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)

Accelerated apoptosis is one mechanism proposed for the loss of CD4+ T-lymphocytes in human immunodeficiency virus type 1 (HIV-1) infection. The HIV-1 envelope glycoprotein, gp160, contains two C-terminal calmodulin-binding domains. Expression of gp160 in Jurkat T-cells results in increased sensitivity to FAS- and ceramide-mediated apoptosis. The pro-apoptotic effect of gp160 expression is blocked by two calmodulin antagonists, tamoxifen and trifluoperazine. This enhanced apoptosis in response to FAS antibody or C(2)-ceramide is associated with activation of caspase 3, a critical mediator of apoptosis. A point mutation in the C-terminal calmodulin-binding domain of gp160 (alanine 835 to tryptophan, A835W) eliminates gp160-dependent enhanced FAS-mediated apoptosis in transiently transfected cells, as well as in vitro calmodulin binding to a peptide corresponding to the C-terminal calmodulin-binding domain of gp160. Stable Tet-off Jurkat cell lines were developed that inducibly express wild type gp160 or gp160A835W. Increasing expression of wild type gp160, but not gp160A835W, correlates with increased calmodulin levels, increased apoptosis, and caspase 3 activation in response to anti-FAS treatment. The data indicate that gp160-enhanced apoptosis is dependent upon calmodulin up-regulation, involves the activation of caspase 3, and requires calmodulin binding to the C-terminal binding domain of gp160.
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PMID:Requirement of calmodulin binding by HIV-1 gp160 for enhanced FAS-mediated apoptosis. 1062 68

3-Amino-1,4-dimethyl-5H-pyrido[4,3-b]indole (Trp-P-1), one of the tryptophan pyrolysates, is a dietary carcinogen and is formed in cooked meat and fish in our daily diet. Trp-P-1 will affect the cells in the blood circulation system before it causes carcinogenicity in target organs such as the liver. In this study, the cytotoxicity of Trp-P-1 was investigated in mononuclear cells (MNCs) from blood. Trp-P-1 (10-15 microM) decreased cell viability and induced apoptosis characterized both by morphological changes and by DNA fragmentation 4 h after treatment. DNA fragmentation was also observed following treatment at 1 nM after 24 h in culture. This result suggested that apoptosis would occur in the body following unexpected intake of foods containing Trp-P-1. To determine the mechanism of apoptosis, we investigated the activation of the caspase cascade in MNCs. Trp-P-1 (10-15 microM) activated the caspase cascade, i.e. the activity of caspase-3, -6, -7, -8 and -9 increased dose-dependently using peptide substrates, the active forms of caspase-3, -8 and -9 were detected by immunoblotting, and cleavage of poly(ADP-ribose) polymerase and protein kinase C-delta as the intracellular substrates for caspases was observed. A peptide inhibitor of caspase-8 completely suppressed activation of all other caspases, while an inhibitor of caspase-9 did not. These results indicated that caspase-8 may act as an apical caspase in the Trp-P-1-activated cascade.
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PMID:3-Amino-1,4-dimethyl-5H-pyrido[4,3-b]indole (Trp-P-1) induces caspase-dependent apoptosis in mononuclear cells. 1138 67

This study assessed the changes in the isoprenoid pathway and its metabolites digoxin, dolichol and ubiquinone in neoplasms (CNS astrocytomas - glioblastoma multiforme and high grade non - Hodgkin's lymphoma). The following parameters were assessed-isoprenoid pathway metabolites, tyrosine and tryptophan catabolites, glycoconjugate metabolism, RBC membrane composition and free radical metabolism. There was an elevation in plasma HMG CoA reductase activity, serum digoxin and dolichol and a reduction in RBC membrane Na+-K+ ATPase activity, serum ubiquinone and magnesium levels. Serum tryptophan, serotonin, nicotine and quinolinic acid were elevated while tyrosine, dopamine, noradrenaline and morphine were decreased. The total serum glycosaminoglycans and glycosaminoglycan fractions (except dermatan sulphate in the case of CNS astrocytomas), the activity of GAG degrading enzymes and glycohydrolases, carbohydrate residues of glycoproteins and serum glycolipids were elevated. HDL cholesterol showed a significant decrease and free fatty acids & triglycerides were increased. The RBC membrane glycosaminoglycans, hexose and fucose residues of glycoproteins and phospholipids were reduced. The activity of all free radical scavenging enzymes, concentration of glutathione, iron binding capacity and ceruloplasmin decreased significantly while the concentration of malondialdehyde (MDA), hydroperoxides, conjugated dienes and NO increased. The concentration of alpha tocopherol was unaltered. Membrane Na+-K+ ATPase inhibition due to elevated digoxin, altered membrane structure and digoxin related tyrosine / tryptophan transport defect leading to increased levels of depolarising tryptophan catabolites and decreased levels of hyperpolarising tyrosine catabolites can lead to alteration in intracellular calcium/magnesium ratios and oncogene activation. Intracellular magnesium deficiency can produce defective microtubule related spindle fibre dysfunction and chromosomal non-dysjunction contributing to neoplastic cellular polyploidy and aneuploidy. Digoxin induced tryptophan/tyrosine transport defect can alter neurotransmitter patterns with increased serotonin, quinolinic acid, nicotine & glutamatergic transmission and reduced dopamine, morphine and noradrenaline levels leading to oncogenesis. Glycoconjugate metabolism is altered by elevated dolichol levels and magnesium depletion consequent to Na+-K+ ATPase inhibition. There is a qualitative alteration in proteoglycans and glycoproteins, defective membrane formation and structure and reduced lysosomal stability leading to disordered contact inhibition and tumour antigen presentation contributing to oncogenesis. Digoxin induced alteration in intracellular calcium/magnesium ratios and low ubiquinone levels can lead to a mitochondrial dysfunction resulting in increased free radical generation and reduced scavenging & caspase-3 activation producing a P21 defect contributing to oncogenesis.
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PMID:Hypothalamic digoxin mediated model for oncogenesis. 1187 54

Mitochondria act as a focal point for upstream apoptosis signals by releasing cytochrome c into the cytosol, leading to the activation of caspases and subsequent cell death. Members of the Bcl-2 protein family regulate this phenomenon by heterodimerization via the BH3 domain of proapoptotic members opposing their pro- and antiapoptotic functions. The mechanism of cytochrome c release from mitochondria and of its regulation remains controversial. In vitro binding studies of purified and biologically active proteins should help in understanding the molecular mechanism of interactions and protein functions. In this work, the Bcl-2-related antiapoptotic chicken protein Nr-13 was overexpressed as a highly soluble recombinant protein which showed correct folding as judged by circular dichroism and fluorescence spectroscopy. Purified Nr-13 inhibits caspase-3 activation in a Xenopus egg-derived cell-free system, and neutralizes the proapoptotic activity of a synthetic peptide containing the BH3 domain of Bax. The latter effect correlates with the high-affinity binding of the BH3 peptide to Nr-13 as monitored by the intrinsic tryptophan fluorescence. On the basis of the structural similarity with Bcl-x(L), putative residues involved in this interaction were identified. Nr-13 exhibits a high-affinity interaction with cytochrome c which is prevented by preincubation with the BH3-Bax peptide. These findings are discussed with respect to a model for the regulation of apoptosis in which a direct interaction between the antiapoptotic protein and cytochrome c may prevent the apoptosis.
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PMID:Interaction between the antiapoptotic protein Nr-13 and cytochrome c. Antagonistic effect of the BH3 domain of Bax. 1209 70

The active site of an apoptotic enzyme caspase-3 was characterized by measuring the intrinsic fluorescence of two tryptophan residues. Temperature dependence of the intrinsic fluorescence, the energy homotransfer between the tryptophan residues, and the fluorescence quenching by tetrapeptide inhibitors were investigated by the fluorescence lifetime measurements. It has been observed that the fluorescence lifetimes of caspase-3 in complex with inhibitors were significantly shortened by the electron transfer process.
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PMID:Probing the caspase-3 active site by fluorescence lifetime measurements. 1214 46

3-Amino-1,4-dimethyl-5H-pyrido[4,3-b]indole (Trp-P-1), which is a tryptophan pyrolysate formed during cooking, induces apoptosis in rat splenocytes, thymocytes, and hepatocytes. In this study, we investigated whether Trp-P-1 is transported into these cells and causes apoptosis. Trp-P-1 was immediately incorporated into rat splenocytes, thymocytes, and hepatocytes in a dose- and time-dependent manner. Dopamine and serotonin significantly competed with the uptake of Trp-P-1 into these cells, and nomifensine and indatraline, which are inhibitors of dopamine- and serotonin-transporters, respectively, markedly suppressed the uptake of Trp-P-1. On the other hand, amino acids including tryptophan did not compete with Trp-P-1. Inhibition of monoamine transporters using nomifensine and indatraline partially suppressed Trp-P-1-induced cell death in these cells. In hepatocytes, the inhibition of transporters prevented Trp-P-1-induced morphological changes and activation of caspase-3. These results demonstrated that Trp-P-1 is incorporated into the cells through monoamine transporters and induces apoptosis.
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PMID:3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole (trp-P-1) is incorporated into rat splenocytes, thymocytes, and hepatocytes through monoamine transporters and induces apoptosis. 1216 39

Donor cells can be preserved in University of Wisconsin (UW), histidine-tryptophan-ketoglutarate (HTK), or Celsior solution. However, differences in efficacy and mode of action in preventing hypothermia-induced cell injury have not been unequivocally clarified. Therefore, we investigated and compared necrotic and apoptotic cell death of freshly isolated primary porcine hepatocytes after hypothermic preservation in UW, HTK, and Celsior solutions and subsequent normothermic culturing. Hepatocytes were isolated from porcine livers, divided in fractions, and hypothermically (4 degrees C) stored in phosphate-buffered saline (PBS), UW, HTK, or Celsior solution. Cell necrosis and apoptosis were assessed after 24- and 48-h hypothermic storage and after 24-h normothermic culturing following the hypothermic preservation periods. Necrosis was assessed by trypan blue exclusion, lactate dehydrogenase (LDH) release, and mitochondrial 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) reduction. Apoptosis was assessed by the induction of histone-associated DNA fragments and cellular caspase-3 activity. Trypan blue exclusion, LDH release, and MTT reduction of hypothermically preserved hepatocytes showed a decrease in cell viability of more than 50% during the first 24 h of hypothermic preservation. Cell viability was further decreased after 48-h preservation. DNA fragmentation was slightly enhanced in hepatocytes after preservation in all solutions, but caspase-3 activity was not significantly increased in these cells. Normothermic culturing of hypothermically preserved cells further decreased cell viability as assessed by LDH release and MTT reduction. Normothermic culturing of hypothermically preserved hepatocytes induced DNA fragmentation, but caspase-3 activity was not hanced in these cells. Trypan blue exclusion, LDH leakage, and MTT reduction demonstrated the highest cell viability after storage in Celsior, and DNA fragmentation was the lowest in cells that had been stored in PBS and UW solutions. None of the preservation solutions tested in this study was capable of adequately preventing cell death of isolated porcine hepatocytes after 24-h hypothermic preservation and subsequent 24-h normothermic culturing. Culturing of isolated and hypothermically preserved hepatocytes induces DNA fragmentation, but does not lead to caspase-3 activation. With respect to necrosis and DNA fragmentation of hypothermically preserved cells, UW and Celsior were superior to PBS and HTK solutions in this model of isolated porcine hepatocyte preservation.
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PMID:Induction of necrosis and DNA fragmentation during hypothermic preservation of hepatocytes in UW, HTK, and Celsior solutions. 1269 65

Caspase-3, one of the major apoptotic proteins, is a cysteine protease and exists as an inactive zymogen in healthy cells. In this study, the dynamic nature of the rearrangements of two tryptophan residues (Trp 206 and Trp 214) in the active sites of caspase-3 during the activation was analyzed by measuring the fluorescence lifetimes. Significant changes in the lifetime occurred upon activation by the specific cleavage. In addition, two mutant proteins that have only one tryptophan residue also showed the similar changes. These data indicate that the activation of caspase-3 resulted in the reorganization of both tryptophan residues.
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PMID:Rearrangement of tryptophan residues in caspase-3 active site upon activation. 1521 Jan 19

We have examined the role of a salt bridge between Lys242 and Glu246 in loop L4 of procaspase 3 and of mature caspase 3, and we show that the interactions are required for stabilizing the active site. Replacing either of the residues with an alanine residue results in a complete loss of procaspase 3 activity. Although both mutants are active in the context of the mature caspase 3, the mutations result in an increase in K(m) and a decrease in kcat when compared with the wild-type caspase 3. In addition, the mutations result in an increase in the pK(a) value associated with a change in kcat with pH, but does not affect the transition observed for Km versus pH. The mutations also affect the accessibility of the active-site solvent as measured by tryptophan fluorescence emission in the presence of quenching agents and as a function of pH. We show that, as the pH is lowered, the (pro)caspase dissociates, and the mutations increase the pH-dependent instability of the dimer. Overall, the results suggest that the contacts lost in the procaspase as a result of replacing Lys242 and Glu246 are compensated partially in the mature caspase as a result of new contacts that are known to form on zymogen processing
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PMID:Ionic interactions near the loop L4 are important for maintaining the active-site environment and the dimer stability of (pro)caspase 3. 1531 47

Indoleamine-2,3-dioxygenase (IDO) is an intracellular enzyme present in dendritic cells and macrophages. It is a known modulator of T-cell response and contributes to the UV protection of the lens. There yet is no information on IDO activity in the corneal endothelium, protecting the endothelial cells from light mediated damage. We exposed murine corneal endothelial cells (MCEC) with different doses of UV-B light 280-320 nm, probed for IDO mRNA (real-time PCR) and assessed apoptosis rate (flow cytometry) and caspase-3-activity in the cells. The metabolites of the IDO catalysed reaction, l-kynurenine, was also measured. Malondialdehyde was detected for quantification of UV-B-induced oxidative stress. To investigate specificity, IDO effects were blocked by 1-methyl-tryptophan. The effects of IDO overexpression in the MCEC were assessed by transfection of an expression vector. MCEC consistently express IDO at low levels. Exposure to UV-B light led to a dose-responding upregulation of IDO; IDO was found competent converting l-tryptophan into l-kynurenine. Irradiation led to increased apoptosis and caspase-3-activity of MCEC. Supplementation of l-kynurenine or overexpression of IDO in the MCEC could reduce apoptosis significantly following UV-B irradiation. Inhibition of IDO by 1-MT was potent to reverse this effect. IDO and its metabolite l-kynurenine can protect corneal endothelial cells from UV-B-induced oxidative stress and apoptosis. It may be an active protection mechanism against corneal endothelial damage.
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PMID:Indoleamine 2,3-dioxygenase protects corneal endothelial cells from UV mediated damage. 1631 52


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