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)

HL-60 cells differentiating into neutrophil-like cells die an apoptotic death in vitro. Susceptibility to apoptosis is associated with decreased Bcl-2 protein and mRNA expression; however, the effect of differentiation on the expression of pro-apoptotic caspases is unknown. Spontaneous apoptosis occurred 6 days after retinoic acid treatment. Western blotting showed loss of Bcl-2 by day 7, and new expression of ICE (caspase 1) and CPP32 (caspase 3) protein by day 2. Northern analysis demonstrated loss of Bcl-2 mRNA and increases in ICE mRNA by day 2; CPP32 mRNA was unchanged. Differential Bcl-2 and ICE mRNA expression was also found when granulocytic differentiation was stimulated by DMSO. Differentiated HL-60 cell lysates exhibited functional ICE proteolytic activity. De novo caspase expression was responsible for the development of spontaneous apoptosis, since specific inhibitors of ICE (YVAD-CMK) and CPP32 (DEVD-CHO), inhibited retinoic acid induced spontaneous apoptosis. Functional maturation and susceptibility to apoptosis are both inducible and linked in this granulocyte precursor cell line.
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PMID:Granulocytic differentiation of HL-60 cells results in spontaneous apoptosis mediated by increased caspase expression. 927 75

Six hours after ultraviolet B (UVB) irradiation (11.6 mJ/cm2), the viability of A431 cells decreased, and, at the same time, fragmentation of genomic DNA into nucleosomal units was observed. Z-Asp-CH2-DCB (100 microM), an inhibitor of interleukin-1 beta-converting enzyme (caspase-1) and caspase-1-like proteases, markedly inhibited UVB-induced cell death and DNA fragmentation. Both YVAD-CMK, an inhibitor of caspase-1, and DEVD-CHO, an inhibitor of caspase-3, moderately inhibited the UVB-induced cell death. A combination of YVAD-CMK and DEVD-CHO acted additionally in inhibiting cell death. These observations suggest strongly the cooperative involvement of caspases in the apoptosis induced in A431 cells by UVB.
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PMID:Involvement of caspases in apoptosis induced by ultraviolet B irradiation in A431 human epithelioid tumor cells. 930 45

Intracellular cysteine proteinases (caspases) play key roles in inflammation and apoptosis. Recombinant caspases are typically produced in Escherichia coli expression systems with the attendant problems of solubilization, re-folding and activation of the protease. Here we describe the expression of hexahistidine-tagged caspase-3 (CPP32/Yama/Apopain) in the methylotropic yeast Pichia pastoris, and the purification of soluble enzyme from yeast lysates using cobalt affinity chromatography. The recombinant protease is fully activated, stable, and cleaves the synthetic substrate DEVD-AFC (Km 16.8 microM) but not YVAD-AFC. It mediates the cleavage of the apoptotic death substrate poly(ADP-ribose) polymerase in cell extracts, but does not cleave pro-interleukin-1beta. It is inhibited by the peptide DEVD-CHO (Ki 2.2 nM), far less efficiently by YVAD-CMK (Ki 0.3 microM), and not detectably by CrmA. By these criteria, recombinant caspase-3 is indistinguishable from native caspase-3 purified from apoptotic cell extracts. Activation of recombinant caspase-3 occurs in yeast in the absence of any intrinsic caspase activity, suggesting that caspase-3 can auto-activate. However, the purified enzyme was incapable of cleaving pro-caspase-3 indicating that autoactivation of caspase-3 in vivo is not likely to occur unless very high concentrations are achieved.
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PMID:Recombinant caspase-3 expressed in Pichia pastoris is fully activated and kinetically indistinguishable from the native enzyme. 932 93

Granzyme B (GzmB) is a neutral serine protease found in cytotoxic lymphocytes; this enzyme is critically involved in delivering the rapid apoptotic signal to susceptible target cells. GzmB has been difficult to study and has not yet been produced in non-mammalian systems because of the complex processing events that are thought to be required for its activation. In this report, we have successfully produced fully active, soluble recombinant GzmB (rGzmB) in a yeast-based system by fusing GzmB cDNA in frame with yeast alpha-factor cDNA, using the yeast KEX2 signal peptidase to release the processed enzyme into the supernatant of yeast cultures. We expressed the proenzyme form of GzmB as well and determined that pro-GzmB is efficiently converted to its active form by the cysteine proteinase dipeptidyl peptidase I. The fully processed enzyme was able to hydrolyze the synthetic substrate N-t-butyloxycarbonyl-L-alanyl-L-alanyl-L-aspartyl (Boc-Ala-Ala-Asp) thiobenzyl ester with a kcat of 17 s-1 and catalytic efficiency kcat/Km of 181,237 M-1 S-1; the recombinant enzyme is therefore at least twice as active as purified native GzmB. In addition, the recombinant enzyme hydrolyzes Boc-Ala-Ala-Met thiobenzyl ester with a kcat of 3.2 S-1 and a catalytic efficiency kcat/Km of 65,306 M-1 S-1. Purified rGzmB can also cleave the putative substrate caspase-3 into its signature p20/p10 forms. Unlike caspases, rGzmB is not sensitive to inhibition by several peptide-based inhibitors, including Ac-DEVD-CHO, Ac-YVAD-CMK, and ZIETD-FMK, as well as Zn2+ (a known inhibitor of caspase-3). Structural studies of rGzmB may allow us to better understand the substrate specificity of this enzyme and to design better inhibitors.
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PMID:Production of fully active recombinant murine granzyme B in yeast. 943 Jul 5

We have demonstrated that ginsenoside Rh2 (G-Rh2), a ginseng saponin with a dammarane skeleton, induces apoptosis of human hepatoma SK-HEP-1 cells as evidenced by analyses of DNA fragmentation, flow cytometry and changes in cell morphology. Ac-YVAD-CMK or Ac-DEVD-CHO effectively prevented G-Rh2-induced DNA fragmentation, indicating the involvement of caspase-like proteases in the process of apoptosis. In addition, G-Rh2 induced the processing of caspase-3 to an active form, p17. In stable Bcl-2 transfectants, G-Rh2 also induced DNA fragmentation, while staurosporine-induced DNA fragmentation was totally blocked. As it did in wild-type cells, G-Rh2 induced the proteolytic activation of caspase-3 protease and subsequent cleavage of PARP in the bcl-2 transfectants. In summary, G-Rh2 contains an apoptotic inducing activity in SK-HEP-1 cells which functions via Bcl-2-insensitive activation of caspase-3, followed by proteolytic cleavage of PARP.
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PMID:Activation of caspase-3 protease via a Bcl-2-insensitive pathway during the process of ginsenoside Rh2-induced apoptosis. 945 77

Ceramide, a product of sphingomyelin metabolism, is a novel lipid second messenger that mediates diverse cellular functions. The present study demonstrates the activation of caspase-3/CPP-32beta, during apoptosis induced by cell permeable exogenous ceramides, in AK-5 tumor, a spontaneously regressing rat histiocytoma. The apoptotic events were suppressed by the caspase-3 specific tetrapeptide inhibitor DEVD-CHO but not by the caspase-1 inhibitor YVAD-CMK. In cells overexpressing Bcl-2, a significant decrease in cell death was observed after exogenous addition of ceramides. Furthermore the processing of caspase-3 to its active form upon apoptotic stimulus, and the subsequent cleavage of the substrate PARP, suggested a central role for caspase-3 in the ceramide mediated apoptosis in AK-5 tumor cells.
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PMID:Selective involvement of caspase-3 in ceramide induced apoptosis in AK-5 tumor cells. 984 82

Nonsteroidal antiinflammatory agents (NSAIA) have been shown to exert potent chemopreventive activity against colon, lung, and breast cancers. In this study, we show that at pharmacological concentrations (1 to 3 mmol/L) sodium salicylate (Na-Sal) can potently induce programmed cell death in several human myeloid leukemia cell lines, including TF-1, U937, CMK-1, HL-60, and Mo7e. TF-1 cells undergo rapid apoptosis on treatment with Na-Sal, as indicated by increased annexin V binding capacity, cpp-32 (caspase-3) activation, and cleavage of poly (ADP-ribose) polymerase (PARP) and gelsolin. In addition, the expression of MCL-1, an antiapoptotic member of the BCL-2 family, is downregulated during Na-Sal-induced cell death, whereas the expression of BCL-2, BAX, and BCL-XL is unchanged. Z-VAD, a potent caspase inhibitor, prevents the cleavage of PARP and gelsolin and rescues cells from Na-Sal-induced apoptosis. In addition, we show that Na-Sal accelerates growth factor withdrawal-induced apoptosis and synergizes with daunorubicin to induce apoptosis in TF-1 cells. Thus, our data provide a potential mechanism for the chemopreventive activity of NSAIA and suggest that salicylates may have therapeutic potential for the treatment of human leukemia.
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PMID:Sodium salicylate activates caspases and induces apoptosis of myeloid leukemia cell lines. 1009 Sep 50

Experimental models of sepsis using endotoxin challenges, including studies with sensitized animals with D-galactosamine, have largely contributed to the basic rationale for innovative clinical trials in human septic shock, which have, to date, failed. The ability of these models to reproduce human disease has been highly discussed. We report here that the widely used D-galactosamine/LPS model does not account for septic shock. Treatment with YVAD-CMK, a potent tetrapeptide inhibitor of caspases of the interleukin (IL)-1beta converting enzyme (ICE) family, protects from LPS-induced liver apoptosis and mortality in D-galactosamine-sensitized mice when administered either before or up to 2 h after the lethal challenge. This curative effect is related to complete inhibition of caspase-3 activity in the liver. However, YVAD-CMK does not affect LPS-induced release of IL-1beta and does not protect from a lethal dose of LPS in unsensitized mice. These experiments demonstrate the difference between these two widely recognized experimental models of sepsis. LPS toxicity in D-galactosamine-treated mice, leading to blocked gene transcription, results from tumor necrosis factor (TNF)-alpha-induced caspase-3-dependent liver injury, not from the systemic inflammatory response. These results provide evidence that inhibitors of the ICE caspase family can prevent or even overcome the ongoing hepatic injury induced by TNF-alpha during sepsis, ischemia-reperfusion, or severe hepatitis.
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PMID:LPS challenge in D-galactosamine-sensitized mice accounts for caspase-dependent fulminant hepatitis, not for septic shock. 1019 82

We recently reported that HL-60 cells underwent apoptosis when exposed to room temperature (RT) (21 degrees C). RT-induced apoptosis of HL-60 cells is inhibited by the caspase-1 inhibitor (YVAD-CMK), but not by the caspase-3 inhibitor (DEVD-CHO). In this study, we studied RT-induced apoptosis in 15 human cell lines of hematopoietic lineage and found that the Jurkat cell line also responded to RT by a different apoptotic process. RT-induced apoptosis of Jurkat cells was attenuated by YVAD-CMK as well as DEVD-CHO. Increased caspase activity on DEVD-AMC, which was inhibited by both YVAD-CMK and DEVD-CHO added to the cell culture, was also detected. The involvement of caspase-3 itself, however, was not recognized by Western blot analysis. In contrast, the processing of caspase-3 was observed in the apoptotic HL-60 cells. These data implicate the presence of the redundant processes of apoptosis induced by RT treatment.
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PMID:Room temperature-induced apoptosis of Jurkat cells sensitive to both caspase-1 and caspase-3 inhibitors. 1039 47

Apoptosis is an important mechanism for regulating the numbers of monocytes and macrophages. Caspases (cysteine-aspartate-specific proteases) are key molecules in apoptosis and require proteolytic removal of prodomains for activity. Caspase-1 and caspase-3 have both been connected to apoptosis in other model systems. The present study attempted to delineate what role these caspases play in spontaneous monocyte apoptosis. In serum-free conditions, monocytes showed a commitment to apoptosis as early as 4 h in culture, as evidenced by caspase-3-like activity. Apoptosis, as defined by oligonucleosomal DNA fragmentation, was prevented by a generalized caspase inhibitor, z-VAD-FMK, and the more specific caspase inhibitor, z-DEVD-FMK. The caspase activity was specifically attributable to caspase-3 by the identification of cleavage of procaspase-3 to active forms by immunoblots and by cleavage of the fluorogenic substrate DEVD-AFC. In contrast, a caspase-1 family inhibitor, YVAD-CMK, did not protect monocytes from apoptosis, and the fluorogenic substrate YVAD-AFC failed to show an increase in activity in apoptotic monocytes. When cultured with LPS (1 microgram/ml), monocyte apoptosis was prevented, as was the activation of caspase-3. Unexpectedly, LPS did not change baseline caspase-1 activity. These findings link spontaneous monocyte apoptosis to the proteolytic activation of caspase-3.
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PMID:Spontaneous human monocyte apoptosis utilizes a caspase-3-dependent pathway that is blocked by endotoxin and is independent of caspase-1. 1043 6


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