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Johannes Gerdes - One of the best experts on this subject based on the ideXlab platform.
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cell proliferation associated nuclear antigen defined by antibody ki 67 a new kind of cell cycle maintaining Proteins
Archivum Immunologiae Et Therapiae Experimentalis, 1995Co-Authors: M Duchrow, H D Flad, Claudia Wohlenberg, C Schluter, G Key, M H G Kubbutat, Johannes GerdesAbstract:Abstract A decade of studies on the human nuclear antigen defined by monoclonal antibody Ki-67 (the "Ki-67 Protein") has made it abundantly clear that this structure is strictly associated with human cell proliferation and that the expression of this Protein can be used to assess the growth fraction of a given cell population. Until recently the Ki-67 Protein was described as a Nonhistone Protein that is highly susceptible to protease treatment. We have isolated and sequenced cDNAs encoding for this antigen and found two isoforms of the full length cDNA of 11.5 and 12.5 kb, respectively, sequence and structure of which are thus far unique. The gene encoding the Ki-67 Protein is organized in 15 exons and is localized on chromosome 10. The center of this gene is formed by an extraordinary 6845 bp exon containing 16 successively repeated homologous segments of 366 bp ("Ki-67 repeats"), each containing a highly conserved new motif of 66 bp ("Ki-67 motif"). The deduced peptide sequence of this central exon possess 10 ProGluSerThr (PEST) motifs which are associated with high turnover Proteins such as other cell cycle-related Proteins, oncogenes and transcription factors, etc. Like the latter Proteins the Ki-67 antigen plays a pivotal role in maintaining cell proliferation because Ki-67 Protein antisense oligonucleotides significantly inhibit 3H-thymidine incorporation in permanent human tumor cell lines in a dose-dependent manner.
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immunobiochemical and molecular biologic characterization of the cell proliferation associated nuclear antigen that is defined by monoclonal antibody ki 67
American Journal of Pathology, 1991Co-Authors: Johannes Gerdes, M Duchrow, Li Li, C Schlueter, Claudia Wohlenberg, C Gerlach, I Stahmer, Sabine Kloth, Ernst Brandt, H D FladAbstract:The monoclonal antibody Ki-67 detects a human nuclear antigen that is present in proliferating cells, but absent in quiescent cells. The aim of this study was to characterize the Ki-67 antigen by means of immunobiochemical and molecular biology techniques. Enzymatic digestion experiments showed that this antigen is highly susceptible to protease treatment, and the antigen cannot be extracted by 0.1 normal HCl, indicating that Ki-67 antigen is a Nonhistone Protein. Immunoblot analysis of cell lysates with Ki-67 showed a double band with apparent molecular weights of 395 kd and 345 kd, regardless of whether the gels were run under reducing or nonreducing conditions. It is noteworthy that these bands were exclusively detectable in lysates prepared from proliferating cells, whereas they were absent in lysates obtained from quiescent cells. These immunobiochemical data are further substantiated by our molecular cloning approaches. By means of immunocloning with Ki-67, the authors isolated and sequenced several cDNA fragments from lambda gt11 libraries. A 1095-bp fragment gave a strong hybridization signal at 7.5 to 9.5 kb in Northern blot analysis with RNA prepared from proliferating cells, whereas it was negative with RNA prepared from quiescent cells. This cDNA fragment could be bacterially expressed, and in subsequent immunoblot analysis Ki-67 reacted exclusively with those fusion Proteins that were derived from bacteria containing the insert in the right reading frame.
Jiwang Chern - One of the best experts on this subject based on the ideXlab platform.
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quinazolin 4 one derivatives as selective histone deacetylase 6 inhibitors for the treatment of alzheimer s disease
Journal of Medicinal Chemistry, 2013Co-Authors: Chaowu Yu, Peiteh Chang, Ling-wei Hsin, Jiwang ChernAbstract:Novel quinazolin-4-one derivatives containing a hydroxamic acid moiety were designed and synthesized. All compounds were subjected to histone deacetylase (HDAC) enzymatic assays to identify selective HDAC6 inhibitors with nanomolar IC50 values. (E)-3-(2-Ethyl-7-fluoro-4-oxo-3-phenethyl-3,4-dihydroquinazolin-6-yl)-N-hydroxyacrylamide, 4b, is the most potent HDAC6 inhibitor (IC50, 8 nM). In vitro, these compounds induced neurite outgrowth accompanied by growth-associated Protein 43 expression, and they enhanced the synaptic activities of PC12 and SH-SY5Y neuronal cells without producing toxic or mitogenic effects. Several of the compounds dramatically increased Nonhistone Protein acetylation, specifically of α-tubulin. Some of the more potent HDAC6 inhibitors decreased zinc-mediated β-amyloid aggregation in vitro. N-Hydroxy-3-(2-methyl-4-oxo-3-phenethyl-3,4-dihydro-quinazolin-7-yl)-acrylamide, 3f, the most promising drug candidate, selectively inhibits HDAC6 (IC50, 29 nM), practically does not affect human ...
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quinazolin 4 one derivatives as selective histone deacetylase 6 inhibitors for the treatment of alzheimer s disease
Journal of Medicinal Chemistry, 2013Co-Authors: Peiteh Chang, Ling-wei Hsin, Jiwang ChernAbstract:Novel quinazolin-4-one derivatives containing a hydroxamic acid moiety were designed and synthesized. All compounds were subjected to histone deacetylase (HDAC) enzymatic assays to identify selective HDAC6 inhibitors with nanomolar IC50 values. (E)-3-(2-Ethyl-7-fluoro-4-oxo-3-phenethyl-3,4-dihydroquinazolin-6-yl)-N-hydroxyacrylamide, 4b, is the most potent HDAC6 inhibitor (IC50, 8 nM). In vitro, these compounds induced neurite outgrowth accompanied by growth-associated Protein 43 expression, and they enhanced the synaptic activities of PC12 and SH-SY5Y neuronal cells without producing toxic or mitogenic effects. Several of the compounds dramatically increased Nonhistone Protein acetylation, specifically of α-tubulin. Some of the more potent HDAC6 inhibitors decreased zinc-mediated β-amyloid aggregation in vitro. N-Hydroxy-3-(2-methyl-4-oxo-3-phenethyl-3,4-dihydro-quinazolin-7-yl)-acrylamide, 3f, the most promising drug candidate, selectively inhibits HDAC6 (IC50, 29 nM), practically does not affect human ether-a-go-go-related membrane channel activity (IC50 >10 μM) or cytochrome P450 activity (IC50 >6.5 μM) in vitro, and significantly improves learning-based performances of mice with β-amyloid-induced hippocampal lesions.
Raymond Reeves - One of the best experts on this subject based on the ideXlab platform.
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organization inducible expression and chromosome localization of the human hmg i y Nonhistone Protein gene
Nucleic Acids Research, 1993Co-Authors: Michael Friedmann, Laurel T Holth, Huda Y Zoghbi, Raymond ReevesAbstract:Members of the HMG-I(Y) family of mammalian Nonhistone Proteins are of importance because they have been demonstrated to bind specifically to the minor groove of A.T-rich sequences both in vitro and in vivo and to function as gene transcriptional regulatory Proteins in vivo. Here we report the cloning, sequencing, characterization and chromosomal localization of the human HMG-I(Y) gene. The gene has several potential promoter/enhancer regions, a number of different transcription start sites and numerous alternatively spliced exons making it one of the most complex Nonhistone chromatin Protein-encoding genes so far reported. The putative promoter/enhancer regions each contain a number of conserved nucleotide sequences for potential binding of inducible regulatory transcription factors. Consistent with the presence of these conserved sequences, we found that transcription of the HMG-I(Y) gene is inducible in human lymphoid cells by factors such as phorbol esters and calcium ionophores. Detailed sequence analysis confirms our earlier suggestion that alternative splicing of precursor mRNAs gives rise to the major HMG-I and HMG-Y isoform Proteins found in human cells. Furthermore, the gene's exon-intron arrangement fully accounts for all of the previously cloned human HMG-I(Y) cDNAs (1,2). Also of considerable interest is the fact that each of the three different DNA-binding domain peptides present in an individual HMG-I(Y) Protein is coded for by sequences present on separate exons thus potentially allowing for exon 'shuffling' of these functional domains during evolution. And, finally, we localized the gene to the short arm of chromosome 6 (6p) in a region that is known to be involved in rearrangements, translocations and other abnormalities correlated with a number of human cancers.
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phosphorylation of the dna binding domain of Nonhistone high mobility group i Protein by cdc2 kinase reduction of binding affinity
Proceedings of the National Academy of Sciences of the United States of America, 1991Co-Authors: Raymond Reeves, Thomas A Langan, Mark S NissenAbstract:Mammalian high-mobility group I Nonhistone Protein (HMG-I) is a DNA-binding chromatin Protein that has been demonstrated both in vitro and in vivo to be localized to the A + T-rich sequences of DNA. Recently an unusual binding domain peptide, "the A.T-hook" motif, that mediates specific interaction of HMG-I with the minor groove of DNA in vitro has been described. Inspection of the A.T-hook region of the binding domain showed that it matches the consensus sequence for phosphorylation by cdc2 kinase. Here we demonstrate that HMG-I is a substrate for phosphorylation by purified mammalian cdc2 kinase in vitro. The site of phosphorylation by this enzyme is a threonine residue at the amino-terminal end of the principal binding-domain region of the Protein. Labeling of mitotically blocked mouse cells with [32P]phosphate demonstrates that this same threonine residue in HMG-I is also preferentially phosphorylated in vivo. Competition binding studies show that cdc2 phosphorylation of a synthetic binding-domain peptide significantly weakens its interaction with A + T-rich DNA in vitro, and a similar weakening of DNA binding has been observed for intact murine HMG-I Protein phosphorylated by the kinase in vitro. These findings indicate that cdc2 phosphorylation may significantly alter the DNA-binding properties of the HMG-I Proteins. Because many cdc2 substrates are DNA-binding Proteins, these results further suggest that alteration of the DNA-binding affinity of a variety of Proteins is an important general component of the mechanism by which cdc2 kinase regulates cell cycle progression.
H D Flad - One of the best experts on this subject based on the ideXlab platform.
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cell proliferation associated nuclear antigen defined by antibody ki 67 a new kind of cell cycle maintaining Proteins
Archivum Immunologiae Et Therapiae Experimentalis, 1995Co-Authors: M Duchrow, H D Flad, Claudia Wohlenberg, C Schluter, G Key, M H G Kubbutat, Johannes GerdesAbstract:Abstract A decade of studies on the human nuclear antigen defined by monoclonal antibody Ki-67 (the "Ki-67 Protein") has made it abundantly clear that this structure is strictly associated with human cell proliferation and that the expression of this Protein can be used to assess the growth fraction of a given cell population. Until recently the Ki-67 Protein was described as a Nonhistone Protein that is highly susceptible to protease treatment. We have isolated and sequenced cDNAs encoding for this antigen and found two isoforms of the full length cDNA of 11.5 and 12.5 kb, respectively, sequence and structure of which are thus far unique. The gene encoding the Ki-67 Protein is organized in 15 exons and is localized on chromosome 10. The center of this gene is formed by an extraordinary 6845 bp exon containing 16 successively repeated homologous segments of 366 bp ("Ki-67 repeats"), each containing a highly conserved new motif of 66 bp ("Ki-67 motif"). The deduced peptide sequence of this central exon possess 10 ProGluSerThr (PEST) motifs which are associated with high turnover Proteins such as other cell cycle-related Proteins, oncogenes and transcription factors, etc. Like the latter Proteins the Ki-67 antigen plays a pivotal role in maintaining cell proliferation because Ki-67 Protein antisense oligonucleotides significantly inhibit 3H-thymidine incorporation in permanent human tumor cell lines in a dose-dependent manner.
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immunobiochemical and molecular biologic characterization of the cell proliferation associated nuclear antigen that is defined by monoclonal antibody ki 67
American Journal of Pathology, 1991Co-Authors: Johannes Gerdes, M Duchrow, Li Li, C Schlueter, Claudia Wohlenberg, C Gerlach, I Stahmer, Sabine Kloth, Ernst Brandt, H D FladAbstract:The monoclonal antibody Ki-67 detects a human nuclear antigen that is present in proliferating cells, but absent in quiescent cells. The aim of this study was to characterize the Ki-67 antigen by means of immunobiochemical and molecular biology techniques. Enzymatic digestion experiments showed that this antigen is highly susceptible to protease treatment, and the antigen cannot be extracted by 0.1 normal HCl, indicating that Ki-67 antigen is a Nonhistone Protein. Immunoblot analysis of cell lysates with Ki-67 showed a double band with apparent molecular weights of 395 kd and 345 kd, regardless of whether the gels were run under reducing or nonreducing conditions. It is noteworthy that these bands were exclusively detectable in lysates prepared from proliferating cells, whereas they were absent in lysates obtained from quiescent cells. These immunobiochemical data are further substantiated by our molecular cloning approaches. By means of immunocloning with Ki-67, the authors isolated and sequenced several cDNA fragments from lambda gt11 libraries. A 1095-bp fragment gave a strong hybridization signal at 7.5 to 9.5 kb in Northern blot analysis with RNA prepared from proliferating cells, whereas it was negative with RNA prepared from quiescent cells. This cDNA fragment could be bacterially expressed, and in subsequent immunoblot analysis Ki-67 reacted exclusively with those fusion Proteins that were derived from bacteria containing the insert in the right reading frame.
M Duchrow - One of the best experts on this subject based on the ideXlab platform.
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cell proliferation associated nuclear antigen defined by antibody ki 67 a new kind of cell cycle maintaining Proteins
Archivum Immunologiae Et Therapiae Experimentalis, 1995Co-Authors: M Duchrow, H D Flad, Claudia Wohlenberg, C Schluter, G Key, M H G Kubbutat, Johannes GerdesAbstract:Abstract A decade of studies on the human nuclear antigen defined by monoclonal antibody Ki-67 (the "Ki-67 Protein") has made it abundantly clear that this structure is strictly associated with human cell proliferation and that the expression of this Protein can be used to assess the growth fraction of a given cell population. Until recently the Ki-67 Protein was described as a Nonhistone Protein that is highly susceptible to protease treatment. We have isolated and sequenced cDNAs encoding for this antigen and found two isoforms of the full length cDNA of 11.5 and 12.5 kb, respectively, sequence and structure of which are thus far unique. The gene encoding the Ki-67 Protein is organized in 15 exons and is localized on chromosome 10. The center of this gene is formed by an extraordinary 6845 bp exon containing 16 successively repeated homologous segments of 366 bp ("Ki-67 repeats"), each containing a highly conserved new motif of 66 bp ("Ki-67 motif"). The deduced peptide sequence of this central exon possess 10 ProGluSerThr (PEST) motifs which are associated with high turnover Proteins such as other cell cycle-related Proteins, oncogenes and transcription factors, etc. Like the latter Proteins the Ki-67 antigen plays a pivotal role in maintaining cell proliferation because Ki-67 Protein antisense oligonucleotides significantly inhibit 3H-thymidine incorporation in permanent human tumor cell lines in a dose-dependent manner.
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immunobiochemical and molecular biologic characterization of the cell proliferation associated nuclear antigen that is defined by monoclonal antibody ki 67
American Journal of Pathology, 1991Co-Authors: Johannes Gerdes, M Duchrow, Li Li, C Schlueter, Claudia Wohlenberg, C Gerlach, I Stahmer, Sabine Kloth, Ernst Brandt, H D FladAbstract:The monoclonal antibody Ki-67 detects a human nuclear antigen that is present in proliferating cells, but absent in quiescent cells. The aim of this study was to characterize the Ki-67 antigen by means of immunobiochemical and molecular biology techniques. Enzymatic digestion experiments showed that this antigen is highly susceptible to protease treatment, and the antigen cannot be extracted by 0.1 normal HCl, indicating that Ki-67 antigen is a Nonhistone Protein. Immunoblot analysis of cell lysates with Ki-67 showed a double band with apparent molecular weights of 395 kd and 345 kd, regardless of whether the gels were run under reducing or nonreducing conditions. It is noteworthy that these bands were exclusively detectable in lysates prepared from proliferating cells, whereas they were absent in lysates obtained from quiescent cells. These immunobiochemical data are further substantiated by our molecular cloning approaches. By means of immunocloning with Ki-67, the authors isolated and sequenced several cDNA fragments from lambda gt11 libraries. A 1095-bp fragment gave a strong hybridization signal at 7.5 to 9.5 kb in Northern blot analysis with RNA prepared from proliferating cells, whereas it was negative with RNA prepared from quiescent cells. This cDNA fragment could be bacterially expressed, and in subsequent immunoblot analysis Ki-67 reacted exclusively with those fusion Proteins that were derived from bacteria containing the insert in the right reading frame.