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

The observation that the nematode cell death effector gene product Ced-3 is homologous to human interleukin-1beta-converting enzyme (caspase-1) has led to the discovery of at least nine other human caspases, many of which are implicated as mediators of apoptosis. Significant interest has been given to aspects of the cell biology and substrate specificity of this family of proteases; however, quantitative descriptions of their biochemical characteristics have lagged behind. We describe the influence of a number of environmental parameters, including pH, ionic strength, detergent, and specific ion concentrations, on the activity and stability of four caspases involved in death receptor-mediated apoptosis. Based on these observations, we recommend the following buffer as optimal for investigation of their characteristics in vitro: 20 mM piperazine-N,N'-bis(2-ethanesulfonic acid) (PIPES), 100 mM NaCl, 10 mM dithiothreitol, 1 mM EDTA, 0.1% 3-[(3-cholamidopropyl)dimethylammonio]-2-hydroxy-1-propanesulfonic acid (CHAPS), 10% sucrose, pH 7.2. Caspase activity is not affected by concentrations of Ca2+ below 100 mM, but is abolished by Zn2+ in the submicromolar range, a common characteristic of cysteine proteases. Optimal pH values vary from 6.8 for caspase-8 to 7.4 for caspase-3, and activity of all is relatively stable between 0 and 150 mM NaCl. Consequently, changes in the physiologic pH and ionic strength would not significantly alter the activity of the enzymes, inasmuch as all four caspases are optimally active within the range of these parameters found in the cytosol of living and dying human cells.
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PMID:Biochemical characteristics of caspases-3, -6, -7, and -8. 932 97

Rat pheochromocytoma (PC12) cells and sympathetic neurons undergo apoptotic cell death upon withdrawal of trophic support. We have shown previously that selective cysteine aspartase (caspase) inhibitors protect PC12 cells and sympathetic neurons from such death, and that the caspase Nedd-2 is required for this type of death to occur. We now show that 4-(2-aminoethyl)benzenesulfonyl fluoride hydrochloride (AEBSF) and N(alpha)-p-tosyl-L-lysine chloromethyl ketone (TLCK), agents that inhibit another class of proteases, the trypsin-like serine proteases, also suppress cell death in this paradigm. The site of action of these agents is upstream of the caspases, because the CPP32-like and Nedd-2-cleaving activities that are induced upon withdrawal of trophic support in PC12 cells are inhibited when AEBSF and TLCK are applied to the cells. Both agents inhibit thymidine incorporation in PC12 cells at concentrations similar to those that promote survival, raising the possibility that they may promote survival in neuronal cells through inhibition of aberrant activation of cell cycle components.
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PMID:Inhibitors of trypsin-like serine proteases inhibit processing of the caspase Nedd-2 and protect PC12 cells and sympathetic neurons from death evoked by withdrawal of trophic support. 932 71

Apoptosis is a major form of cell death, characterized initially by a series of stereotypic morphological changes. In the nematode Caenorhabditis elegans, the gene ced-3 encodes a protein required for developmental cell death. Since the recognition that CED-3 has sequence identity with the mammalian cysteine protease interleukin-1 beta-converting enzyme (ICE), a family of at least 10 related cysteine proteases has been identified. These proteins are characterized by almost absolute specificity for aspartic acid in the P1 position. All the caspases (ICE-like proteases) contain a conserved QACXG (where X is R, Q or G) pentapeptide active-site motif. Capases are synthesized as inactive proenzymes comprising an N-terminal peptide (prodomain) together with one large and one small subunit. The crystal structures of both caspase-1 and caspase-3 show that the active enzyme is a heterotetramer, containing two small and two large subunits. Activation of caspases during apoptosis results in the cleavage of critical cellular substrates, including poly(ADP-ribose) polymerase and lamins, so precipitating the dramatic morphological changes of apoptosis. Apoptosis induced by CD95 (Fas/APO-1) and tumour necrosis factor activates caspase-8 (MACH/FLICE/Mch5), which contains an N-terminus with FADD (Fas-associating protein with death domain)-like death effector domains, so providing a direct link between cell death receptors and the caspases. The importance of caspase prodomains in the regulation of apoptosis is further highlighted by the recognition of adapter molecules, such as RAIDD [receptor-interacting protein (RIP)-associated ICH-1/CED-3-homologous protein with a death domain]/CRADD (caspase and RIP adapter with death domain), which binds to the prodomain of caspase-2 and recruits it to the signalling complex. Cells undergoing apoptosis following triggering of death receptors execute the death programme by activating a hierarchy of caspases, with caspase-8 and possibly caspase-10 being at or near the apex of this apoptotic cascade.
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PMID:Caspases: the executioners of apoptosis. 933 44

Cysteine proteases of the CED-3 and ICE family have been recently proposed as the ultimate executioners in several mammalian cell death pathways. Among them, the cysteine protease CPP32 has been shown to participate in programmed cell death (PCD), or apoptosis, affecting lymphoid cells in vitro. In the thymus, negative selection is a mechanism through which developing thymocytes expressing a TcR with high affinity for self peptide-MHC complexes are eliminated by PCD. In order to investigate the role of CPP32 in thymic apoptosis, isolated thymocytes were submitted to cell surface CD3 crosslinking by immobilized anti-CD3 mAb or to dexamethasone treatment. Although apoptosis occurred in the absence or after crosslinking with anti-CD3 mAb, specific activation of CPP32, as assessed by the extent of proteolytic cleavage of the p32 zymogen, was only detected in thymocytes cultured in the presence of the immobilized antibody or dexamethasone. This activation was a very early event during apoptosis as it occurred before the exposure of phosphatidyl serine to the upper side of the cell membrane. This was observed both in anti-CD3- and dexamethasone-induced apoptosis. Moreover, using mice transgenic for pigeon cytochrome C (PCC)-specific TcR, we were able to show that, after injection of PCC, the activation of CPP32 and cleavage of its substrate occurred in thymocytes obtained from mice expressing a permissive MHC haplotype for PCC presentation (H-2k). Moreover, PCC induced apoptosis was blocked by the caspase inhibitor zVAD. While spontaneous apoptosis was not accompanied by detectable levels of CPP32 processing, it was characterized by the proteolysis of poly(ADP-ribose) polymerase (PARP) and was blocked by the cysteine protease inhibitor, zVAD-CH2F. Taken together, these results support the concept that CPP32 is among the earliest effectors of the pathway leading to negative selection of autoreactive thymocytes. Our results also suggest the involvement of a distinct CPP32-like cysteine protease in spontaneous apoptosis of thymocytes.
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PMID:Specific activation of the cysteine protease CPP32 during the negative selection of T cells in the thymus. 934 8

IL-1beta converting enzyme (ICE) family cysteine proteases are subdivided into three groups; ICE-, CPP32-, and Ich-1-like proteases. In Fas-induced apoptosis, activation of ICE-like proteases is followed by activation of CPP32-like proteases which is thought to be essential for execution of the cell death. It was recently reported that two subfamily members of the mitogen-activated protein kinase superfamily, JNK/SAPK and p38, are activated during Fas-induced apoptosis. Here, we have shown that MKK7, but not SEK1/ MKK4, is activated by Fas as an activator for JNK/ SAPK and that MKK6 is a major activator for p38 in Fas signaling. Then, to dissect various cellular responses induced by Fas, we used several peptide inhibitors for ICE family proteases in Fas-treated Jurkat cells and KB cells. While Z-VAD-FK which inhibited almost all the Fas-induced cellular responses blocked the activation of JNK/SAPK and p38, Ac-DEVD-CHO and Z-DEVD-FK, specific inhibitors for CPP32-like proteases, which inhibited the Fas-induced chromatin condensation and DNA fragmentation did not block the activation of JNK/SAPK and p38. Interestingly, these DEVD-type inhibitors did not block the Fas-induced morphological changes (cell shrinkage and surface blebbing), induction of Apo2.7 antigen, or the cell death (as assessed by the dye exclusion ability). These results suggest that the Fas-induced activation of the JNK/SAPK and p38 signaling pathways does not require CPP32-like proteases and that CPP32-like proteases, although essential for apoptotic nuclear events (such as chromatin condensation and DNA fragmentation), are not required for other apoptotic events in the cytoplasm or the cell death itself. Thus, the Fas signaling pathway diverges into multiple, separate processes, each of which may be responsible for part of the apoptotic cellular responses.
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PMID:Fas induces cytoplasmic apoptotic responses and activation of the MKK7-JNK/SAPK and MKK6-p38 pathways independent of CPP32-like proteases. 936 18

Neuroblastomas frequently show spontaneous regression and differentiation, which may at least partly be regulated by signaling through nerve growth factor and its receptors, TRK-A and p75LNTR. We studied 52 neuroblastic tumors to test whether the cell death-related proteases, interleukin-1 beta converting enzyme (ICE), CPP32, and Ich-1, were involved in the regression of the tumors. High levels of expression of ICE and CPP32 were significantly correlated with a high level of TRK-A expression, single copy of N-myc, younger age, lower stages, and better prognosis. The immunohistochemical studies and Western analyses as well as the terminal dUTP-biotin nick end labeling (TUNEL) method revealed that both ICE and CPP32 were translocated from the cytoplasm into the nuclei in regressing, apoptotic tumor cells. Our results suggest that ICE and CPP32 cysteine proteases may play an important role in regulating the apoptotic process of the favorable neuroblastomas.
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PMID:High levels of expression and nuclear localization of interleukin-1 beta converting enzyme (ICE) and CPP32 in favorable human neuroblastomas. 937 72

Caspases are a family of cysteine proteases implicated in the biochemical and morphological changes that occur during apoptosis (programmed cell death). The loop domain of Bcl-2 is cleaved at Asp34 by caspase-3 (CPP32) in vitro, in cells overexpressing caspase-3, and after induction of apoptosis by Fas ligation and interleukin-3 withdrawal. The carboxyl-terminal Bcl-2 cleavage product triggered cell death and accelerated Sindbis virus-induced apoptosis, which was dependent on the BH3 homology and transmembrane domains of Bcl-2. Inhibitor studies indicated that cleavage of Bcl-2 may further activate downstream caspases and contribute to amplification of the caspase cascade. Cleavage-resistant mutants of Bcl-2 had increased protection from interleukin-3 withdrawal and Sindbis virus-induced apoptosis. Thus, cleavage of Bcl-2 by caspases may ensure the inevitability of cell death.
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PMID:Conversion of Bcl-2 to a Bax-like death effector by caspases. 939 3

There is increasing evidence that programmed cell death (PCD) depends on a novel family of intracellular cysteine proteases, called caspases, that includes the Ced-3 protease in the nematode Caenorhabditis elegans and the interleukin-1beta-converting enzyme (ICE)-like proteases in mammals. Some developing cells, including lens epithelial cells, erythroblasts, and keratinocytes, lose their nucleus and other organelles when they terminally differentiate, but it is not known whether the enzymatic machinery of PCD is involved in any of these normal differentiation events. We show here that at least one CPP32 (caspase-3)-like member of the caspase family becomes activated when rodent lens epithelial cells terminally differentiate into anucleate lens fibers in vivo, and that a peptide inhibitor of these proteases blocks the denucleation process in an in vitro model of lens fiber differentiation. These findings suggest that at least part of the machinery of PCD is involved in lens fiber differentiation.
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PMID:A role for caspases in lens fiber differentiation. 942 63

Nitric oxide has been shown to inhibit apoptosis of human umbilical venous endothelial cells (HUVEC). Therefore we investigated the effect of different NO donors, PAPA NONOate (NOC-15; NO.) and nitrosodium tetrafluoroborate (NOBF4, NO+), and the reaction product of NO and O2-, peroxynitrite (ONOO- ), on TNF-alpha- or serum depletion-induced apoptosis of HUVEC. TNF-alpha-induced DNA fragmentation, determined by ELISA, was inhibited by NOC-15, NOBF4, and ONOO- in a concentration-dependent manner (maximal effects with 10 microM NO. and ONOO- and 100 microM NO+). The inhibition of apoptosis correlated with a protective effect on cell viability. The caspases, a cysteine protease family, play an important role in apoptotic processes. To determine whether the different NO donors and ONOO- regulate this enzyme, caspase-3-like activity was measured in homogenates of TNF-alpha-treated HUVEC. The TNF-alpha-induced enzyme activity was abrogated by NO., NO+, and ONOO-. Furthermore, caspase-3 activity was determined in vitro by reconstitution of the separately cloned, bacterially expressed, and purified active p17 and p12 subunits. The reconstituted caspase-3 exhibited enzyme activity, which was suppressed by the different NO donors and ONOO- with an IC50 of 50 microM for NOC-15, 1 mM for NOBF4, and 50 microM for ONOO-. The inhibition of caspase-3 activity correlated with a S-nitrosylation of the reactive cysteine residue and was reversed by further addition of dithiothreitol. This study suggests that the cellular regulatory processes of NO to protect cells from apoptosis may be independent of the redox state and that low concentrations of NO and ONOO- inhibit the cellular suicide program in HUVEC via S-nitrosylation of members of the caspase family.
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PMID:Effects of redox-related congeners of NO on apoptosis and caspase-3 activity. 944

Dolichyl phosphate, an essential carrier lipid in the biosynthesis of N-linked glycoprotein, has been found to induce apoptosis in rat glioma C6 cells and human monoblastic leukemia U937 cells. In the present study, dolichyl phosphate and structurally related compounds were examined regarding their apoptosis-inducing activities in U937 cells. Dihydroheptaprenyl and dihydrodecaprenyl phosphates, of which isoprene units are shorter than that of dolichyl phosphate, induced apoptosis in U937 cells. This phenomenon occurred in a dose- and time-dependent manner, as seen with dolichyl phosphate-induced apoptosis. Derivatives of the same isoprene units of dolichyl phosphate, such as dolichol, dolichal or dolichoic acid, did not induce DNA fragmentation. Farnesyl phosphate and geranylgeranyl phosphate also failed to induce apoptosis. During apoptosis, the caspase family of cysteine proteases play important roles. We observed that apoptosis induced by dihydroprenyl phosphate was mediated by caspase-3-like (CPP32-like) activation but not by caspase-1-like (ICE-like) activation. This caspase-3-like activation was inhibited by a specific inhibitor of caspase-3, DEVD-CHO, but not by an caspase-1 inhibitor YVAD-CHO. We interpret these results to mean that dihydroprenyl phosphates with more than seven isoprene units have apoptosis-inducing activity and that their signal is mediated by caspase-3-like activation.
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PMID:Dihydroheptaprenyl and dihydrodecaprenyl monophosphates induce apoptosis mediated by activation of caspase-3-like protease. 946 Dec 54


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