Proteins with sulfhydryl groups in nuclei and nuclear matrix of normal and tumour cells.
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Animals↗
Biomedical subjects
Z Kiliańska
Publications and source records attributed to Z Kiliańska.
Nuclear proteins of hamster hepatoma after administration of antitumour agents.
One- and two-dimensional gel electrophoretic analysis of nuclear proteins of Kirkman-Robbins hepatoma was used to study the effects of the tumour growth inhibitors methotrexate (MTX) and acyclonucleoside (DHPQtB) on protein composition. MTX and DHPQtB inhibited Kirkman-Robbins hepatoma growth by 89.2 +/- 3.5% and 16.3 +/- 6.1% respectively. The biosynthesis and/or metabolism of some polypeptide spots was affected by these antitumour agents, especially among components with molecular wt/isoelectric points of 52,000-64,000/4.9-5.5, 69,000-78,000/5.0-5.9 and 88,000-100,000/5.1-5.9.
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Studies on low molecular weight nuclear protein of tumour and normal cells.
1. Preliminary results of comparative electrophoretical and immunological analyses of the components of two classes of non-histone proteins, i.e. NHP1 and NHP2 eluted from hydroxyapatite allowed to suppose that protein of Mw 18,000 is specific for animal tumour cells. 2. However, the studies on cellular distribution of this polypeptide indicated that it is exclusively located in nuclei of hepatoma and normal liver as well. 3. The former observation seems to be the result of changes of the affinity of this protein to DNA during neoplastic transformation.
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Hamster liver chromatin immunospecific non-histone proteins.
Three antisera were prepared against non-histone protein classes named NHCP1, NHCP2 and dehistonized chromatin (with different affinity to DNA) from hamster liver. Two main antigenic bands of MW 17,000 and 36,000 were specific in the NHCP1 fraction and one antigen of MW 56,000 was specific for the NHCP2 fraction from nuclease-sensitive and especially nuclease-resistant chromatin. Other NHCP2 liver antigens of MW 22,000, 27,000, 30,000, 36,000, 37,000, 40,000, 45,000, 46,000, 51,000, 98,000 and 100,000 were present only in nuclease-resistant chromatin of hamster liver. Immunologically specific hamster liver non-histone proteins within the NHCP1 and NHCP2 fractions seem to be restricted to nuclease-resistant chromatin fraction of this tissue. The above mentioned liver specific antigens are absent or present only at trace amounts in analogous Kirkman-Robbins hepatoma fractions.
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Identification of a nuclear antigen with molecular weight of 48,000 differentially expressed in tumour and normal cells.
A non-histone protein with mol. wt of 48,000 differentially expressed in normal and tumour cells was identified using immunological criteria. Antibodies were raised against a component specific for Kirkman-Robbins hepatoma of mol. wt about 48,000 separated from hepatoma non-histone proteins by preparative electrophoresis in polyacrylamide gel. It was demonstrated by immunoblotting that Morris hepatoma 7777 and Ehrlich ascites cells share an antigenic non-histone protein with Kirman-Robbins hepatoma. Tumour cells when compared with normal cells, i.e. hamster and rat liver, are characterized by significant enrichment of this component. Intracellular distribution of the polypeptide with mol. wt 48,000 suggests that this component may be a structural protein the biosynthesis of which increases or the antigenic determinants of which change in tumour cells.
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Molecular characterization of non-histone chromatin proteins from experimental tumours.
Using two-dimensional (2-D) electrophoresis, two non-histone chromatin protein fractions (NHCP1 and NHCP2) from three animal tumours (Kirkman-Robbins hepatoma, Morris hepatoma 7777 and Ehrlich ascites cells) and normal hamster liver were analyzed. Apart from many common components several tissue specific polypeptides of the NHCP1 and NHCP2 fractions were detected. It was found that some spots present in electropherograms of non-histone proteins of tumour cells (M X 10(-3)/pI): 17-24/4.9-6.5 (NHCP1 and NHCP2); 34-41/4.9-6.0 (HCP1 and NHCP2); 44-46/5.3-7.5 (HCP2); 46-49/5.0-7.5 (NHCP1); 49/5.9-7.5 (NHCP2) and 102-134/5.6-7.0 (NHCP1) were absent from normal liver.
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Specificity of non-histone protein antigens in chicken liver nuclei.
1. Chicken liver nuclei were fractionated by disruption with ultrasound and subsequent precipitation with divatent cations. A small, protein-rich fraction (CS), representing less than 5% of the total nuclear DNA reacted strongly with antisera to dehistonized chicken liver chromatin. 2. Further fractionation of the CS by DNA-affinity chromatography yielded two DNA-binding immunoreactive proteins (approximate Mr 35 and 56 kD). 3. We have shown previously (Kiliańska et al., 1981, Kiliańska, 1984) that this DNA-binding component strongly inhibited the in vitro transcription of RNA.
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Molecular and functional diversity of non-histone protein fraction NHCP1 from hamster Kirkman-Robbins hepatoma and liver.
Non-histone protein fraction NHCP1 of micrococcal nuclease-sensitive and nuclease-resistant chromatin from Kirkman-Robbins hepatoma and hamster liver was studied by two-dimensional electrophoresis followed by Coomassie and silver staining and by microcomplement fixation technique in the presence of antibodies elicited against NHCP1 of both tissues. Apart from many common spots several tissue specific components associated with either nuclease-sensitive or nuclease-resistant chromatin were found. The presence of tissue specific components among NHCP1 from hepatoma and liver was confirmed by immunological analysis. It was stated that these components are exclusively localized in nuclease-resistant part of chromatin from neoplastic and normal tissues thus suggesting their structural function.
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Specificity of Kirkman-Robbins hepatoma non-histone chromatin proteins: electrophoretic and immunological analyses.
The specificity of Kirkman-Robbins hepatoma and hamster liver non-histone chromatin proteins has been studied by comparing polypeptide patterns in polyacrylamide gel electrophoresis and by their immunological activity in the complement fixation test. Non-histone proteins were separated from DNA with a polyethylene glycol-dextran mixture and fractionated by hydroxylapatite chromatography into three classes named NHCP1, NHCP2, and NHCP3. Electrophoretic analysis indicated that among the non-histone proteins of Kirkman-Robbins hepatoma and hamster liver differences mainly of a quantitative nature can be observed. However, the polypeptides with molecular weight 25 000, 31 000, 36 000, 73 000 in NHCP1; 20 000, 40 000 in NHCP2 and 20 000, 23 000, 32 000, 38 000, 44 000, 75 000, 80 000 in NHCP3 were found to be specific for hepatoma chromatin. Application of antibodies against NHCP1, NHCP2 and dehistonized chromatin of Kirkman-Robbins hepatoma revealed that the highest specificity of NHCP2 eluted from hydroxylapatite with 100 mM phosphate buffer at pH 6.8. The NHCP1 of hepatoma shares some common antigenic determinants with analogous proteins of liver. On the other hand non-histone proteins specific for hepatoma dehistonized chromatin can be localized in the NHCP3 and partially in the NHCP1 fractions.
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Comparison of non-histone proteins from hamster Kirkman-Robbins hepatoma and liver.
Three classes of non-histone proteins were obtained from hamster Kirkman-Robbins hepatoma and liver nuclei following separation of nucleic acids with the polyethylene glycol-dextran mixture and fractionation of nuclear proteins on hydroxylapatite in a salt-glycerol-phenylmethylsulphonyl fluoride system at increasing concentrations of Na+ and K+ phosphate buffer, pH 6.8. Two-dimensional polyacrylamide gel electrophoresis of these proteins documented their high heterogeneity; many spots were common but some spots specific only for neoplastic or normal tissue were also observed.
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Distribution of non-histone proteins between micrococcal nuclease sensitive and nuclease resistant chromatin from chicken cells with active and inactive genomes.
Chicken liver and erythrocyte nuclei were separated by mild treatment with micrococcal nuclease into nuclease sensitive (NS) and nuclease resistant (NR) fractions, differing in chemical composition and transcriptional activity in vitro. Nuclei, NS and NR fractions of both tissues were fractionated by hydroxyapatite chromatography into three groups of non-histone chromatin proteins (NHCP) and characterized by SDS-polyacrylamide gel electrophoresis. Some differences in the molecular distribution of non-histone proteins of chicken liver and erythrocytes between nuclease sensitive and resistant parts of chromatin have been described.
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Chicken liver non-histone protein fraction repressing DNA transcription in vitro.
A chicken liver non-histone protein fraction repressing DNA transcription in vitro was isolated using ultrasound shearing followed by precipitation with divalent cations. It was characterized by SDS-polyacrylamide electrophoresis and isoelectric focusing. This inhibitory protein fraction showed strong affinity to DNA and represented non-histone proteins with main bands of mol. wt about 35000 and 56000 and isoelectric points about 5.7 and 6.2. The inhibitory non-histone protein fraction seems to repress DNA transcription by interacting with DNA template rather than with RNA polymerase.
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Chromatin proteins associated with micrococcal nuclease-sensitive and nuclease-resistant chromatin fractions of Kirkman-Robbins hepatoma and hamster liver.
Micrococcal nuclease-sensitive (SP) and nuclease-resistant (PP) chromatin fractions from Kirkman-Robbins hepatoma and hamster liver were obtained. The molecular distribution of three non-histone proteins (NHCP1, NHCP2 and NHCP3), histones, and chromatin-bound protease activity between SP and PP fractions of both tissues was compared. Differences, mainly of quantitative nature, among non-histone proteins of neoplastic and normal tissue were observed. Moreover, it was found that polypeptides with mol. wt 81 000 (NHCP1), 39 000 (NHCP2) and 21 000, 35 000, 37 000 (NHCP1), 70 000, 112 000, 141 000, 157 000 (NHCP2), 30 000-33 000 (NHCP3) were associated only with the nuclease-sensitive part of chromatin of hepatoma and normal tissue, respectively. A major difference in histone composition of hamster hepatoma and liver concerns histones H2A and H1. Furthermore, an enrichment of high mobility group proteins as well as other soluble non-histone proteins in an acid extract of the SP fraction was observed. Apparently chromatin-bound protease activity can be found in both fractions of chromatin.
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Distribution of chromatin proteins between fractions of hamster liver chromatin differing in their susceptibility to micrococcal nuclease.
Hamster liver nuclei were fractionated by digestion with micrococcal nuclease into nuclease released (SP) and nuclease resistant (PP) fractions varying in chemical composition and transcriptional activity. Electrophoretic analysis of histones from SP and PP showed no qualitative and quantitative differences. Apparently chromatin-bound protease activity can be found in both fractions. Nonhistone chromatin proteins isolated from SP and PP under mild conditions were fractionated by hydroxyapatite chromatography into NHCP1, NHCP2, NHCP3 and molecular heterogeneity and specificity were tested by SDS-polyacrylamide gel electrophoresis. The differences observed in nonhistone proteins are mainly of quantitative nature, however some specific polypeptides for SP and PP are observed.
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Changes in non-histone chromatin proteins during the storage of chromatin.
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Animals↗
An electrophoretic comparison of non-histone proteins from rat liver total chromatin and chromatin depleted of 0.35 M NaCl soluble proteins.
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Animals↗
Characterization of nuclear non-histone proteins from rat liver isolated in the polyethylene glycol-dextran system and separated by hydroxyapatite chromatography.
1. Three groups of non-histone proteins were obtained from rat liver nuclei by separation of nucleic acids with the polyethylene glycol - dextran polymer mixture and fractionation of nuclear proteins on hydroxyapatite in the salt - glycerol - phenylmethylsulphonyl fluoride system at increasing concentration of Na+ and K+ phosphate buffer, pH 6.8. 2. Non-histone proteins of rat liver represent a heterogeneous complex of polypeptides devoid of DNA, of molecular weight ranging from 12000 to 130000, and isoelectric points from pH 4.0 to 7.8.
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Specificity of non-histone proteins from hamster Kirkman-Robbins hepatoma.
Nuclear phosphoproteins from Syrian hamster liver and Kirkman-Robbins hepatoma were obtained by phenol method of TENG et al. and their chemical composition was investigated. Amino acid composition of phosphoproteins from both tissues resembled each other to a great extent. There was almost twice more phosphorus in the preparations from hepatoma than in the ones from liver. Using two electrophoretic techniques: SDS-polyacrylamide gel electrophoresis and isoelectric focusing some differences between the examined proteins of neoplastic and normal tissue were found. Three additional fractions (molecular weights 29 000, 89 000 and 93 000, respectively) could be observed in case of hepatoma proteins in comparison with the ones from liver. The phosphoproteins of Kirkman-Robbins hepatoma revealed three additional bands with isoelectric points 6.6, and 7.1.
Amino Acids↗