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  • 1
    Electronic Resource
    Electronic Resource
    DNA fragmentation during thymic apoptosis is catalyzed by DNase γ (1996)
    Springer
    Apoptosis 1 (1996), S. 147-152 
    Details
    ISSN: 1573-675X
    Keywords: Apoptosis ; DNA fragmentation ; DNase γ ; programmed cell death ; thymocytes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract The physiological and pathological importance of cell death by apoptosis has recently been recognized. One of the hallmarks of apoptosis is the enzymatic cleavage of genomic DNA into nucleosomal oligomers. The identification of an endonuclease responsible for apoptosis might help to explain how this cell suicide is regulated and why DNA is cleaved. Here, we found that γ type of DNase was retained in apoptotic rat thymocyte nuclei. Homogeneously purified DNase γ (Mr = 33 kDa) from the apoptotic nuclei was revealed to be a Ca2+/Mg2+-dependent endonuclease and inhibited by Zn2+. This enzyme cleaved chromosomal DNA with 3′-hydroxyl (OH) and 5′-phosphoryl (P) ends. The cleavage ends and its divalent cation dependencies match those observed in apoptotic thymocytes induced by X-irradiation or glucocorticoid treatment, indicating that this endonuclease is a central component of the thymic apoptosis machinery.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF01321021
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  • 2
    Electronic Resource
    Electronic Resource
    The restrictive proteolysis of α-fodrin to a 120 kDa fragment is not catalyzed by calpains during thymic apoptosis (1997)
    Springer
    Apoptosis 2 (1997), S. 84-90 
    Details
    ISSN: 1573-675X
    Keywords: Apoptosis ; α-fodrin ; calpain ; thymocytes ; x-irradiation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract The α-subunit (240 kDa) of fodrin was found to be digested selectively to a 120 kDa fragment during apoptosis of rat thymocytes in vivo and in vitro. This fragment was detected by an antibody (Ab) against full length α-fodrin, but not by the anti-N-terminal sequence (GMMPR) of the μ-calpain-generated 150 kDa fragment Ab or the anti-PEST sequence of α-fodrin Ab. On the other hand, basal levels of the 150 kDa fragment were constantly recognized by these three antibodies during apoptosis. The production of the 120 kDa fragment during apoptosis was not affected by the addition of calpain inhibitors such as Ac-LLLnal and E-64d, despite inhibition of the generation of the 150 kDa fragment. When x-irradiated thymocytes were incubated in the presence of N-tosyl-L-phenylalanyl chloromethyl ketone (TPCK), both production of the 120 kDa fragment and apoptosis were suppressed. Purified μ- and m-calpain did not catalyze the formation of the 120 kDa fragment from purified α-fodrin in vitro. These results suggest that a protease different from calpains is involved in the major process of α-fodrin proteolysis to a 120 kDa fragment during thymic apoptosis.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1026443926962
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  • 3
    Electronic Resource
    Electronic Resource
    Purification and characterization of a DNase γ-like endonuclease from Xenopus laevis liver (1998)
    Springer
    Apoptosis 3 (1998), S. 145-153 
    Details
    ISSN: 1573-675X
    Keywords: Apoptosis ; DNA fragmentation ; DNase γ ; endonuclease ; Xenopus laevis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract We recently found that two apoptotic DNase γ-like endonucleases (36 and 38kDa DNases) were present in Xenopus laevis larval and adult liver cell nuclei and that their activities increased in metamorphic climax. Here, we purified the main DNase γ-like endonuclease from Xenopus laevis liver cell nuclei and characterized its physical and enzymatic properties in detail. The molecular mass of Xenopus liver nuclear endonuclease was 38,000 daltons as determined by SDS-polyacrylamide gel electrophoresis. A native molecular mass of 35,000 was estimated by gel filtration. The purified Xenopus liver endonuclease was a neutral one and required both Ca2+ and Mg2+ for DNase activity. Unlike the mammalian DNase γ, the Ca2+/Mg2+ requirement could not be supplied by Mn2+. The inhibition profiles by aurintricarboxylic acid, sodium citrate and divalent metal ions such as Co2+, Ni2+, Cu2+ and Zn2+ were similar to those of mammalian DNase γ. These results suggest that this endonuclease is a Xenopus laevis homolog of the mammalian apoptotic endonuclease DNase γ.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1009663204558
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  • 4
    Electronic Resource
    Electronic Resource
    Fas-mediated apoptosis in Jurkat cells is suppressed in the pre-G2/M phase (1999)
    Springer
    Apoptosis 4 (1999), S. 255-261 
    Details
    ISSN: 1573-675X
    Keywords: Apoptosis ; cell cycle ; G2/M phase ; Fas ; Jurkat cell.
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract The relationship between the cell cycle and Fas-mediated apoptosis was investigated using Jurkat cells. Analysis of the inducibility of apoptosis by anti-Fas antibody during the cell cycle synchronized by the thymidine double-block method, showed that apoptosis was induced in only 50% of the G2/M phase cells, while most of cells in the other phases underwent apoptosis. These observations indicate that G2/M phase cells are more resistant to Fas-mediated apoptosis than cells in other phases. Furthermore, a detailed analysis of G2/M phase found that only 20–30% of the cells underwent apoptosis 12 h after the removal of the second thymidine block (pre-G2/M phase). This suggests that Fas-mediated apoptosis is potently suppressed during the pre-G2/M phase. A possible explanation for the observation that cells in the pre-G2/M phase are less sensitive to anti-Fas antibody is lower expression level of Fas. To test this possibility, Fas expression levels on the cell surface during the cell cycle were examined. The content of Fas on the cell surface, however, did not change appreciably during the cell cycle. Thus, the suppression of apoptosis in the pre-G2/M phase is determined downstream after the receipt of the apoptotic signal through Fas.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1009652825846
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  • 5
    Electronic Resource
    Electronic Resource
    cDNA Cloning of Human DNase γ: Chromosomal Localization of Its Gene and Enzymatic Properties of Recombinant Protein (1998)
    Springer
    Apoptosis 3 (1998), S. 89-95 
    Details
    ISSN: 1573-675X
    Keywords: Apoptosis ; DNA fragmentation ; DNase γ ; endonuclease
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract We report here on the nucleotide sequence of the cDNA encoding human DNase γ, which is a candidate for an apoptotic endonuclease. The cDNA clone isolated from a human spleen cDNA library is composed of a 918 bp open reading frame encoding a 305 amino acid precursor protein for DNase γ. Northern blot analysis reveals that the expression of a single transcript of 1.5 kb DNase γ mRNA is detected in the spleen and liver. The chromosomal localization of DNase γ gene is mapped to chromosome 3 at region p21.1-p14.2 by fluorescence in situ hybridization (FISH). Characterization of thioredoxin-DNase γ fusion protein (Trx-hDNase γ) shows that the recombinant protein has a Ca2+/Mg2+- or Mn2+-dependent endonuclease activity that cleaves chromatin DNA to nucleosomal units. The optimum pH is around 7.2. Zn2+ and aurintricarboxylic acid (ATA) inhibits the activity in dose-dependent manners. These properties are identical to those of purified DNase γ.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1009692807692
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  • 6
    Electronic Resource
    Electronic Resource
    Presence of DNase γ-Like Endonuclease in Nuclei of Neuronal Differentiated PC12 Cells (1998)
    Springer
    Apoptosis 3 (1998), S. 97-103 
    Details
    ISSN: 1573-675X
    Keywords: Apoptosis ; DNA fragmentation ; DNase γ ; neurogenesis ; PC12 cells ; programmed cell death
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract DNase γ, which cleaves chromosomal DNA into nucleosomal units (DNA ladder formation), has been suggested to be the critical component of apoptotic machinery. Using rat pheochromocytoma PC12 cells, which are differentiated to sympathetic neurons by nerve growth factor (NGF), we investigated whether DNase γ-like enzyme is present in neuronal cells and is involved in neuronal cell death. The nuclear auto-digestion assay for DNase catalyzing internucleosomal DNA cleavage revealed that nuclei from neuronal differentiated PC12 cells contain acidic and neutral endonucleases, while nuclei from undifferentiated PC12 cells have only acidic endonuclease. The DNA ladder formation observed in isolated nuclei from neuronal differentiated PC12 cells at neutral pH requires both Ca2+ and Mg2+, and is sensitive to Zn2+. The molecular mass of the neutral endonuclease present in neuronal differentiated PC12 cell nuclei is 32000 as determined by activity gel analysis (zymography). The properties of the neuronal endonuclease present in neuronal differentiated PC12 cell nuclei were similar to those of purified DNase γ from rat thymocytes and splenocytes. Interestingly, in neuronal differentiated PC12 cells, internucleosomal DNA fragmentation is observed following NGF deprivation, whereas undifferentiated PC12 cells fail to exhibit DNA ladder formation during cell death by serum starvation. These results suggest that the DNase γ-like endonuclease present in neuronal differentiated PC12 cell nuclei is involved in internucleosomal DNA fragmentation during apoptosis, induced by NGF deprivation.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1009644924530
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  • 7
    Electronic Resource
    Electronic Resource
    Mesangial cell apoptosis induced by a fibronectin fragment (1998)
    Springer
    Apoptosis 3 (1998), S. 407-412 
    Details
    ISSN: 1573-675X
    Keywords: Apoptosis ; cell adhesion ; fibronectin fragment ; matrix metalloproteinase ; mesangial cell.
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract We previously showed that in passive Heymann nephritis (PHN) rats, a large quantity of fibronectin (FN) fragments containing the central cell-binding (CCB) domain and adjacent domains are generated in the kidney and excreted into urine (Nishizawa et al., Biol Pharm Bull 1998; 21: 429–433). To ascertain whether the FN fragments could affect the progression of PHN, we investigated the effect of a 150 K FN fragment containing the CCB and carboxyl-terminal heparin-binding (Hep 2) domains on cultured rat mesangial cells. When rat mesangial cells cultured on FN-coated plates were exposed to the 150 K FN fragment, some mesangial cells detached from the FN substrate and then underwent apoptosis as judged by nuclear and DNA fragmentations. The 150 K FN fragment competitively inhibited the mesangial cell adhesion to the FN substrate in a dose-dependent manner. Furthermore, gelatinzymography of the conditioned medium of mesangial cells showed that the 150 K FN fragment induced and/or poteintiated the extracellular matrix (ECM)-degrading proteinases including matrix metalloproteinases (MMPs) of mesangial cells. These results indicate that the 150 K FN fragment may elicit mesangial cell apoptosis by disrupting the mesangial cell adhesion through two distinct ways: the inhibition of mesangial cell adhesion and the ECM-degradation by the 150 K FN fragment-induced MMPs. Thus, FN fragments containing the CCB and adjacent domains generated in the kidneys of PHN rats may be involved in the evolution of the renal injury.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1009606502160
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  • 8
    Electronic Resource
    Electronic Resource
    Calculations of Electron Inelastic Mean Free Paths (IMFPs) VI. Analysis of the Gries Inelastic Scattering Model and Predictive IMFP Equation (1997)
    Chichester [u.a.] : Wiley-Blackwell
    Surface and Interface Analysis 25 (1997), S. 25-35 
    Details
    ISSN: 0142-2421
    Keywords: electron inelastic mean free path ; inelastic electron scattering ; calculated IMFP ; Chemistry ; Polymer and Materials Science
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Physics
    Notes: Gries has recently reported [Surf. Interface Anal. 24, 38 (1996)] an atomistic model for inelastic electron scattering relevant to Auger electron spectroscopy and x-ray photoelectron spectroscopy and has derived an equation (designated G1) for the estimation of inelastic mean free paths (IMFPs). We present an analysis of the Gries model and the G1 equation in terms of the similarities and differences of inelastic electron scattering by free atoms and by solids. We also compare the G1 equation with our TPP-2M equation for estimation of IMFPs. The former equation was developed from fits to our published IMFPs over the 200-2000 eV energy range, and is identical in its energy dependence to the Bethe equation for inelastic scattering cross-sections and to a simplification of our TPP-2M equation for the same energy range. Comparison of parameters indicates that the Gries fitting parameter k1 should be approximately 0.0016 and 0.0022 for non-transition and transition elements, respectively. We find that the G1 equation could be improved by allowing the Gries fitting parameter k2 to depend on density (as recommended for the equivalent parameter in TPP-2M). Although we believe that the Gries model is inconsistent with current theories for the electronic structure of metals, semiconductors and inorganic compounds, we find (from sum-rule considerations) that the G1 equation can provide an approximate guide to IMFP values. For some compounds, however, there were unexplained deviations (as found by Gries). In contrast to the G1 equation, the TPP-2M equation provides useful IMFP estimates for all materials over the parameter range that has been explored. Gries claims that the G1 equation can be extrapolated to energies lower than 200 eV on the basis of limited agreement with some experimental IMFPs over the 10-100 eV range for Be and the alkali metals, and has questioned the reliability of our IMFPs for energies below 200 eV. We consider this comparison to be inadequate, and we recommend that the G1 equation not be used in the 50-200 eV range. © 1997 by John Wiley & Sons, Ltd.
    Additional Material: 4 Ill.
    Type of Medium: Electronic Resource
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