcinosarcoma tumors that is absent from normal and regenerating rat liver.

Antibodies to chromatin proteins of Novikoff hepatoma cells formed precipitin bands in the double-diffusion immunoprecipitation assay with chromatin proteins of Novikoff hepatoma, Walker 256 carcinosarcoma, and 18-day fetal rat liver. The antigen used for preparation of antiserum was the chromatin proteins initially extracted with 3 M NaCl-7 M urea and soluble after dialysis to 0.14 M NaCl-0.35 M urea. The chromatin proteins used for analytical studies were extracted with 0.6 M NaCl containing 0.01 M Tris-HCl (pH 8) and 100 muM phenylmethylsulfonyl fluoride. Corresponding chromatin proteins of normal and 18-hr regenerating rat liver, heart, and kidney did not form precipitin bands. The antigen was purified from the chrmatin of Novikoff hepatoma cells by exclusion chromatography on Sephadex G-150 and preparative nondenaturing polyacrylamide gel electrophoresis. Its migration on denaturing sodium dodecyl sulfate-polyacrylamide gels corresponded to a molecular weight of 26,000. Amino acid analysis showed that the ratio of acidic to basic amino acids was 1.4 to 1.0. Evidence for its homogeneity included its migration as a single protein spot on two-dimensional polyacrylamide gel electrophoresis and its single lysine amino-terminal amino acid. This protein is a glycoprotein, as shown by the presence of 15 moles of galactosamine per mole of antigen. These studies demonstrate the presence of a fetal glycoprotein in the chromatin of two tumors that may have an important role in determining their gene products.

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PubMed1976

Homochromatographic and immunological analysis of controls of nucleolar gene function.

To compare regulation of nucleolar function of tumors and other tissues, it was necessary to develop assays of the fidelity of ribosomal DNA readouts. For this purpose, homochromatography analyses of complete T1 ribonuclease digestion products of the in vivo labeled 45 S preribosomal RNA were compared with those of 18S and of 28 S ribosomal RNA. Homochromatography analysis of the in vitro readout product of isolated nucleoli showed the presence of many large marker nucleotides of the in vivo 45 S preribosomal RNA. Moreover, no other large oligonucleotides were detected. The in vitro readout product of nucleolar chromatin had the same T1 ribonuclease digestion products, including the large marker of oligonucleotides. However, the in vitro readout product of nucleolar DNA contained no large marker T1 ribonuclease oligonucleotides. These results indicate that the fidelity of nucleolar readouts is controlled by regulatory proteins of the nucleolar chromatin. Differences were found in nucleolar proteins of normal rat liver and Novikoff hepatoma by immunological analyses. The possibility exists that differences in readout rates of tumor and other nucleoli are related to the protein difference detected by these immunological studies.

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PubMed1976

Antigenically active nonhistone chromatin proteins in cancer cells.

Two-dimensional polyacrylamide gel electrophoresis shows that in nuclei of Novikoff hepatoma ascites cells there are approximately 75 proteins in the chromatin fraction soluble in 3 M NaCl:7 M urea. Dialysis of this fraction to an ionic strength of 0.15 produces a soluble fraction and a precipitate. The proteins in the soluble fraction have been reported to be active in gene control. Antibodies to the soluble fraction distribute diffusely throughout the nucleus, and antibodies to the precipitate localized primarily in the nucleolus and the nuclear ribonucleoprotein network. The nucleolar proteins differ from the extranucleolar proteins in antigenicity and labeling patterns. The development of methods for isolation, purification, and identification of nuclear proteins provided the opportunity for analysis of chromatin antigens in tumor cells. Utilizing two-dimensional preparative polyacrylamide gel techniques as well as conventional procedures, several nuclear proteins have been isolated in electrophoretically homogeneous states including protein A-24, a histone-like nonhistone protein; C-14, a protein that stimulates nucleolar RNA polymerase; and a chromatin antigen soluble in 3 M NaCl:7 M urea that remains soluble after dialysis to 0.15 M NaCl to precipitate the histones and the DNA. This antigen has been found in the chromatin of both the Novikoff hepatoma and the Walker 256 carcinosarcoma but not in the chromatin of either normal or regenerating liver. It is a nonhistone nuclear protein as indicated by its amino acid analysis in which the ratio of the number of acidic to basic amino acids is approximately 1.4. Further studies are in progress on the function and structure of this chromatin protein. As an approach to analysis of relative rates of synthesis of this antigen and otherproteins, the products of translation of messenger RNA of Novikoff hepatoma and normal liver are being analyzed by autoradiography of two-dimensional electrophoretic gels.

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PubMedDate not supplied

Antigenic proteins of nucleolar chromatin of Novikoff hepatoma ascites cells.

Nucleolar chromatin of Novikoff hepatoma ascites cells contains an antigen (no-Ag1) detected with antinucleolar antibodies by the immunodiffusion technique. This antigen was distinguished from the previously reported nuclear chromatin antigen NAg-1 (19) by the findings that tumor nucleolar antibodies which formed immunoprecipitin bands with no-Ag1 did not do so with NAg-1 and that tumor cytosol, which contains NAg-1, formed immunoprecipitin bands with tumor chromatin antibodies but not with antibodies to tumor nucleoli. Tumor nucleolar chromatin contains both NAg-1 and no-Ag1, but only no-Ag1 formed bands with tumor nucleolar antibodies, no-Ag1 is a component of tumor nucleolar chromatin that was not soluble in 0.075 M NaCl-0.025 M EDTA, pH 8, and only slightly soluble in 0.01 M Tris-HCl, pH 8. NO-Ag1 was not found in liver nucleoli. Antibodies to liver nucleoli formed immunoprecipitin bands with liver nucleolar antigens but none were confluent with those formed between tumor nucleolar antibodies and antigens of tumor nucleolar chromatin. Absorption of the tumor nucleolar antibodies with whole tumor cells or whole liver pressate did not alter band formation with no-Ag1. Three antigens in liver nucleoli were not found in tumor nucleoli.

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