Mercuric chloride induces increases in both cytoplasmic and nuclear free calcium ions through a protein phosphorylation-linked mechanism

The mechanism of the lymphocyte stimulatory action of sulfhydryl group-reactive mercuric ions was studied with respect to its potential ability to induce a protein tyrosine phosphorylation-linked signal for mobilization of free Ca²⁺ into cytoplasm and nucleus of the cell. Exposure of human leukamic T cell line (Jurkat) cells to high (1 mM) and low (0.01 mM) concentrations of HgCl₂ induced tyrosine phosphorylation of multiple proteins in a concentration-dependent manner. Confocal microscopy directly visualized the time course localization of Ca²⁺ inside the cells after exposure to HgCl₂. The onset and level of Ca²⁺ mobilization following HgCl₂ exposure were in parallel to those of protein tyrosine phosphorylation. Interestingly, by either concentration of HgCl₂, Ca²⁺ was mobilized in both cytoplasm and nucleus almost simultaneously, and the level of Ca²⁺ mobilization in the nucleus was more than that in the cytoplasm. All the HgCl₂-mediated Ca²⁺ mobilization was prevented by addition of protein kinase inhibitor staurosporin prior to HgCl₂. These results suggest that heavy metal stress triggers a protein tyrosine phosphorylation-linked signal that leads to a nuclear event-dominant Ca²⁺ mobilization.     

Effect of Ca2+ on programmed death of guard and epidermal cells of pea leaves

The effect of Ca²⁺ on programmed death of guard cells (GC) and epidermal cells (EC) determined from destruction of the cell nucleus was investigated in epidermis of pea leaves. Ca²⁺ at concentrations of 1–100 μM increased and at a concentration of 1 mM prevented the CN—induced destruction of the nucleus in GC, disrupting the permeability barrier of GC plasma membrane for propidium iodide (PI). Ca²⁺ at concentrations of 0.1–1 mM enhanced drastically the number of EC nuclei stained by PI in epidermis treated with chitosan, an inducer of programmed cell death. The internucleosomal DNA fragmentation caused by CN^? was suppressed by 2 mM Ca²⁺ on 6 h incubation, but fragmentation was stimulated on more prolonged treatment (16 h). Presumably, the disruption of the permeability barrier of plasma membrane for PI is not a sign of necrosis in plant cells. Quinacrine and diphenylene iodonium at 50 μM concentration prevented GC death induced by CN^? or CN^? + 0.1 mM Ca²⁺ but had no influence on respiration and photosynthetic O₂ evolution in pea leaf slices. The generation of reactive oxygen species determined from 2′,7′-dichlorofluorescein fluorescence was promoted by Ca²⁺ in epidermal peels from pea leaves.     

Adenosine cyclic 3′,5′-monophosphate-dependent protein kinase from human platelets

A single cyclic AMP-dependent protein kinase (EC 2.7.1.37) has been isolated from human platelets by using DEAE-cellulose ion-exchange chromatography and Sephadex G-150 gel filtration. The molecular weight of the protein kinase was estimated to be 86 490. In the presence of cyclic AMP, the protein kinase could be dissociated into a catalytic subunit of molecular weight 50 000, and either one regulatory subunit of molecular weight 110 000 or two regulatory subunits of molecular weights 110 000 and 38 100, depending on the pH used. Recombination of either of the regulatory subunits with the catalytic subunit restored cyclic AMP-dependency in the catalytic subunit.The apparent K_m for ATP in the presence of 10 μM Mg²⁺ was 4 μM (plus cyclic AMP) and 4.3 μM (minus cyclic AMP). The concentration of cyclic AMP needed for half-maximal stimulation of the protein kinase was 0.172 μM and apparent dissociation constants of 3.7 nM (absence of MgATP) and 0.18 μM (presence of MgATP) were exhibited by the “protein kinase-cyclic AMP complex”. The enzyme required Mg²⁺ for maximum activity and showed a pH optimum of 6.2 with histone as substrate.In addition to four major endogenous platelet protein acceptors of apparent molecular weights 45 000, 28 000, 18 500, and 11 100, the platelet protein kinase also phosphorylated the exogenous acceptor proteins thrombin, collagen and histone, all capable of inducing platelet aggregation. Prothrombin, a nonaggregating agent, was not phosphorylated.     

Kaempferol promotes primordial follicle activation through the phosphatidylinositol 3‐kinase/protein kinase B signaling pathway and reduces DNA fragmentation of sheep preantral follicles cultured in vitro

The aim of this study was to evaluate the effects of kaempferol on the morphology, follicular activation, growth, and DNA fragmentation of ovine preantral follicles cultured in situ, and the effects of a phosphatidylinositol 3 kinase (PI3K) inhibitor and the expression of phosphorylated protein kinase B (pAKT) after culture. Ovine ovarian fragments were fixed for histological and terminal deoxynucleotidyl transferase‐mediated dUTP nick‐end labeling (TUNEL) analyses (fresh control) or cultured in α‐MEM+ alone (control) or with different concentrations of kaempferol (0.1, 1, 10, or 100 μM) for 7 days. Follicles were classified as normal or atretic, primordial or growing, and the oocyte and follicle diameters were measured. Proliferating cells were analyzed and DNA fragmentation was evaluated by the TUNEL assay. Inhibition of PI3K activity was performed through pretreatment in media added with 50 µM LY294002 for 1 hr and pAKT immunohistochemistry was performed after culture in the absence or presence of LY294002. After culture, the percentage of normal follicles was similar among the treatments (p > 0.05), except for 100 µM kaempferol, which had less normal follicles (p < 0.05). Moreover, kaempferol at 10 μM showed a higher percentage of follicular activation and cell proliferation than the other treatments (p < 0.05) and a percentage of TUNEL‐positive cells similar to that in the fresh control and lower than other treatments (p < 0.05). LY294002 significantly inhibited primordial follicle activation stimulated by α‐MEM+ and 10 μM kaempferol and reduced pAKT expression in those follicles. In conclusion, 10 μM kaempferol promotes primordial follicle activation and cell proliferation through the PI3K/AKT pathway and reduces DNA fragmentation of ovine preantral follicles cultured in vitro.     

Intracellular mobilization of Ca by the insect steroid hormone 20-hydroxyecdysone during programmed cell death in silkworm anterior silk glands

20-Hydroxyecdysone (20E) triggers programmed cell death (PCD) and regulates de novo gene expression in the anterior silk glands (ASGs) of the silkworm Bombyx mori. PCD is mediated via a nongenomic pathway that includes Ca²⁺ as a second messenger and the activation of protein kinase C/caspase-3-like protease; however, the steps leading to a concomitant buildup of intracellular Ca²⁺ are unknown. We employed pharmacological tools to identify the components of this pathway. ASGs were cultured in the presence of 1 μM 20E and one of the following inhibitors: a G-protein-coupled receptor (GPCR) inhibitor, a phospholipase C (PLC) inhibitor, an inositol 1,4,5-trisphosphate receptor (IP₃R) antagonist, and an L- or T-type Ca²⁺ channel blocker. The T-type Ca²⁺ channel blocker inhibited 20E-induced nuclear and DNA fragmentation; in contrast, PCD was induced by 20E in Ca²⁺-free medium, indicating that the source of Ca²⁺ is an intracellular reservoir. The IP₃R antagonist inhibited nuclear and DNA fragmentation, suggesting that the endoplasmic reticulum may be the Ca²⁺ source. Finally, the GPCR and PLC inhibitors effectively blocked nuclear and DNA fragmentation. Our results indicate that 20E increases the intracellular level of Ca²⁺ by activating IP₃R, and that this effect may be brought about by the serial activation of GPCR, PLC, and IP₃.     

Phosphorylation of myosin light chain by protease activated kinase I

Protease activated kinase I from rabbit reticulocytes has been shown to phosphorylate the P-light chain of myosin light chains isolated from rabbit skeletal muscle. The enzyme is not activated by Ca²⁺ and calmodulin or phospholipids. Protease activated kinase I is not inhibited by trifluoperazine at concentrations up to 200 μM or by the antibody to the Ca²⁺, calmodulin-dependent myosin light chain kinase from rabbit skeletal muscle. Two-dimensional peptide mapping of chymotryptic digests of myosin P-light chain show the site phosphorylated by the protease activated kinase is different from that phosphorylated by the Ca²⁺, calmodulin-dependent myosin light chain kinase.     

Regulation of DNA fragmentation: the role of caspases and phosphorylation

DNA fragmentation is a hallmark of apoptosis that is induced by apoptotic stimuli in various cell types. Apoptotic signal pathways, which eventually cause DNA fragmentation, are largely mediated by the family of cysteinyl aspartate-specific protease caspases. Caspases mediate apoptotic signal transduction by cleavage of apoptosis-implicated proteins and the caspases themselves. In the process of caspase activation, reversible protein phosphorylation plays an important role. The activation of various proteins is regulated by phosphorylation and dephosphorylation, both upstream and downstream of caspase activation. Many kinases/phosphatases are involved in the control of cell survival and death, including the mitogen-activated protein kinase signal transduction pathways. Reversible protein phosphorylation is involved in the widespread regulation of cellular signal transduction and apoptotic processes. Therefore, phosphatase/kinase inhibitors are commonly used as apoptosis inducers/inhibitors. Whether protein phosphorylation induces apoptosis depends on many factors, such as the type of phosphorylated protein, the degree of activation and the influence of other proteins. Phosphorylation signaling pathways are intricately interrelated; it was previously shown that either induction or inhibition of phosphorylation causes cell death. Determination of the relationship between protein and phosphorylation helps to reveal how apoptosis is regulated. Here we discuss DNA fragmentation and protein phosphorylation, focusing on caspase and serine/threonine protein phosphatase activation.     

Isolation of brain Ca2+-calmodulin tubulin kinase containing calmodulin binding proteins

Ca²⁺-calmodulin tubulin kinase activity was isolated from brain cytosol and separated from its substrate protein, tubulin, and Ca²⁺ regulatory protein, calmodulin. Characterization of the Ca²⁺-tubulin kinase system revealed a Km of 4 μM, 0.5 μM, 60 μM for Ca²⁺, calmodulin and ATP, respectively. The tubulin kinase system bound to a calmodulin affinity column in the presence of Ca²⁺ and was released from the column by chelation with EGTA. A major 55,000 and a minor 65,000 dalton peptide were identified as the only calmodulin binding proteins in the enzyme fraction, indicating that one or both of these peptides represent the calmodulin binding subunit of the Ca²⁺-calmodulin tubulin kinase system.     

Lysine-rich histone H1 kinase from soybean hypocotyl.

A protein kinase (ATP: histone phosphotransferase) with high specificity for the phosphorylation of the very lysine-rich histone H1 has been partially purified and characterized from soybean hypocotyl. The enzyme has a molecular weight of about 48,500. Its activity and sedimentation behavior are refractory to cyclic nucleoside monophosphates. No significant amount of cyclic AMP or cyclic GMP binding activity could be detected in the crude or partially purified enzyme preparations. Km for ATP and histone H1 are 0.4 μM and 0.7 μM, respectively. The enzyme requires Mg²⁺ or Mn²⁺ for activity, while addition of 0.5 mM Ca²⁺, Zn²⁺ or Hg²⁺ results in 50% inhibition. Arginine-rich histones H3 and H4 are inhibitory to histone H1 phosphorylation; these histones affect the Vmax of the enzyme, but not the Km for histone H1.     

A new rhodamine‐based fluorescent chemodosimeter for mercuric ions in water media

A new rhodamine–ethylenediamine–nitrothiourea conjugate (RT) was synthesized and its sensing property as a fluorescent chemodosimeter toward metal ions was explored in water media. Analytical results from absorption and fluorescence spectra revealed that the addition of Hg²⁺ ions to the aqueous solution of the chemodosimeter RT caused a distinct fluorescence OFF–ON response with a remarkable visual color change from colorless to pink; however, no clear spectral and color changes were observed from other metal ions including: Zn²⁺, Cu²⁺, Cd²⁺, Pb²⁺, Ag⁺, Fe²⁺, Cr³⁺, Co³⁺, Ni²⁺, Ca²⁺, Mg²⁺, K⁺ and Na⁺. The sensing results and the molecular structure suggested that a Hg²⁺‐induced a desulfurization reaction and cyclic guanylation of the thiourea moiety followed by ring‐opening of the rhodamine spirolactam in RT are responsible for a distinct fluorescence turn‐on signal, indicating that RT is a remarkably sensitive and selective chemodosimeter for Hg²⁺ ions in aqueous solution. Hg²⁺ within a concentration range from 0.1 to 25 μM can be determined using RT as a chemodosimeter and a detection limit of 0.04 μM is achieved. Copyright © 2014 John Wiley & Sons, Ltd.     

Investigation of carvedilol-evoked Ca2+ movement and death in human oral cancer cells

The effect of carvedilol on cytosolic free Ca²⁺ concentrations ([Ca²⁺]_i) in OC2 human oral cancer cells is unknown. This study examined if carvedilol altered basal [Ca²⁺]_i levels in suspended OC2 cells by using fura-2 as a Ca²⁺-sensitive fluorescent probe. Carvedilol at concentrations between 10 and 40 µM increased [Ca²⁺]_i in a concentration-dependent fashion. The Ca²⁺ signal was decreased by 50% by removing extracellular Ca²⁺. Carvedilol-induced Ca²⁺ entry was not affected by the store-operated Ca²⁺ channel blockers nifedipine, econazole, and SK&F96365, but was enhanced by activation or inhibition of protein kinase C. In Ca²⁺-free medium, incubation with the endoplasmic reticulum Ca²⁺ pump inhibitor thapsigargin did not change carvedilol-induced [Ca²⁺]_i rise; conversely, incubation with carvedilol did not reduce thapsigargin-induced Ca²⁺ release. Pretreatment with the mitochondrial uncoupler carbonylcyanide m-chlorophenylhydrazone (CCCP) inhibited carvedilol-induced [Ca²⁺]_i release. Inhibition of phospholipase C with U73122 did not alter carvedilol-induced [Ca²⁺]_i rise. Carvedilol at 5–50 µM induced cell death in a concentration-dependent manner. The death was not reversed when cytosolic Ca²⁺ was chelated with 1,2-bis(2-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid acetoxymethyl ester (BAPTA/AM). Annexin V/propidium iodide staining assay suggests that apoptosis played a role in the death. Collectively, in OC2 cells, carvedilol induced [Ca²⁺]_i rise by causing phospholipase C-independent Ca²⁺ release from mitochondria and non-endoplasmic reticulum stores, and Ca²⁺ influx via protein kinase C-regulated channels. Carvedilol (up to 50 μM) induced cell death in a Ca²⁺-independent manner that involved apoptosis.     

Effect of phorbol 12-myristate 13-acetate on Ca2+-ATPase activity in rat liver nuclei

The effect of phorbol 12-myristate 13-acetate (PMA) on Ca²⁺-ATPase activity in rat liver nuclei was investigated. Ca²⁺-ATPase activity was calculated by subtracting Mg²⁺-ATPase activity from (Ca²⁺-Mg²⁺)-ATPase activity. The nuclear Ca²⁺-ATPase activity was significantly increased by the presence of PMA (2–20 μM) in the enzyme reaction mixture; the maximum effect was seen at 10 μM. The PMA (10 μM)-increased Ca²⁺-ATPase activity was not blocked by the presence of staurosporine (2 μM) or dibucaine (2 and 10 μM), an inhibitor of protein kinase. Meanwhile, vanadate (20 and 100 μM) caused a significant reduction in the nuclear Ca²⁺-ATPase activity increased by PMA (10 μM). The present finding suggests that PMA has an activating effect on liver nuclear Ca²⁺-ATPase independent of protein kinase. © 1994 Wiley-Liss, Inc.     

Cadmium-induced apoptosis in C6 glioma cells: Mediation by caspase 9-activation

The induction of apoptotic cell death by cadmium was investigated in eight mammalian cell lines. Great differences in the cytotoxicity of cadmium were found with different cell lines: Rat C6 glioma cells turned out to be most sensitive with an IC₅₀-value of 0.7 M, while human A549 adenocarcinoma cells were relatively resistant with an IC₅₀-value of 164 M CdCl₂. The mode of cadmium-induced cellular death was identified to involve apoptotic DNA fragmentation in three cell lines, i.e., in C6 glioma cells, E367 neuroblastoma cells and NIH3T3 fibroblasts. In C6 glioma cells, this process was investigated in detail. Internucleosomal DNA-fragmentation occurred 40 h after application of CdCl₂ and was concentration-dependent between 1–100 M CdCl₂, followed by a decrease at higher concentrations due to necrotic processes. Apoptotic chromatin-condensation and nuclear fragmentation was observed 48 h after application of 2.5 M CdCl₂. Furthermore, cadmium (1 M, 48 h) caused a breakdown of the mitochondrial membrane potential as shown by the decline in mitochondrial uptake of rhodamine 123. Also, we found an activation of caspase 9, a protease known to be activated in apoptotic processes following mitochondrial damage. Besides Cd²⁺, other toxic heavy metal ions (Hg²⁺, Pb²⁺, Ni²⁺, Fe²⁺, CrO₄ ^2–, Cu²⁺ or Co²⁺) did not induce apoptotic DNA fragmentation in C6 cells. The only exception was Zn²⁺ which caused apotosis at high concentrations (>150 M) whereas it protected against cadmium-induced apoptosis at low concentrations (10–50 M).     

Inhibitory effect of calcium-binding protein regucalcin on protein kinase activity in the nuclei of regenerating rat liver

The effect of Ca²⁺-binding protein regucalcin on protein kinase activity in the nuclei of normal and regenerating rat livers was investigated. Protein kinase activity in the nuclei isolated from normal rat liver was significantly increased by addition of Ca²⁺ (500 μM) and calmodulin (10 μg/ml) in the enzyme reaction mixture. Nuclear protein kinase activity was significantly decreased in the presence of EGTA (1.0 mM), trifluoperazine (TFP; 20 μM), dibucaine (10⁻⁴ M), or staurosporine (10⁻⁷ M), indicating that Ca²⁺-dependent protein kinases are present in the nuclei. Protein kinase activity was significantly elevated in the liver nuclei obtained at 6 to 48 h after a partial hepatectomy. Hepatectomy-increased nuclear protein kinase activity was significantly decreased in the presence of EGTA (1.0 mM), TFP (20 μM), or staurosporine (10⁻⁷ M) in the enzyme reaction mixture. The presence of regucalcin (0.1–0.5 μM) caused a significant decrease in protein kinase activity in the nuclei obtained from normal and regenerating rat livers. Meanwhile, the nuclear protein kinase activity from normal and regenerating livers was significantly elevated in the presence of anti-regucalcin monoclonal antibody (50–200 ng/ml). The present study suggests that regucalcin plays a role in the regulation of protein kinase activity in the nuclei of proliferative liver cells. J. Cell. Biochem. 71:569–576, 1998. © 1998 Wiley-Liss, Inc.     

Stimulation by phspholipid of calcium-dependent phosphorylation of endogenous proteins from mammalian tissues

The occurrence of endogenous substrate proteins for calcium-dependent protein kinase, augmented by either phospholipid or calmodulin, was examined in extracts of several tissues. Pancreas, vas deferens, adrenal and liver were found to contain substrate proteins for phospholipid-sensitive protein kinase. Phosphorylation of pancreatic substrate protein for phospholipid-senstivie protein kinase was rapid and highly sensitive to Ca²⁺, being detectable within 15 s a following exposure to Ca²⁺ and phosphatidylserine and at concentrations of Ca²⁺ as low as 0.5 μM. These findings suggest that the phospholipid-sensitive protien kinase system may serve to mediate some effects of Ca²⁺ in a variety of mammalian cell types.     

Influence of Mercury on the Green Alga Haematococcus lacustris. Inhibition Effects and Recovery of Impact

The influence of mercury ions on germination of the resting cells (aplanospores) and on cell division, cell structure, phototaxis, and photosynthesis during the flagellate stage of Hoemotococcus lacustris was investigated. Aplanospores possess a higher tolerance against mercury ions than flagellates. The reason could be seen in the thicker wall of the resting cells which possibly provide a detoxifying effect by immobilisation of Hg²⁺. This is confirmed by a normal phototactic orientation of flagellates formed from Hg ²⁺-influenced aplanospores. In contrast, a direct addition of Hg²⁺ (0.1 to 1 /*M) to the flagellate stage induced an immediate loss of the flagellates to react phototactically, but it was interesting that the inhibition was overcome with time. Obviously, these Hg²⁺ concentrations influence only the sensory transduction chain, whereas the energetic background is not injured because the velocity of movement and the percentage of motile cells were scarcely affected. This is supported by the high level of the photochemical efficiency of photosystem II, which remains unchanged at 1 uM Hg²⁺. Recovery of photosynthesis from inhibition by 10/iM Hg²⁺ suggests a connection between Hg²⁺ influence and metabolism of the D, protein in the reaction centre of photosystem II. The Hg²⁺ effect was reversed with time in light, but not in darkness, and streptomycin, an inhibitor of chloroplast protein synthesis, prevented recovery. In flagellates, showing no reactivation, exposure to 10uA/l Hg²⁺ caused cell swelling, a loss of the flagella, and a disorganisation in structure of the chloroplast and of nucleus.     

Bcl-2 Protects Against Apoptosis in Neuronal Cell Line Caused by Thapsigargin-Induced Depletion of Intracellular Calcium Stores

Abstract: The toxicity of thapsigargin, a selective inhibitor of endoplasmic reticular Ca²⁺-ATPase, was investigated in GT1-7 cells, a murine hypothalamic cell line. Treatment of these cells with 50 or 100 nM thapsigargin greatly reduced cell viability at 24 and 48 h. These doses of thapsigargin induced a rapid rise in free cytosolic Ca²⁺ ([Ca²⁺]_i), followed by a sustained increase. Addition of EGTA to chelate extracellular Ca²⁺ diminished somewhat the size of the initial increase of [Ca²⁺]_i caused by thapsigargin, and abolished the sustained increase. The sustained increase could also be abolished by addition of La³⁺ and by SKF 96365, a drug selective for receptor-mediated calcium entry, but not by verapamil or flunarizine. Pretreatment with 50 µM BAPTA/AM, a cytosolic Ca²⁺ chelator, inhibited the peak [Ca²⁺]_i caused by thapsigargin but did not inhibit the sustained elevation of [Ca²⁺]_i. Neither EGTA nor BAPTA/AM inhibited the cell death induced by thapsigargin. The cell death was characterized by DNA fragmentation (“laddering”), nuclear condensation and fragmentation, and was inhibited by protein synthesis inhibitor cycloheximide, all characteristic of apoptotic cell death. Overexpression of the proto-oncogene bcl-2 in GT1-7 cells inhibited significantly DNA fragmentation, nuclear condensation and fragmentation, and cell death induced by thapsigargin. However, Bcl-2 did not alter either basal [Ca²⁺]_i or the elevation of [Ca²⁺]_i induced by thapsigargin. Our results suggest that abnormal Ca²⁺ release from endoplasmic reticulum caused by thapsigargin induces GT1-7 death by apoptosis and that this effect does not depend on Ca²⁺ influx from the extracellular space. Bcl-2 inhibited apoptosis induced by thapsigargin, but the mechanism is unlikely to be inhibition of endoplasmic reticular Ca²⁺ release in GT1-7 neuronal cells.     

Apoptosis and Its Modulation in Human Promyelocytic HL-60 Cells Treated with DNA Topoisomerase I and II Inhibitors

Electron microscopy studies demonstrate unequivocally that the observed oligonucleosome-sized secondary DNA fragmentation in human promyelocytic HL-60 cells treated with the topoisomerase inhibitors camptothecin and teniposide is correlated with the morphological changes in cell structure typical of programmed cell death (apoptosis). Since apoptosis has been associated with potential involvement of intracellular signaling linked to the Ca²⁺/calmodulin and protein kinase C transduction pathways, we also investigated the effects of signaling modulators on camptothecin- and teniposide-induced secondary DNA fragmentation in HL-60 cells. Neither calcium chelators, calcium/calmodulin inhibitors (calmidazolium or cyclosporine A), protein kinase C stimulation by TPA, protein phosphatase inhibition by okadaic acid, protein kinase inhibition by staurosporine, calphostin C, genistein or H7, nor cell cycle alterations by caffeine had any detectable effect. Interestingly, most of these intracellular signaling modulators were able to induce DNA fragmentation in HL-60 cells by themselves. These results may suggest that even though modulation of these signaling pathways was unable to prevent topoisomerase inhibitor-induced apoptosis, their sole deregulations could induce apoptosis in HL-60 cells. In contrast, aphidicolin blocked camptothecin-induced secondary DNA fragmentation, indicating that replication-induced DNA damage is required for camptothecin- but not teniposide-induced secondary DNA fragmentation. Zinc, 3-aminobenzamide, and spermine also modulated both camptothecin- and teniposide-induced secondary DNA fragmentation without significant alteration of topoisomerase-mediated primary DNA strand breaks. Hence, poly(ADP-ribosyl)ation and chromatin structure may be important in modulating oligonucleosomesized DNA fragmentation associated with apoptosis in HL-60 cells treated with topoisomerase inhibitors.