Isolation and purification of calcium and magnesium dependent endonuclease from rat liver nuclei

Ishida, Ryoji; Akiyoshi, Hiroto; Takahashi, Taijo

doi:10.1016/0006-291x(74)90662-7

Cited by 98 publications

(25 citation statements)

References 10 publications

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“…This wo-ld be possible only when CME is presented in tile nuclei in th~ fr~ form (not bound to chromatin) and every linker ~egtaent is accessible to the enzyme. However, biochemical e;4-dence has shown that the enzyme is bound to ehromatin and can only be elated with salt [18].…”

Section: Discussionmentioning

confidence: 99%

Internucleosomal chromatin degradation in myeloma and B‐hybridoma cell cultures

et al. 1992

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The activity of Ca/Mg-dependent endonuelease (CME) is strongly inhibited in mycloma X-63.Agg.653 and B-hybridoma MLC-Ic as compared with mouse splenocytes. Nevertheless, pronounced internu¢leo~omal chromatin degradation oecur~ in both cell lines during long-term cultivation without passing. In isolated cell nuclei of X-63 the activation of CM E, which precedes chromatin fra~nentation in vivo and loss of cell viability, is revealed. The time--course of CME activation is opposite to cell proliferation and is not accompanied by alterations in enzyme quantity. The results suggest that cell death of X-63 and MLC-lc occurs via apoptosis0 and involves the mechanisms controlling the activation and/or interaction of CME with chromatin.

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Section: Discussionmentioning

confidence: 99%

Internucleosomal chromatin degradation in myeloma and B‐hybridoma cell cultures

et al. 1992

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“…For example, it is well known that Ca2+/Mg2+-dependent nuclease can be poly(ADP-ribosy1)ated and that this modification inhibits its activity [31,401. A lot of preparations of enzymes belonging to this group of nucleases have been described [22,23,[25][26][27][28][29][30][41][42][43]. They are variable, in their affinity to different ion exchangers, molecular mass and some other properties but differ from DNase-I-like nucleases by showing the maximum activity in the presence of a combination of Ca2+ and Mg2+ or Mn2+, insensitivity to G-actin, inhibition of their activity by iodacetamide and N-ethylmaleimide [25,26,301.…”

Section: Discussionmentioning

confidence: 99%

Properties of some nuclear nucleases of rat thymocytes and their changes in radiation‐induced apoptosis

Nikonova

Beletsky

Umansky

1993

European Journal of Biochemistry

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Three nuclease activites have been found and characterized in rat thymocyte nuclear extracts. A Mn2+-dependent nuclease is loosely bound to nuclear components and can be extracted with 0.35 M NaCI. The enzyme is activated by MnZ+ but not by Mg", Ca2+, or both. Its molecular mass is 36-40 kDa when measured by gel filtration and 37 kDa by SDSPAGE. An acidic nuclease is independent of divalent ions, produces DNA strand breaks with 5'-OH ends, its molecular mass is about 37 kDa. Two fractions of Ca2+Nn2+-dependent nuclease, differing in binding to CM-Sepharose but identical in other respects, are active in the presence of Mn2+ but can be additionally activated by Ca2+. They are inactive in the presence of Mg'' or Ca2+ but cleave DNA in Ca2'/Mg2'-containing medium. The molecular mass of the enzyme is 22 kDa as determined by both gel filtration and electrophoresis. The dependence of nuclease activities on pH, ions, and sulfhydryl reagents is described. Cycloheximide injection to both control and irradiated animals strongly inhibits the activities of Ca2+/Mn2+-dependent nuclease from thymocyte nuclei separated by chromatography on CMSepharose and does not change the activities of Mn2+-dependent and acidic nucleases. Nuclease activity in thymocyte nuclei from irradiated rats is increased in CaZ+Ng2'-containing and Ca2+/ Mn"-containing media whereas there is no change in the activity of acidic nuclease. CaZ+/Mn2+-dependent nuclease is extracted from thymocyte nuclei of irradiated rats with 0.35 M NaCl but from control nuclei only with 0.5 M NaCl. Possible reasons of labilization of Ca*+Nn*+-dependentnuclease binding to the nuclear structures in dying thymocytes are discussed.In the past decade the concept has been formulated that in most cases the death of cells is the result of their active response to physiological or damaging factors [ 1 -31. This type of cell death was called apoptosis [l] and the recent literatur data suggests that, besides the programmed and physiological cell death [l -31, cell death [5-81. Data indicate the involvement of nuclear nucleases, particularly, Ca2+Ng2+-dependent endonuclease [7, 21 -241 but the mechanism of itdtheir activation in dying cells remain unclear. A number of preparations of this enzyme were purified from different sources but they usually differed in molecular mass, substrate specificity, binding to ion-exchange columns and so on [25-301. Ca2+/Mg2+-nucleases with molecular masses in the range 25-120 kDa with preference for single-stranded or double-stranded DNA are described. The mechanisms of regulation of the nuclease activity are poorly understood. Even the ion dependence of the nuclease differs between various studies. The enzyme is normally inactive in the presence of Mg2+ or Ca" only but is activated by addition of both ions. At the same time the nuclease is normally active in the presence of Mn2+ and the maximal activity is manifested with Mn' + and Caz+ [25, 301. Previously it has been shown that there are different nucleases in rat thymus nuclei but only a CaZ...

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“…It is possible that larger DS DNA could be integrated into the chromosome in an unstable or abortive fashion such that it is not ligated into DNA of the DI fraction. Indeed, specific chromosome-associated nucleases (30)(31)(32) (33).…”

Section: Discussionmentioning

confidence: 99%

“…Is this a special class of nucleosomes, released perhaps through endogenous nuclease action (30)(31)(32)? Or is it possible that aphidicolin blocks endogenous nucleases such that radiolabeled DNA is prevented from appearing in the large size classes of the DS fraction, even though the radiolabeled oligonucleotides are processed to DI DNA?…”

Section: Discussionmentioning

confidence: 99%

Kinetics of thymidine incorporation into detergent-soluble DNA of mouse lymphocytes.

Lagree¹,

Mui²

1984

Proc. Natl. Acad. Sci. U.S.A.

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We reported recently that splenocytes from concanavalin A-stimulated mice rapidly incorporated [3HJthy- In experiments testing the effects of aphidicolin (5 or 10 ,ug/ml) or hydroxyurea (2 or 5 mM), the inhibitor was added 15-60 min before addition of [3H]dThd and its presence was maintained throughout the pulse-chase period.Isolation and Analysis of DNA from the DS and DetergentInsoluble (DI) Fractions. DS DNA was purified from NP-40 supernate as described (9), except that cesium chloride density centrifugation and dialysis were omitted. Each isolate from 5 x 107 cells was dissolved in 100 ,ul of 10 mM Tris-HCl (pH 8.0). Aliquots were resolved by electrophoresis using native 1.2% agarose gels. After visualization of the DNA with ethidium bromide, the gel was cut into 2.0-mm slices.Each slice was dissolved as described (9) 7056The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

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Isolation and purification of calcium and magnesium dependent endonuclease from rat liver nuclei

Cited by 98 publications

References 10 publications

Internucleosomal chromatin degradation in myeloma and B‐hybridoma cell cultures

Internucleosomal chromatin degradation in myeloma and B‐hybridoma cell cultures

Properties of some nuclear nucleases of rat thymocytes and their changes in radiation‐induced apoptosis

Kinetics of thymidine incorporation into detergent-soluble DNA of mouse lymphocytes.

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