Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.22.62 (caspase-9)
 7,507 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Apoptosis is an evolutionarily conserved process that is critical for tissue homeostasis and development including sex determination in essentially all multicellular organisms. Here, we report the cloning of an ankyrin repeat-containing protein, termed F1Aalpha, in a yeast two-hybrid screen using the cytoplasmic domain of Fas (CD95/APO-1) as bait. Amino acid sequence analysis indicates that F1Aalpha has extensive homology to the sex-determining protein FEM-1 of the Caenorhabditis elegans, which is required for the development of all aspects of the male phenotype. F1Aalpha associates with the cytoplasmic domains of Fas and tumor necrosis factor receptor 1, two prototype members of the "death receptor" family. The F1Aalpha protein also oligomerizes. Overexpression of F1Aalpha induces apoptosis in mammalian cells, and co-expression of Bcl-XL or the dominant negative mutants of either FADD or caspase-9 blocks this effect. Deletion analysis revealed the center region of F1Aalpha, including a cluster of five ankyrin repeats to be necessary and sufficient for maximum apoptotic activity, and the N-terminal region appears to regulate negatively this activity. Furthermore, F1Aalpha is cleaved by a caspase-3-like protease at Asp(342), and the cleavage-resistant mutant is unable to induce apoptosis upon overexpression. F1Aalpha is therefore a member of a growing family of death receptor-associated proteins that mediates apoptosis.
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PMID:F1Aalpha, a death receptor-binding protein homologous to the Caenorhabditis elegans sex-determining protein, FEM-1, is a caspase substrate that mediates apoptosis. 1054 91

The caspase recruitment domain (CARD) of Apaf-1 binds to the CARD of caspase-9 to trigger a proteolytic cascade that leads to apoptotic cell death. We report the crystal structure of the Apaf-1 CARD at 1. 3 A resolution, solved in a two-element multiwavelength anomalous dispersion (MAD) X-ray diffraction experiment. This CARD adopts a six-helix bundle fold with Greek key topology surrounding an extensive hydrophobic core. This fold, which we call the "death fold", is found in other domains that mediate interactions in apoptotic signaling despite very low sequence identity. From a structure-based alignment, we identify conserved patterns that characterize the death fold and its subclasses. Like the Ig-fold, it provides a rigid structural scaffold upon which diverse recognition surfaces are assembled.
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PMID:Crystal structure of Apaf-1 caspase recruitment domain: an alpha-helical Greek key fold for apoptotic signaling. 1054 41

We have previously shown that the small heat shock protein HSP27 inhibited apoptotic pathways triggered by a variety of stimuli in mammalian cells. The present study demonstrates that HSP27 overexpression decreases U937 human leukemic cell sensitivity to etoposide-induced cytotoxicity by preventing apoptosis. As observed for Bcl-2, HSP27 overexpression delays poly(ADP-ribose)polymerase cleavage and procaspase-3 activation. In contrast with Bcl-2, HSP27 overexpression does not prevent etoposide-induced cytochrome c release from the mitochondria. In a cell-free system, addition of cytochrome c and dATP to cytosolic extracts from untreated cells induces the proteolytic activation of procaspase-3 in both control and bcl-2-transfected U937 cells but fails to activate procaspase-3 in HSP27-overexpressing cells. Immunodepletion of HSP27 from cytosolic extracts increases cytochrome c/dATP-mediated activation of procaspase-3. Overexpression of HSP27 also prevents procaspase-9 activation. In the cell-free system, immunodepletion of HSP27 increases LEDH-AFC peptide cleavage activity triggered by cytochrome c/dATP treatment. We conclude that HSP27 inhibits etoposide-induced apoptosis by preventing cytochrome c and dATP-triggered activity of caspase-9, downstream of cytochrome c release.
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PMID:HSP27 inhibits cytochrome c-dependent activation of procaspase-9. 1054 89

ELL-12, a liposome formulation of the ether-lipid 1-O-octadecyl-2-O-methyl-sn-glycero-3-phosphocholine (ET-18-OCH(3)), is a nonmyelosuppressive antiproliferative agent that is more effective and less toxic than the ether lipid itself in tumor model systems. We found that ELL-12 induced apoptosis in Jurkat, H9, and U-937 cells that was preceded by activation of executioner caspases. In addition, ELL-12 triggered release of cytochrome c from mitochondria to the cytoplasm before caspase-9 activation. Apoptosis, activation of caspases, and cytochrome c release were blocked by Bcl-x(L) overexpression in Jurkat T cells, suggesting a critical role for mitochondria in ELL-12-triggered cell death. Furthermore, ELL-12 had no effect on expression of CD95 ligand, and inhibition of the Fas signaling pathway with antagonistic anti-CD95 antibody did not affect apoptosis induced by ELL-12. Hence, ELL-12 could be a promising adjunct for the treatment of tumors in addition to myelosuppressive chemotherapeutic drugs and/or those that use the CD95-ligand/receptor system to trigger apoptosis.
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PMID:Liposomal ET-18-OCH(3) induces cytochrome c-mediated apoptosis independently of CD95 (APO-1/Fas) signaling. 1055 70

In monolayer cultures of P19 EC cells treated with both all-trans retinoic acid (RA) and bone morphogenetic protein (BMP)-4 (RA/BMP-4 treatment), many non-adherent apoptotic cells and activated caspase-3-positive cells were observed, but they were not observed in cells treated with RA or BMP-4 alone. Consistent with the appearance of activated caspase-3-positive cells, BMP-4 and RA together induced processing of caspase-9, Ac-DEVD-MCA cleavage activity and DNA fragmentation. These three activities were observed infrequently or not at all when cells were treated with RA or BMP-4 alone. In the RA/BMP-4 treatment-induced apoptosis, caspase-9 was upstream of caspase-3 in the enzyme cascade, and the caspase-9 to -3 step was key in the apoptotic pathway. Bcl-xL inhibited processing of caspase-9, Ac-DEVD-MCA cleavage activity and DNA fragmentation induced by RA/BMP-4 treatment. However, unlike staurosporine-induced apoptosis, cytochrome c, which activates caspase-9, was not detected in the cytosol of RA/BMP-4-treated cells. RA and BMP-4 may activate caspase-9 through an apoptotic pathway other than the Apaf-1/cytochrome c pathway. The prominent decrease of X-chromosome-linked inhibitory apoptosis protein (XIAP) in the cytosol may explain the activation of caspase-9 induced by RA and BMP-4 treatment.
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PMID:BMP-4 and retinoic acid synergistically induce activation of caspase-9 and cause apoptosis of P19 embryonal carcinoma cells cultured as a monolayer. 1057 80

alpha-Fetoprotein (AFP) is an oncoembryonal protein with multiple cell growth regulating, differentiating and immunosuppressive activities. Previous studies have shown that treatment of tumor cells in vitro with 1-10 microM AFP produces significant suppression of tumor cell growth by inducing dose-dependent cytotoxicity, but the molecular mechanisms underlying these AFP functions are obscure. Here, we show that AFP cytotoxicity is closely related to apoptosis, as shown by cell morphology, nuclear DNA fragmentation and caspase-3-like activity resulting in cleavage of poly(ADP-ribose) polymerase. Apoptosis was significantly inhibited by a CPP32 family protease inhibitor whereas a general caspase inhibitor had no inhibitory effect, showing some enhancement of AFP-mediated cell death. Using fluorogenic caspase substrates, we found that caspase-3-like proteases were activated as early as 4 h after treatment of Raji cells with 15 microM AFP, whereas caspase-1, caspase-8, and caspase-9-like activity was not detected during the time interval 0.5-17 h. AFP treatment of Raji cells increased Bcl-2 protein, showing that AFP-induced apoptosis is not explained by downregulation of the Bcl-2 gene. This also suggests that AFP operates downstream of the Bcl-2-sensitive step. AFP notably decreased basal levels of soluble and membrane-bound Fas ligand. Incubation of AFP-sensitive tumor cells (HepG2, Raji) with neutralizing anti-Fas, anti-tumor necrosis factor receptor (TNFR)1 or anti-TNFR2 mAb did not prevent AFP-induced apoptosis, demonstrating its independence of Fas-dependent and TNFR-dependent signaling. In addition, it was found that cells resistant to TNF-induced (Raji) or Fas-induced (MCF-7) apoptosis are, nevertheless, sensitive to AFP-mediated cell death. In contrast, cells sensitive to Fas-mediated cell death (Jurkat) are completely resistant to AFP. Taken as a whole, our data demonstrate that: (a) AFP induces apoptosis in tumor cells independently of Fas/Fas ligand or TNFR/TNF signaling pathways, and (b) AFP-mediated cell death involves activation of the effector caspase-3-like proteases, but is independent of upstream activation of the initiator caspase-1, caspase-8, and caspase-9-like proteases.
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PMID:alpha-fetoprotein causes apoptosis in tumor cells via a pathway independent of CD95, TNFR1 and TNFR2 through activation of caspase-3-like proteases. 1058 68

Ceramide is characterized as a second messenger of apoptosis induced by various agents such as tumor necrosis factor (TNF-alpha), Fas ligand, hydrogen peroxide, heat shock and ionizing radiation. In this study, we investigated the mechanism of ceramide-induced apoptosis using a human neuroblastoma cell line, SK-N-MC. N-Acetyl-sphingosine (C2-ceramide), a cell-permeable ceramide analogue, was able to induce apoptosis in SK-N-MC cells as estimated by DNA fragmentation and chromatin condensation. C2-ceramide-induced DNA fragmentation was blocked by caspase inhibitor (Z-Asp-CH(2)-DCB). An increase in caspase-3 (CPP32)-like protease activity was evident during C2-ceramide-induced apoptosis, suggesting that caspases are involved in this apoptosis. Moreover, enzymatic cleavage of VDVAD-AFC and LEHD-AFC (specific substrates for caspase-2 and -9, respectively) was increased by treatment with C2-ceramide. To elucidate which types of caspase are activated in C2-ceramide-treated cells, we performed Western blot analysis using antibodies against each isoform. Both proforms of caspase-2 and -3 were decreased in response to C2-ceramide in a time-dependent manner. Mitochondrial cytochrome c is also time-dependently released into the cytosol in response to treatment with C2-ceramide. Addition of cytochrome c into the S-100 fractions prepared from SK-N-MC cells could activate caspase-2 in cell-free systems. These results suggest the possibility that cytochrome c released to the cytosol can activate caspases (caspase-9, -3, and -2) during C2-ceramide-induced apoptosis of SK-N-MC cells.
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PMID:Possible involvement of cytochrome c release and sequential activation of caspases in ceramide-induced apoptosis in SK-N-MC cells. 1059 Mar 15

Apoptosis induced by T cell receptor (TCR) triggering in T lymphocytes involves activation of cysteine proteases of the caspase family through their proteolytic processing. Caspase-3 cleavage was also reported during T cell stimulation in the absence of apoptosis, although the physiological relevance of this response remains unclear. We show here that the caspase inhibitor benzyloxycarbonyl (Cbz)-Val-Ala-Asp(OMe)-fluoromethylketone (zVAD) blocks proliferation, major histocompatibility complex class II expression, and blastic transformation during stimulation of peripheral blood lymphocytes. Moreover, T cell activation triggers the selective processing and activation of downstream caspases (caspase-3, -6, and -7), but not caspase-1, -2, or -4, as demonstrated even in intact cells using a cell-permeable fluorescent substrate. Caspase-3 processing occurs in different T cell subsets (CD4(+), CD8(+), CD45RA(+), and CD45RO(+)), and in activated B lymphocytes. The pathway leading to caspase activation involves death receptors and caspase-8, which is also processed after TCR triggering, but not caspase-9, which remains as a proenzyme. Most importantly, caspase activity results in a selective substrate specificity, since poly(ADP-ribose) polymerase (PARP), lamin B, and Wee1 kinase, but not DNA fragmentation factor (DFF45) or replication factor C (RFC140), are processed. Caspase and substrate processing occur in nonapoptotic lymphocytes. Thus, caspase activation is an early and physiological response in viable, stimulated lymphocytes, and appears to be involved in early steps of lymphocyte activation.
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PMID:Early activation of caspases during T lymphocyte stimulation results in selective substrate cleavage in nonapoptotic cells. 1060 47

Antigen-induced apoptosis of B cells serves to deplete the immune repertoire of anti-self specificities leading to central and peripheral B cell tolerance. However, the mechanism of B cell receptor (BCR)-mediated apoptosis is widely unknown. By using the human Burkitt lymphoma cell line BL60 as a model system for human germinal center B cells we show here that BCR-mediated apoptosis requires transcriptional activity but, in contrast to activation-induced T cell apoptosis, is neither mediated via known death receptor systems nor does it involve initial activation of caspase-8. Moreover, during BCR-induced apoptosis cytochrome c release and mitochondrial permeability transition (PT) precedecaspase activation. Although caspase inhibition after BCR stimulation blocks cleavage of caspase substrates and DNA fragmentation it does not prevent mitochondrial PT, cytochrome c release and cell death. Thus, BCR-mediated apoptosis is initiated by the caspase-independent induction of mitochondrial PT resulting in release of cytochrome c and subsequent activation of caspase-9, downstream caspases and apoptosis.
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PMID:Critical role for mitochondria in B cell receptor-mediated apoptosis. 1060 28

In the present study, we examined the direct cytotoxic effects of cocaine on fetal cardiac myocytes. Cocaine treatment of cultured fetal rat (21 days) myocardial cells (FRMCs) induced a time- and concentration-dependent increase in apoptotic cells in FRMCs. Cocaine induced surface exposure of phosphatidylserine in FRMCs at 12-h treatment and increased apoptotic cells up to 96 h. Corresponding DNA fragmentation induced by cocaine in these cells was demonstrated in situ by terminal deoxynucleotidyl transferase biotin-dUTP nick end labeling assay and by electrophoresis of labeled DNA fragments, showing the characteristic apoptotic ladders. The pD(2) and maximum increase of cocaine-induced apoptosis in FRMCs were 4.3 and 3.2-fold, respectively. Both caspase-9 and caspase-3 inhibitors (Z-LEHD-FMK and Ac-DEVD-CHO, respectively) blocked cocaine-induced apoptosis. In addition, cyclosporin A inhibited cocaine-induced apoptosis in a concentration-dependent manner with an IC(50) value of 0.1 microM. The maximum of 86% inhibition was obtained with 3 microM cyclosporin A. Cocaine induced the release of cytochrome c from the mitochondria and increased its levels in the cytosol by 3.1-fold. In accordance, the level of cytochrome c in the mitochondria fraction decreased by approximately 60%. Cocaine-induced translocation of cytochrome c was inhibited by cyclosporin A. The results indicate that cocaine has a direct cytotoxic effect on fetal cardiomyocytes by inducing apoptosis in the cells. Furthermore, the release of cytochrome c from the mitochondria and its subsequent activation of caspase-9 and caspase-3 play a key role in cocaine-induced apoptosis.
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PMID:Cocaine induces apoptosis in fetal myocardial cells through a mitochondria-dependent pathway. 1060 26


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