Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P42574 (caspase-3)
 45,978 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

DNA fragmentation factor (DFF) comprises DFF45 and DFF40 subunits, the former of which acts as an inhibitor of the latter (the catalytic subunit) and whose cleavage by caspase-3 results in DFF activation. Disruption of the DFF45 gene blocks the generation of 50-kb DNA fragments and confers resistance to apoptosis. We recently suggested that the early fragmentation of DNA by DFF and the consequent activation of poly(ADP-ribose) polymerase-1 (PARP-1), mitochondrial dysfunction, and activation of caspase-3 contribute to an amplification loop in the apoptotic process. To verify the existence of such a loop, we have now examined the effects of restoring DFF expression in DFF45-deficient fibroblasts. Co-transfection of mouse DFF45(-/-) fibroblasts with plasmids encoding human DFF40 and DFF45 reversed the apoptosis resistance normally observed in these cells. The DFF45(-/-) cells regained the ability to fragment their DNA into 50-kb pieces in response to TNF, which resulted in a marked activation of PARP-1 and a concomitant depletion of intracellular NAD. DFF expression also resulted in an increase both in cytochrome c release into the cytosol and in caspase-3 activation triggered by TNF. These results support the importance of DFF, PARP-1, mitochondria, and caspase-3 in an amplification phase of TNF-induced apoptosis.
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PMID:Roles of DNA fragmentation factor and poly(ADP-ribose) polymerase-1 in sensitization of fibroblasts to tumor necrosis factor-induced apoptosis. 1178 71

In Jurkat cells Bid was cleaved upon activation of the Fas receptor with an anti-Fas antibody. The caspase-8 inhibitor benzyloxycarbonyl-Ile-Glu(OMe)-Thr-Asp(OMe)-CH(2)F (IETD) prevented the cleavage of Bid and the loss of viability. The nuclear enzyme poly(ADP-ribose)polymerase (PARP) was also cleaved upon the activation of caspases, and IETD similarly prevented PARP cleavage. The PARP inhibitor 3-aminobenzamide (3-AB) restored the cell killing in the presence of IETD, an effect that occurred without restoration of the cleavage of Bid or PARP. In the presence of 3-AB and IETD, translocation occurred of full-length Bid to the mitochondria. The induction of the mitochondrial permeability transition (MPT) was documented by the cyclosporin A (CyA) sensitivity of the release of cytochrome c, the release of malate dehydrogenase from the mitochondrial matrix, the loss of the mitochondrial membrane potential, and the pronounced swelling of these organelles, as assessed by electron microscopy. In addition to preventing all evidence of the MPT, CyA prevented the loss of cell viability, without effect on the cleavage of either Bid or PARP. The prevention of PARP cleavage by inhibition of caspase-3 resulted in a 10-fold activation of the enzyme and a resultant depletion of NAD and ATP. The PARP inhibitor 3-AB prevented the loss of NAD and ATP. Depletion of ATP by metabolic inhibitors similarly prevented the cell killing. It is concluded that the cleaving of PARP in Fas-mediated apoptosis allowed expression of an energy-dependent cell death program that included the translocation of full-length Bid to the mitochondria with induction of the MPT.
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PMID:Cytochrome c release upon Fas receptor activation depends on translocation of full-length bid and the induction of the mitochondrial permeability transition. 1179 Jul 91

Poly(ADP-ribose) polymerase-1 (PARP-1) is a chromatin-associated enzyme that is activated by DNA strand breaks and catalyzes the transfer of ADP-ribose groups from NAD to itself and other nuclear proteins. Although caspase-mediated PARP-1 cleavage occurs during almost all forms of apoptosis, the contribution of PARP-1 activation and cleavage to this cell death process remains unclear. Using immortalized fibroblasts from wild-type (PARP-1(+/+)) and PARP-1 knockout (PARP-1(-/-)) mice, and a mouse neuroblastoma cell line (N18), the role that poly(ADP-ribosyl)ation plays in Sindbis virus (SV)-induced apoptosis was examined. Robust PARP-1 activation occurred in SV-infected cells prior to morphologic changes associated with apoptotic cell death and PARP-1 activity ceased simultaneously with caspase-3 activation and PARP-1 proteolysis. PARP-1 activity was maximal before detectable DNA fragmentation, but was absent when DNA damage was most intense. SV and staurosporine-induced cell death was delayed in fibroblasts lacking PARP-1 activity, suggesting that PARP-1 activation contributes to apoptotic cell death induced by these stimuli. SV replication was not affected by lack of PARP-1 activity, but DNA fragmentation and caspase-3 activation were delayed and occurred at lower levels in PARP-1-deficient fibroblasts. Early virus-induced PARP-1 activation may represent a novel way by which cells signal to the nucleus to regulate protein function by poly(ADP-ribosyl)ation in response to virus infection.
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PMID:Rapid activation of poly(ADP-ribose) polymerase contributes to Sindbis virus and staurosporine-induced apoptotic cell death. 1185 9

Disruption of mitochondrial electron transport and opening of the so-called mitochondrial permeability transition pores (PTPs) are early events in apoptotic cell death and may be caused by the uncoupler of mitochondrial oxidation and phosphorylation, carbonyl cyanide p-trifluoromethoxyphenylhydrazone (FCCP). We investigated the cellular toxicity of FCCP in HL60 and CCRF-CEM cells alone or in combination with the known apoptosis inducers such as inhibitor of serine/threonine protein kinases staurosporine (Sts) and protein kinase C inhibitor chelerythrine. FCCP induced apoptotic cell death in both cell lines in a dose-dependent manner, and we were able to demonstrate an appearance of caspase-3-dependent PARP cleavage fragments with Western blot and the appearance of large (15-50 kb) DNA fragments using pulsed-field gel electrophoresis. After 2 hr of incubation with Che or Sts more than half of the cells had died by apoptosis. We observed a statistically significant delay in Sts- and Che-induced apoptotic cell death in CCRF-CEM cells when the cells were preincubated with FCCP but not with zVAD-FMK: about 50% more cells survived after pre-treatment with FCCP, as compared to 1 hr treatment with Che alone (P<0.05), and 25% more cells were alive after 6 hr of treatment, as compared to 6 hr exposure to Sts alone (P<0.05). The protective effect of FCCP was, however, transient and lasted only 6 hr. Treatment with aurintricarboxylic acid completely prevented Che- and Sts-induced apoptotic cell death in CCRF-CEM and HL60 cells. Incubation with Che resulted in a drop in the intracellular ATP content, predominantly distinctive in HL60, and in NAD(+) content in CCRF-CEM cells. Both ATP and NAD(+) drop were prevented with ATA, but not with FCCP or zVAD. Our data suggest that treatment with uncouplers of oxidative phosphorylation can induce apoptotic cell death in haematopoietic cell lines. However, when used in combination with serine/threonine protein kinase inhibitors FCCP can even prevent apoptosis.
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PMID:Modulation of apoptosis by mitochondrial uncouplers: apoptosis-delaying features despite intrinsic cytotoxicity. 1185 98

We recently developed a class of novel antitumor agents that elicit a potent growth-inhibitory response in many tumor cells cultured in vitro. WK175, a member of this class, was chosen as a model compound that showed strong in vitro efficacy. WK175 interferes with the intracellular steady-state level of NAD(+), resulting in a decreased cellular NAD(+) concentration. We found that WK175 induces apoptotic cell death without any DNA-damaging effect. The apoptotic death signaling pathway initiated by WK175 was examined in detail: mitochondrial membrane potential, cytochrome c release, caspase 3 activation, caspase 3 and poly(ADP-ribose) polymerase cleavage, and the appearance of a sub-G(1) cell cycle population were determined in time course studies in THP-1 (a human monocytic leukemia cell line) cells. We found activation of this cascade after 24 h of treatment with 10 nM WK175. Induction of apoptosis was prevented by bongkrekic acid, Z-Asp-Glu-Val-Asp-fluoromethylketone, and Z-Leu-Glu-His-Asp-fluoromethylketone, inhibitors of the mitochondrial permeability transition and of caspase 3 and 9, respectively, but not by Ac-Tyr-Val-Ala-Asp-CHO, a specific caspase 1 inhibitor, suggesting the involvement of the permeability transition pore, caspase 3, and caspase 9 in the WK175-induced apoptotic cascade. These results imply that decreased NAD(+) concentration initiates the apoptotic cascade, resulting in the antitumor effect of WK175.
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PMID:WK175, a novel antitumor agent, decreases the intracellular nicotinamide adenine dinucleotide concentration and induces the apoptotic cascade in human leukemia cells. 1186 82

TAS-103, a new anticancer drug, induces DNA cleavage by inhibiting the activities of topoisomerases I and II. We investigated the mechanism of TAS-103-induced apoptosis in human cell lines. Pulsed field gel electrophoresis revealed that in the leukemia cell line HL-60 and the H(2)O(2)-resistant subclone, HP100, TAS-103 induced DNA cleavage to form 1-2-Mb fragments at 1 h to a similar extent, indicating that the DNA cleavage was induced independently of H(2)O(2). TAS-103-induced DNA ladder formation in HP100 cells was delayed compared with that seen at 4 h in HL-60 cells, suggesting the involvement of H(2)O(2)-mediated pathways in apoptosis. Flow cytometry revealed that H(2)O(2) formation preceded increases in mitochondrial membrane potential (DeltaPsim) and caspase-3 activation. Inhibitors of poly(ADP-ribose) polymerase (PARP) prevented both TAS-103-induced H(2)O(2) generation and DNA ladder formation. The levels of NAD(+), a PARP substrate, were significantly decreased in HL-60 cells after a 3-h incubation with TAS-103. The decreases in NAD(+) levels preceded both increases in DeltaPsim and DNA ladder formation. Inhibitors of NAD(P)H oxidase prevented TAS-103-induced apoptosis, suggesting that NAD(P)H oxidase is the primary enzyme mediating H(2)O(2) formation. Expression of the antiapoptotic protein, Bcl-2, in BJAB cells drastically inhibited TAS-103-induced apoptosis, confirming that H(2)O(2) generation occurs upstream of mitochondrial permeability transition. Therefore, these findings indicate that DNA cleavage by TAS-103 induces PARP hyperactivation and subsequent NAD(+) depletion, followed by the activation of NAD(P)H oxidase. This enzyme mediates O(2)(-)-derived H(2)O(2) generation, followed by the increase in DeltaPsim and subsequent caspase-3 activation, leading to apoptosis.
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PMID:Mechanism of apoptosis induced by a new topoisomerase inhibitor through the generation of hydrogen peroxide. 1206 15

Toxic reactive oxygen species (ROS) such as hydrogen peroxide, nitric oxide, superoxide, and the hydroxyl radical are generated in a variety of neuropathological conditions and cause significant DNA damage. We determined the effects of 3-aminobenzamide (AB), an inhibitor of the DNA repair enzyme poly(ADP-ribose) polymerase (PARP), on cell death in differentiated PC12 cells, a model of sympathetic neurons, after H(2) O(2) injury. Exposure to 0.5 mm H(2) O(2) resulted in a significant decrease in intracellular NAD(H), NADP(H), and ATP levels. This injury resulted in the death of 90% of the cells with significant necrosis early (2 h) after injury and increased apoptosis (12-24 h after injury), as measured by PS exposure and the presence of cytoplasmic oligonucleosomal fragments. Treatment with 2.5 mm AB restored pyridine nucleotide and ATP levels and ameliorated cell death (65% versus 90%) by decreasing the extent of both necrosis and apoptosis. Interestingly, we observed that H(2) O(2) -induced injury caused a delayed cell death exhibiting features of apoptosis but in which caspase-3 like activity was absent. Moreover, pretreatment with AB restored caspase-3-like activity. Our results suggest that apoptosis and necrosis are both triggered by PARP overactivation, and that maintenance of cellular energy levels after injury by inhibiting PARP shifts cell death from necrosis to apoptosis.
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PMID:Poly(ADP-ribose) polymerase inhibition prevents both apoptotic-like delayed neuronal death and necrosis after H(2)O(2) injury. 1209 61

Previous studies have shown that the liver is the first organ to display signs of injury during hemorrhagic shock. We examined the mechanism by which pyruvate can prevent liver damage during hemorrhagic shock in swine anesthetized with halothane. Thirty minutes after the induction of a 240-min controlled arterial hemorrhage targeted at 40 mmHg, hypertonic sodium pyruvate (0.5 g. kg(-1). h(-1)) was infused to achieve an arterial concentration of 5 mM. The volume and osmolality effects of pyruvate were matched with 10% saline (HTS) and 0.9% saline (NS). Although the peak hemorrhage volume increased significantly in both the pyruvate and HTS group, only the pyruvate treatment was effective in delaying cardiovascular decompensation. In addition, pyruvate effectively maintained the NADH/NAD redox state, as evidenced by increased microdialysate pyruvate levels and a significantly lower lactate-to-pyruvate ratio. Pyruvate also prevented the loss of intracellular antioxidants (GSH) and a reduction in the GSH-to-GSSG ratio. These beneficial effects on the redox environment decreased hepatic cellular death by apoptosis. Pyruvate significantly increased the ratio of Bcl-Xl (antiapoptotic molecule)/Bax (proapoptotic molecule), prevented the release of cytochrome c from mitochondria, and decreased the fragmentation of caspase 3 and poly(ADP ribose) polymerase (DNA repair enzyme). These beneficial findings indicate that pyruvate infused 30 min after the onset of severe hemorrhagic shock is effective in maintaining the redox environment, preventing the loss of the key antioxidant GSH, and decreasing early apoptosis indicators.
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PMID:Pyruvate improves redox status and decreases indicators of hepatic apoptosis during hemorrhagic shock in swine. 1223 18

The regulation of enzyme activity function is a major factor in the cellular response to a changing environment. One mechanism of enzyme activity regulation includes post-translational protein thiol modification by nitric oxide (NO) or its redox species. Major routs used by NO to modify cysteine residues of proteins include S-nitrosation, oxidation, mixed disulfide formation with glutathione, and the covalent attachment of nucleotide cofactors, i.e NAD(+)/NADH. Critical thiol centers serve as recognition sites for NO, thus channeling the NO signal through post-translational modifications and oxidation into cellular functions. Here, we summarize current knowledge on active site thiol modification of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and caspase-3 by nitric oxide. Although very different in their cellular function, both enzymes contain highly reactive cysteines which represent sensitive targets for NO. Our studies are supportive of a potential role of S-nitrosation and mixed disulfide formation as a general signaling mechanism that allows sensing of nitrosative stress. At the same time, modification of GAPDH and caspase-3 by NO show the diversity of mechanisms (S-nitrosation versus oxidations) that we are confronted with as a result of NO delivery, especially comparing in vitro studies with cellular systems. In the future it will be challenging to dissect how nitrosative and oxidative signaling mechanisms overlap and how intracellular communication systems allow their activation in a selective way.
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PMID:Protein thiol modification of glyceraldehyde-3-phosphate dehydrogenase and caspase-3 by nitric oxide. 1236 1

Aldehyde dehydrogenase 3A1 (ALDH3A1) is one of the most abundant proteins found in corneal epithelial cells of mammalian species, with several postulated protective roles that include detoxification of peroxidic aldehydes, scavenging of free radicals, and direct absorption of ultraviolet (UV) radiation. In the present study, the protective role of ALDH3A1 against UV- and 4-hydroxy-2-nonenal- (4-HNE-) induced oxidative damage was studied. For this purpose, human ALDH3A1 was stably transfected in a human corneal epithelial cell line (HCE) lacking endogenous enzyme. Cells transfected with ALDH3A1 were more resistant to UV- and 4-HNE-induced cytotoxicity than mock-transfected cells. DNA fragmentation assays revealed that both treatments induced apoptosis in mock-transfected cells, but not in ALDH3A1-expressing cells. Apoptosis appeared to occur via caspase-3 activation and subsequent PARP cleavage. The Michaelis-Menten constant (K(m)) for 4-HNE was 54 microM in ALDH3A1-transfected cells; the addition of 100 microM 4-HNE increased NAD(P)H levels by 50% above that in mock-transfected cells. We also found that ALDH3A1 expression prevented 4-HNE-induced protein adduct formation. Taken together, these data suggest that ALDH3A1 is a regulatory element of the cellular defense system that protects corneal epithelium against UV- and 4-HNE-induced oxidative damage.
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PMID:Aldh3a1 protects human corneal epithelial cells from ultraviolet- and 4-hydroxy-2-nonenal-induced oxidative damage. 1270 98


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