Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.22.56 (caspase-3)
 35,750 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Children with congenital homozygous deficiency of purine nucleoside phosphorylase (PNP) have abnormalities in purine metabolism that result in T-cell selective immune deficiency. The mechanism of action for cell death has been attributed to intracellular accumulation of dGTP, a potent inhibitor of ribonucleotide reductase and subsequently DNA synthesis, in thymocytes and T-cells but not B-cells. However, the mode of cell death has not been determined to be either necrosis or apoptosis. To examine the involvement of apoptosis in T-cells following PNP inhibition, MOLT-4 cells, a human T cell leukemia cell line, were co-treated with the PNP inhibitor, CI-1000 (2-amino 3,5-dihydro-7-(3-thienylmethyl)-4H-pyrrolo[3,2-d]-pyrimidin-4-one HCl), and 2'-deoxyguanosine (dGuo) which resulted in a concentration-dependent loss of cell viability (trypan blue) and inhibition of tritiated thymidine ([3H]-TdR) uptake. Staining of cells with the DNA dye Hoechst 33,258 showed nuclear morphology characteristic of apoptosis. Western blots (24 h lysates) were probed with antibodies against several proteins implicated in apoptosis. Anti-PARP revealed the presence of an 85 kD PARP breakdown product while, anti-alpha-spectrin revealed the accumulation a 120 kD breakdown product, both suggestive of CPP32 cleavage (caspase-3; an ICE-like cysteine protease). Western blots also detected the loss of the intact 32 kD caspase-3 isoform, a biochemical event associated with caspase-3 activation. Corresponding fluorometric activity assays detected a marked increase in caspase-3-like activity using the substrate Ac-DEVD-MCA. Lastly, a pan caspase inhibitor (Z-D-DCB) and 2'-deoxycytidine (dCyd), which is known to prevent dGTP accumulation following PNP inhibition, were able to prevent cell death and all indicators of caspase-3-like activity in MOLT-4 cells co-treated with dGuo and CI-1000. In summary, we provided several lines of evidence for the role of apoptosis and the contribution of caspase-3-like proteases in T-cell death following PNP inhibition.
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PMID:A purine nucleoside phosphorylase (PNP) inhibitor induces apoptosis via caspase-3-like protease activity in MOLT-4 T cells. 940 42

The mechanism of apoptosis induced by 2-chloro-2'-deoxyadenosine (2CdA) in human leukemia cell line MOLT-4 was investigated. 2CdA induced increases of 3'-OH ends of genomic DNA, ladder-like DNA fragmentation and phosphatidylserine translocation to the outer membrane, which are apoptotic characteristics. These apoptotic phenomena induced by 2CdA were inhibited by cycloheximide (CHX; a protein synthesis inhibitor), deoxycytidine (dC; a substrate of deoxycytidine kinase), acetyl Ile-Glu-Thr-Asp aldehyde (Ac-IETD-CHO; a caspase-8 inhibitor) and acetyl Asp-Glu-Val-Asp aldehyde (Ac-DEVD-CHO; a caspase-3 inhibitor). The protein synthesis-dependent expression of Fas and Fas ligand (Fas-L) was detected by treatment with 2CdA. The proteolytic processing of procaspases-8 and -3 to produce active fragments, caspases-8 (p18) and -3 (p17), respectively, was observed after treatment with 2CdA, and suppressed by cycloheximide. Increases in the activities of caspases-8 and -3 were observed after 2CdA treatment. Their activation was also dependent on protein synthesis. These results indicated that 2CdA-induced apoptosis was triggered by phosphorylation of 2CdA followed by the protein synthesis-dependent expression of Fas and Fas-L and activation of caspases-8 and -3.
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PMID:2-Chloro-2'-deoxyadenosine induces apoptosis through the Fas/Fas ligand pathway in human leukemia cell line MOLT-4. 1067 48

Methylglyoxal (MG) is a physiological metabolite, but it is known to be toxic, inducing stress in cells and causing apoptosis. This study examines molecular mechanisms in the MG-induced signal transduction leading to apoptosis, focusing particularly on the role of JNK activation. We first confirmed that MG caused apoptosis in Jurkat cells and that it was cell type dependent because it failed to induce apoptosis in MOLT-4, HeLa, or COS-7 cells. A caspase inhibitor, Z-DEVD-fmk, completely blocked MG-induced poly(ADP-ribose)polymerase (PARP) cleavage and apoptosis, showing the critical role of caspase activation. Inhibition of JNK activity by a JNK inhibitor, curcumin, remarkably reduced MG-induced caspase-3 activation, PARP cleavage, and apoptosis. Stable expression of the dominant negative mutant of JNK also protected cells against apoptosis notably, although not completely. Correspondingly, loss of the mitochondrial membrane potential induced by MG was decreased by the dominant negative JNK. These results confirmed a crucial role of JNK working upstream of caspases, as well as an involvement of JNK in affecting the mitochondrial membrane potential.
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PMID:Methylglyoxal induces apoptosis in Jurkat leukemia T cells by activating c-Jun N-terminal kinase. 1072 98

The ubiquitin-proteasome pathway is responsible for selective degradation of short-lived cellular proteins and is critical for the regulation of many cellular processes. We previously showed that ubiquitin (Ub) secreted from hairy cell leukemia cells had inhibitory effects on clonogenic growth of normal hematopoietic progenitor cells. In this study, we examined the effects of exogenous Ub on the growth and survival of a series of human hematopoietic cells, including myeloid cell lines (HL-60 and U937), a B-cell line (Daudi), and T-cell lines (KT-3, MT-4, YTC-3, and MOLT-4). Exogenous Ub inhibited the growth of various hematopoietic cell lines tested, especially of KT-3 and HL-60 cells. The growth-suppressive effects of Ub on KT-3 and HL-60 cells were almost completely abrogated by the proteasome inhibitor PSI or MG132, suggesting the involvement of the proteasome pathway in this process. Furthermore, exogenous Ub evoked severe apoptosis of KT-3 and HL-60 cells through the activation of caspase-3. In interleukin-6 (IL-6)-dependent KT-3 cells, STAT3 was found to be conjugated by exogenous biotinylated Ub and to be degraded in a proteasome-dependent manner, whereas expression levels of STAT1, STAT5, or mitogen-activated protein kinase were not affected. Moreover, IL-6-induced the up-regulation of Bcl-2 and c-myc, and JunB was impaired in Ub-treated KT-3 cells, suggesting that the anti-apoptotic and mitogenic effects of IL-6 were disrupted by Ub. These results suggest that extracellular Ub was incorporated into hematopoietic cells and mediated their growth suppression and apoptosis through proteasome-dependent degradation of selective cellular proteins such as STAT3. (Blood. 2000;95:2577-2585)
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PMID:Induction of apoptosis by extracellular ubiquitin in human hematopoietic cells: possible involvement of STAT3 degradation by proteasome pathway in interleukin 6-dependent hematopoietic cells. 1075 37

Caspases, a family of cysteine proteases, play a central role in the pathways leading to apoptosis. Recently, it has been reported that a broad spectrum inhibitor of caspases, the tripeptide Z-VAD-fmk, induced a switch from apoptosis to necrosis in dexamethasone-treated B lymphocytes and thymocytes. As such a cell death conversion could increase the efficiency of radiation therapy and in order to identify the caspases involved in this cell death transition, we investigated the effects of caspase-3-related proteases inhibition in irradiated MOLT-4 cells. Cells were pretreated with Ac-DEVD-CHO, an inhibitor of caspase-3-like activity, and submitted to X-rays at doses ranging from 1 to 4 Gy. Our results show that the inhibition of caspase-3-like activity prevents completely the appearance of the classical hallmarks of apoptosis such as internucleosomal DNA fragmentation or hypodiploid particles formation and partially the externalization of phosphatidylserine. However, this was not accompanied by any persistent increase in cell survival. Instead, irradiated cells treated by this inhibitor exhibited characteristics of a necrotic cell death. Therefore, functional caspase-3-subfamily not only appears as key proteases in the execution of the apoptotic process, but their activity may also influence the type of cell death following an exposure to ionizing radiation.
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PMID:Caspase-3-like activity determines the type of cell death following ionizing radiation in MOLT-4 human leukaemia cells. 1094 6

The alkylating agent, nitrogen mustard (HN2), is thought to cause apoptosis through production of free oxygen radicals. To explore the mechanism of HN2-induced apoptosis, we utilized ebselen, a selenoorganic compound with potent antioxidant activity. We examined whether ebselen would inhibit apoptosis in BALB/c mouse spleen lymphocytes and human MOLT-4 leukemia cells treated with HN2 (2.5 microM) in vitro. Non-toxic concentrations (<50 microM) of ebselen were found to prevent HN2-induced apoptosis of murine lymphocytes in a dose-dependent manner, as measured by cell viability, hypodiploid DNA formation, and phosphatidylserine externalization. However, ebselen was ineffective at preventing spontaneous apoptosis in these cells, pointing to the selectivity of its action. Furthermore, pretreatment with ebselen at 1-10 microM for 72 hr protected MOLT-4 cells from HN2-induced apoptosis and maintained cell viability and proliferation as monitored by the above-mentioned parameters. This was accompanied by the preservation of mitochondrial transmembrane potential and elevated glutathione levels and by a blockage of caspase-3 and -9 activation. In vivo, ebselen also had a marked protective effect against spleen weight loss associated with lymphocyte apoptosis in mice treated by HN2. Therefore, ebselen provides an efficient protection against HN2-induced cell death in normal and tumoral lymphocytes and might prove useful as an antidote against alkylating agents.
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PMID:Prevention of nitrogen mustard-induced apoptosis in normal and transformed lymphocytes by ebselen. 1107 38

Lipoproteins of Mycoplasma salivarium and Mycoplasma fermentans preferentially induced necrotic cell death in lymphocytic cell lines, MOLT-4 and Raji, and in one monocytic cell line, THP-1, whereas they preferentially induced apoptotic cell death in another monocytic cell line, HL-60. These findings were also supported by ultrastructural observations by the use of scanning and transmission electron microscopes and by agarose gel electrophoresis of the genomic DNA. The lipoproteins activated caspase-3 in both MOLT-4 and HL-60 cells, which was assessed by the cleavage of the synthetic substrate DEVD-pNA and the endogenous substrate poly(ADP-ribose) polymerase. The cytotoxicity to MOLT-4 and HL-60 cells was inhibited by various caspase inhibitors, Ac-DMQD-CHO, Ac-IETD-CHO, and Z-VAD-FMK. The cytotoxicity was also partially suppressed by the monoclonal antibody to Toll-like receptor 2. Thus this study demonstrated that mycoplasmal lipoproteins induce caspases-dependent necrotic and apoptotic cell death in lymphocytes and monocytes/macrophages, which is partially induced by TLR2-mediated signaling.
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PMID:Mycoplasmal lipoproteins induce toll-like receptor 2- and caspases-mediated cell death in lymphocytes and monocytes. 1206 29

We have previously reported that in a MOLT-4 leukemia cell line the acquired resistance to 9-beta-D-arabinofuranosylguanine (Ara-G) is due to deficiency of the activating enzymes deoxyguanosine kinase and deoxycytidine kinase [Biochem. Biophys. Res. Commun. 293 (5) (2002) 1489]. In this study we investigated whether apoptotic pathways are affected in two human T-cell lymphoblastic MOLT-4 cell lines with acquired resistance to Ara-G. In contrast to the MOLT-4 wild type cells, Ara-G resistant cells displayed no increase in caspase-3 or caspase-9 activity, DNA fragmentation, cytochrome c release or a drop in the mitochondrial membrane potential (DeltaPsi(mito)) upon Ara-G treatment. A drop in the DeltaPsi(mito) was induced in wild type cells after treatment with tributyltin, an inducer of mitochondrial permeability transition, and with carbonyl cyanide m-chlorophenylhydrazone, an uncoupling agent that reduces the DeltaPsi(mito), although not in Ara-G resistant cells. Ara-G resistant cells displayed higher levels of the anti-apoptotic protein Bcl-xL in immunoblots. A recent study indicates that Ara-G-induced apoptosis is mediated in part via the Fas pathway [Cancer Res. 43 (2047) (2002) 411]. When cells were treated with anti-Fas antibody, the wild type cell line exhibited increased caspase-3-like activity but the Ara-G resistant cells did not. Using FACS analysis and semi-quantitative PCR, 3-6-fold decreased protein levels and almost no detectable mRNA levels of Fas in the resistant cells were recorded. These data indicate that the inability to induce apoptosis via both the apoptosome pathway and the Fas pathway, due to increased levels of Bcl-xL and a lack of Fas, contributes to Ara-G resistance. This resistance to apoptosis in Ara-G resistant cells may serve to explain the overall resistance to a variety of anti-neoplastic drugs.
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PMID:Resistance to mitochondrial- and Fas-mediated apoptosis in human leukemic cells with acquired resistance to 9-beta-D-arabinofuranosylguanosine. 1241 45

It has been demonstrated that caspase-3 is responsible for determining the mode of cell death, i.e., apoptosis or necrosis. To characterize the mode of cell death induced by the inhibition of caspase-3, we have studied the effects of Ac-DEVD-CHO, Ac-YVAD-CHO, and Ac-IETD-CHO, inhibitors of caspases, on structural changes in X-irradiated human leukemic MOLT-4 cells. When cells were irradiated with X-rays and incubated in the presence of Ac-DEVD-CHO, the expression of cell death, as measured by the dye exclusion test, was inhibited, whereas no such change was observed in colony-forming ability. The hallmarks of apoptosis, i.e., nuclear condensation and DNA ladder formation, were depressed. However, a new type of nuclear morphology appeared. The sum of the frequencies of apoptosis and this new type of nuclear structure corresponded to the frequency of X-ray-induced apoptosis for cells incubated in the absence of Ac-DEVD-CHO. Removal of Ac-DEVD-CHO during the course of post-irradiation incubation increased apoptotic nuclear condensation accompanied by a slight decrease in the frequency of the new type of nuclear structure. When Ac-IETD-CHO was used in place of Ac-DEVD-CHO, inhibition of cell death (apoptosis) was also observed, but not in the case of Ac-YVAD-CHO. These results suggest that the inhibition of caspase-3 diminishes the expression of apoptotic hallmarks with no effect on cell survival, that the morphology observed in the presence of Ac-DEVD-CHO is an apoptosis-related structure, and that the cell death observed is a programmed cell death independent of caspase-3. The development of this mode of cell death was slower than that of apoptosis by 4 h.
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PMID:Time sequence analysis of caspase-3-independent programmed cell death and apoptosis in X-irradiated human leukemic MOLT-4 cells. 1245 29

The cytotoxicities of lipoproteins of Mycoplasma fermentans and Mycoplasma salivarium to a lymphocytic cell line, MOLT-4, and a monocytic cell line, HL-60, was upregulated by ATP added extracellularly in a dose-dependent manner. These lipoproteins induced ATP release and plasma membrane permeability increase in these cell lines. In addition, periodate-oxidized ATP, an antagonist for P2X purinergic receptors, suppressed the cytotoxicity of the lipoproteins, suggesting the possibility that P2X receptors for ATP play crucial roles in the cytotoxicity. Activation of caspase-3 induced by the lipoproteins, which was assessed by the cleavage of the synthetic substrate DEVD-pNA and the endogenous substrate poly(ADP-ribose) polymerase, was also upregulated and downregulated by extracellular ATP and periodate-oxidized ATP, respectively. On the basis of these results, this study suggests that mycoplasmal lipoproteins induce the permeability increase in lymphocytes and monocytes, by which ATP is released, and the ATP regulates the cytotoxicities of the lipoproteins to the cells, possibly by interaction with ATP receptors such as P2X purinergic receptors.
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PMID:Extracellular ATP regulates cell death of lymphocytes and monocytes induced by membrane-bound lipoproteins of Mycoplasma fermentans and Mycoplasma salivarium. 1247 45


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