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)

Tumor necrosis factor (TNF) is one of the most potent activators of nuclear transcription factor NF-kappaB, c-Jun N-terminal protein kinase (JNK), and apoptosis in a wide variety of cells. The biological effects of TNF are mediated through sequential interactions of various cytoplasmic proteins with intracellular domains of TNF receptors. Whether signal transducer and activator of transcription-1 (STAT1), which mediates interferon (IFN) signaling, also plays any role in the TNF-mediated activation of NF-kappaB, JNK, and apoptosis has not been established. Here, we report our investigation of the role of STAT1 in TNF signaling using STAT1-deficient U3A and STAT1-stably transfected U3A-PSG91 cells. IFNalpha inhibited the proliferation of STAT1-expressing U3A-PSG91 cells but had no effect on STAT1-negative U3A cells. TNF alone, even up to 10 nM, had no effect on the proliferation of either U3A-PSG91 or U3A cells. Irrespective of STAT1 status, TNF induced cytotoxic effects in the presence of cycloheximide (CHX) in both cell types. Additionally, TNF-induced caspase-3 and caspase-8 activation and TNF-induced PARP cleavage were unaffected by the presence or absence of STAT1. TNF activated NF-kappaB, consisting of p50 and p65, in both U3A and U3A-pSG91 cells in a dose- and time-dependent manner, but the degree and rate of activation were slightly lower in U3A cells, as were IkappaBalpha degradation and NF-kappaB-dependent reporter gene expression. STAT1 was, however, required for IFNalpha-mediated downregulation of TNF-induced NF-kappaB activation. TNF activated JNK in both cell types, but dose and time of exposure required for optimum activation differed slightly. Thus, overall our results indicate that STAT1 plays a minimal role in TNF-mediated cellular responses.
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PMID:Lack of requirement of STAT1 for activation of nuclear factor-kappaB, c-Jun NH2-terminal protein kinase, and apoptosis by tumor necrosis factor-alpha. 1183 5

TNF family receptors can lead to the activation of NF-kappaB and this can be a prosurvival signal in some cells. Although activation of NF-kappaB by ligation of Fas (CD95/Apo-1), a member of the TNFR family, has been observed in a few studies, Fas-mediated NF-kappaB activation has not previously been shown to protect cells from apoptosis. We examined the Fas-induced NF-kappaB activation and its antiapoptotic effects in a leukemic eosinophil cell line, AML14.3D10, an AML14 subline resistant to Fas-mediated apoptosis. EMSA and supershift assays showed that agonist anti-Fas (CH11) induced nuclear translocation of NF-kappaB heterodimer p65(RelA)/p50 in these cells in both a time- and dose-dependent fashion. The influence of NF-kappaB on the induction of apoptosis was studied using pharmacological proteasome inhibitors and an inhibitor of IkappaBalpha phosphorylation to block IkappaBalpha dissociation and degradation. These inhibitors at least partially inhibited NF-kappaB activation and augmented CH11-induced cell death. Stable transfection and overexpression of IkappaBalpha in 3D10 cells inhibited CH11-induced NF-kappaB activation and completely abrogated Fas resistance. Increases in caspase-8 and caspase-3 cleavage induced by CH11 and in consequent apoptotic killing were observed in these cells. Furthermore, while Fas-stimulation of resistant control 3D10 cells led to increases in the antiapoptotic proteins cellular inhibitor of apoptosis protein-1 and X-linked inhibitor of apoptosis protein, Fas-induced apoptosis in IkappaBalpha-overexpressing cells led to the down-modulation of both of these proteins, as well as that of the Bcl-2 family protein, Bcl-x(L). These data suggest that the resistance of these leukemic eosinophils to Fas-mediated killing is due to induced NF-kappaB activation.
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PMID:Fas resistance of leukemic eosinophils is due to activation of NF-kappa B by Fas ligation. 1224 43

BAG-1 protein can be expressed as four isoforms of 50, 46, 33 and 29 kDa with different subcellular localizations, which may have different functions in anti-apoptosis, but the exact mechanism remains unclear. We constructed BAG-1 full length and deletion mutated plasmids in a pCR3.1 vector and established stable transfections of BAG-1 isoforms in low BAG-1 expressing C33A cells. Treatment of the transfected cells with cisplatin, staurosporine, paclitaxel and doxorubicine showed that BAG-1 p50, p46 and p33 isoforms enhanced the resistance to apoptosis. BAG-1 p50, p46 and p33 exhibited different degrees of apoptosis inhibition in the transfected cells and BAG-1 p46 isoform had the most pronounced effect on anti-apoptosis. BAG-1 p29 failed to protect the transfected cells from apoptosis. Resistance to apoptosis by BAG-1 isoforms was correlated with decreased caspase-3 activation. We also detected the expression of Bax, Bak, p53, Bcl-2, Bcl-X(L), AIF and MRP1 by Western blots. Bcl-2 protein expression was significantly increased in p50, p46 and p33 transfected cells, while the expression of Bax, Bak, p53, Bcl-X(L) and MRP1 was essentially unchanged. These in vitro results suggest that distinct isoforms of BAG-1 have different anti-apoptotic functions and their functions may be correlated to increased Bcl-2 expression.
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PMID:Distinct BAG-1 isoforms have different anti-apoptotic functions in BAG-1-transfected C33A human cervical carcinoma cell line. 1237 Aug 27

Reactive oxygen species are recognized as important mediators of biological responses. Hyperglycemia promotes the intracellular generation of superoxide anion and hydrogen peroxide. In several cell lines, oxidant stress has been linked to the activation of death programs. Here, we report for the first time that high ambient glucose concentration induces apoptosis in murine and human mesangial cells by an oxidant-dependent mechanism. The signaling cascade activated by glucose-induced oxidant stress included the heterodimeric redox-sensitive transcription factor NF-kappaB, which exhibited an upregulation in p65/c-Rel binding activity and suppressed binding activity of the p50 dimer. Recruitment of NF-kappaB and mesangial cell apoptosis were both inhibited by antioxidants, implicating oxidant-induced activation of NF-kappaB in the transmission of the death signal. The genetic program for glucose-induced mesangial cell apoptosis was characterized by an upregulation of the Bax/Bcl-2 ratio. In addition, phosphorylation of the proapoptotic protein Bad was attenuated in mesangial cells maintained at high-glucose concentration, favoring progression of the apoptotic process. These perturbations in the expression and phosphorylation of the Bcl-2 family were coupled with the release of cytochrome c from mitochondria and caspase activation. Our findings indicate that in mesangial cells exposed to high ambient glucose concentration, oxidant stress is a proximate event in the activation of the death program, which culminates in mitochondrial dysfunction and caspase-3 activation, as the terminal event.
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PMID:High glucose promotes mesangial cell apoptosis by oxidant-dependent mechanism. 1241 73

Evidence from live cell bioassays shows that the flat mucosa from patients with colon cancer exhibits resistance to bile salt-induced apoptosis. Three independent cell lines derived from the colonic epithelial cell line HCT-116 were selected for resistance to bile salt-induced apoptosis. These cell lines were developed as tissue culture models of apoptosis resistance. Selection was carried out for resistance to apoptosis induced by sodium deoxycholate (NaDOC), the bile salt found in highest concentrations in human fecal water. Cultures of HCT-116 cells were serially passaged in the presence of increasing concentrations of NaDOC. The resulting apoptosis resistant cells were able to grow at concentrations of NaDOC (0.5 mM) that cause apoptosis in a few hours in unselected HCT-116 cells. These cells were then analyzed for changes in gene expression. Observations from cDNA microarray, 2-D gel electrophoresis/MALDI-mass spectroscopy, and confocal microscopy of immunofluorescently stained preparations indicated underexpression or overexpression of numerous genes at either the protein or mRNA level. Genes that may play a role in apoptosis and early stage carcinogenesis have been identified as upregulated in these cell lines, including Grp78, Bcl-2, NF-kappaB(p50), NF-kappaB(p65), thioredoxin peroxidase (peroxiredoxin) 2, peroxiredoxin 4, maspin, guanylate cyclase activating protein-1, PKCzeta, EGFR, Ras family members, PKA, PI(4,5)K, TRAF2 and BIRC1 (IAP protein). Under-expressed mRNAs included BNIP3, caspase-6, caspase-3 and serine protease 11. NF-kappaB was constitutively activated in all three resistant cell lines, and was responsible, in part, for the observed apoptosis resistance, determined using antisense oligonucleotide strategies. Molecular and cellular analyses of these resistant cell lines has suggested potential mechanisms by which apoptosis resistance may develop in the colonic epithelium in response to high concentrations of hydrophobic bile acids that are associated with a Western-style diet. These analyses provide the rationale for the development of hypothesis-driven intermediate biomarkers to assess colon cancer risk on an individual basis.
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PMID:Development and molecular characterization of HCT-116 cell lines resistant to the tumor promoter and multiple stress-inducer, deoxycholate. 1250 30

The mechanism of induction of apoptosis by double-stranded RNA (dsRNA) is not fully characterized. The dsRNA is normally present in extremely low quantities in cells, but following infection with RNA viruses, large quantities of the dsRNA viral replicative intermediate may be produced triggering the antiviral response as well as cell death. In this report, transfection of polyinosinic-polycytidylic acid [poly(I:C)] into NIT 1 cells has been used as a model of intracellular dsRNA-induced beta-cell apoptosis. At 18 h post transfection, 45% of the cells were apoptotic as indicated by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end-labeling (TUNEL) staining, and this was accompanied by an increase in nuclear factor kappaB (NF-kappaB) p50/p65 nuclear translocation and cleavage of caspases 3 and 8. The NF-kappaB inhibitor peptide, SN50, significantly reduced caspase-3 activity and the percentage of TUNEL-positive cells, substantiating a role for NF-kappaB in inducing intracellular dsRNA-mediated apoptosis. Concomitantly, RNA-dependent protein kinase activity was observed at 3 h post transfection along with phosphorylation and degradation of inhibitory kappaB-alpha. Expression of TRAIL (TNF-related apoptosis-inducing ligand), Fas, IL-15, and caspase-12 mRNAs was up-regulated in the presence of poly(I:C) but not when SN50 was also added. In contrast, there was no change detected in Fas, Fas-associated death domain, Bcl-2, Bcl-xl, Bax, p53, or XIAP(X-linked inhibitor of apoptosis protein) expression up to 12 h after poly(I:C) transfection. In addition, caspase-12 was cleaved, and phosphorylation of eukaryotic initiation factor 2alpha occurred, suggesting that an endoplasmic reticulum stress pathway was involved in addition to NF-kappaB induction of an extrinsic pathway, possibly mediated by TNF-related apoptosis-inducing ligand.
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PMID:Nuclear factor-kappaB translocation mediates double-stranded ribonucleic acid-induced NIT-1 beta-cell apoptosis and up-regulates caspase-12 and tumor necrosis factor receptor-associated ligand (TRAIL). 1296 48

The transcription factor NF-kappa B is elevated in murine T-cell lymphoma lines compared with normal thymic lymphocytes, and may play a role in the neoplastic transformation of these cells. When T lymphoma cells were treated with the soy isoflavone genistein, a marked reduction in nuclear NF-kappa B levels was detectable predominantly for the p50/p50 homodimer and p50/p65 heterodimer. To examine the mechanism by which NF-kappa B is reduced by genistein, we analyzed the NF-kappa B inhibitor, I kappa B alpha, and detected a 34 kDa cleavage product Delta I kappa B alpha, which was induced by genistein in a dose-dependent manner. Our observation that a pan-caspase inhibitor could inhibit the induction of Delta I kappa B alpha by genistein suggested that caspase activity was responsible for this cleavage product. In support of this idea, we detected an increase in caspase-3 activity in response to increasing time of genistein exposure. When the induction of Delta I kappa B alpha was prevented, we detected no reduction of NF-kappa B levels by genistein. These results support a direct role for Delta I kappa B alpha in the reduction of NF-kappa B by genistein. To determine the effect of genistein on some NF-kappa B target gene products, we examined the antiapoptotic proteins Bcl-2, Bcl-X(L), A1, and cIAP-1. Only changes in A1 and cIAP-1 levels were affected with significant reductions in response to genistein. Generation of the repressive activity of Delta I kappa B alpha on NF-kappa B is a novel mechanism for the reduction of this transcription factor by genistein and the possible effect this may have on the ability of genistein to induce apoptosis in tumor cells.
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PMID:Genistein reduces NF-kappa B in T lymphoma cells via a caspase-mediated cleavage of I kappa B alpha. 1296 87

Members of the NF-kappaB family of transcription factors cause transcriptional activation of anti-apoptotic genes. Here we determined whether survival of biotin-deficient cells is mediated by nuclear translocation of NF-kappaB. Human T (Jurkat) cells were cultured in biotin-deficient or biotin-supplemented media; nuclear translocation of NF-kappaB was stimulated with phytohemagglutinin and phorbol-12-myristate-13-acetate. Nuclear abundance of two members (p50 and p65) of the NF-kappaB family was greater in biotin-deficient compared to biotin-supplemented cells; this effect was mediated by phosphorylation of IkappaBalpha. The nuclear enrichment of p50 and p65 in biotin-deficient cells was associated with transcriptional activation of NF-kappaB-depedent genes such as the tumor suppressor gene p53 and the anti-apoptotic gene Bfl-1/A1. Biotin-deficient cells exhibited smaller activities of the apoptotic enzyme caspase-3 in response to treatment with tumor necrosis factor alpha, and decreased cell death in response to serum starvation compared to biotin-supplemented cells. These findings suggest that NF-kappaB mediates survival of biotin-deficient cells.
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PMID:Jurkat cells respond to biotin deficiency with increased nuclear translocation of NF-kappaB, mediating cell survival. 1529 80

Akt, also known as protein kinase B, is a serine/threonine protein kinase with antiapoptotic activities; also, it is a downstream target of phosphatidylinositol 3-kinase. Here we show that Akt1/Akt2 play a critical role in osteoclast differentiation but not cell survival and that mammalian target of rapamycin (mTOR) and Bim, a pro-apoptotic Bcl-2 family member, are required for cell survival in isolated osteoclast precursors. To investigate the function of Akt1, Akt2, mTOR, and Bim, we employed a retroviral system for delivery of small interfering RNA into cells. Loss of Akt1 and/or Akt2 protein inhibited osteoclast differentiation due to down-regulation of IkappaB-kinase (IKK) alpha/beta activity, phosphorylation of IkappaB-alpha, nuclear translocation of nuclear factor-kappaB (NFkappaB) p50, and NFkappaB p50 DNA-binding activity. Surprisingly, deletion of Akt1 and/or Akt2 protein did not stimulate cleaved caspase-3 activity and failed to promote apoptosis. Conversely, loss of mTOR protein induced apoptosis due to up-regulation of cleaved caspase-3 activity. In addition, we found that mTOR is downstream of phosphatidylinositol 3-kinase (but not Akt) and that macrophage colony-stimulating factor regulates Bim expression through mTOR activation for cell survival. These results demonstrate that Akt1/Akt2 are key elements in osteoclast differentiation and that the macrophage colony-stimulating factor stimulation of mTOR leading to Bim inhibition is essential for cell survival in isolated osteoclast precursors.
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PMID:Akt1/Akt2 and mammalian target of rapamycin/Bim play critical roles in osteoclast differentiation and survival, respectively, whereas Akt is dispensable for cell survival in isolated osteoclast precursors. 1554 69

Hypericin is the presumed active moiety within Saint John's wort. Extracts of Saint John's wort are widely used as an effective treatment for depression. Available as "over-the-counter" drugs, they are frequently part of the self-medication of patients undergoing radiation therapy for malignant diseases. In addition to antidepressive properties, hypericin has been shown to be able to induce apoptosis and radiosensitize tumor cells, and to have antiinflammatory and phototoxic skin effects. However, the underlying mechanisms are not clear. In this study, we investigated possible inhibitory effects of hypericin on proteasome function and related pathways. Extracts from U373 human glioma cells were incubated with different concentrations of hypericin. Three proteasome activities were monitored using a fluorogenic peptide assay. Activity of the transcription factor NF-kappaB and protein levels of p65, p50, IkappaBalpha and caspase-3 were investigated by EMSA and Western blotting, respectively. Hypericin caused a dose-dependent and photoactivation-independent inhibition of proteasome function. Hypericin treatment (6.25-50 microM) inhibited NF-kappaB, caused accumulation of phosphorylated IkappaBalpha, decreased p50 protein levels and induced cleavage of p65 protein in U373 cells. These effects were observed in MCF-7 cells only at higher concentrations of hypericin (12.5-50 microM). Additionally, inhibition of NF-kappaB activity in U373 cells by hypericin was prevented by caspase inhibition. Although hypericin clearly inhibits proteasome function, its effect NF-kappaB DNA-binding activity was not exclusively proteasome-dependent. The underlying mechanism might also involve caspase activation, a consequence of proteasome inhibition.
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PMID:Hypericin-an inhibitor of proteasome function. 1567 61


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