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)

Recent evidence suggests that members of the interleukin-1-beta-converting enzyme (ICE)/Ced-3 family are key mediators of mammalian apoptosis. The known members of the ICE/Ced-3 cysteine protease family are synthesized as proenzymes and require proteolytic processing to produce active, heterodimeric enzymes. The baculovirus protein P35 has recently been shown to inhibit several members of the ICE/Ced-3 cysteine protease family. The importance of ICE/Ced-3 cysteine proteases in programmed cell death prompted us to investigate the role of the apoptotic mediator, CPP32, in the glucocorticoid-mediated cell death pathway. Glucocorticoids induce growth inhibition and apoptosis in sensitive leukemic cell lines, immature thymocytes, and eosinophils. In this report, we demonstrate the enzymatic cleavage of proCPP32 to its active subunits in cells undergoing glucocorticoid-induced apoptotic cell death. Concurrently, in apoptotic cells, PARP, a 116-kilodalton (kDa) human poly(ADP-ribose) polymerase, is proteolytically cleaved to its signature 85-kDa fragment. The proteolytic processing of PARP (the nuclear DNA repair enzyme known to be cleaved in association with apoptosis) is catalyzed by members of the ICE/Ced-3 family. Importantly, stable transfection of the antiapoptotic baculovirus P35 inhibits glucocorticoid-induced apoptotic cell death, proteolytic processing of proCPP32, and cleavage of the 116kDa PARP. We conclude that activation of CPP32 is a critical event in glucocorticoid-induced apoptosis and that this pathway is inhibited at or upstream of CPP32 by baculovirus P35. These data demonstrate that PARP cleavage occurs during glucocorticoid-induced apoptotic cell death and show that this proteolytic process is blocked by the expression of baculovirus P35, supporting a role for activation of the ICE/Ced-3-like cysteine protease during glucocorticoid-induced apoptosis.
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PMID:Baculovirus P35 inhibits the glucocorticoid-mediated pathway of cell death. 898 38

We have investigated the ability of Sf-caspase-1 and two mammalian caspases, caspase-1 and caspase-3, to induce apoptosis in Spodoptera frugiperda Sf-21 insect cells. While the transient expression of the pro-Sf-caspase-1 did not induce apoptosis, expression of the pro-domain deleted form, p31, or coexpression of the two subunits of mature Sf-caspase-1, p19 and p12, induced apoptosis in Sf-21 cells. The behavior of Sf-caspase-1 resembled that of the closely related mammalian caspase, caspase-3, and contrasted with that of the mammalian caspase-1, the pro-form of which was active in inducing apoptosis in Sf-21 cells. The baculovirus caspase inhibitor P35 blocked apoptosis induced by active forms of all three caspases. In contrast, members of the baculovirus inhibitor of apoptosis (IAP) family failed to block active caspase-induced apoptosis. However, during viral infection, expression of OpIAP or CpIAP blocked the activation of pro-Sf-caspase-1 and the associated induction of apoptosis. Thus, the mechanism by which baculovirus IAPs inhibit apoptosis is distinct from the mechanism by which P35 blocks apoptosis and involves inhibition of the activation of pro-caspases like Sf-caspase-1.
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PMID:Baculovirus inhibitors of apoptosis (IAPs) block activation of Sf-caspase-1. 939 Oct 73

The P35 protein derived from the baculovirus Autographa californica NPV has been characterized as an inhibitor of apoptotic cell death in a great number of organisms and situations. This potential has been further mapped to the capacity of P35 to inhibit all caspases investigated. Here we show that P35 does not inhibit caspase-9 activity in a cell-free system of mammalian caspase activation. In cell extracts, cytochrome c addition led to the activation of caspase-9, -3 and -7. When cytosolic extract from cells expressing P35 was added, caspase-9-mediated maturation of caspase-3 proceeded normally but caspase-3-mediated further events were prevented, such as complete processing of caspase-3, processing of caspase-7 and the appearance of DEVD-cleaving activity. The P35 protein from Bombyx mori NPV, which has been reported to have a much weaker anti-apoptosis activity in vivo, was found also to have significant caspase-3-inhibiting activity. These data suggest that P35 evolved specifically to inhibit effector rather than initiator caspases.
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PMID:Baculovirus P35 protein does not inhibit caspase-9 in a cell-free system of apoptosis. 1102 59

Baculovirus P35 is a universal suppressor of apoptosis that stoichiometrically inhibits cellular caspases in a novel cleavage-dependent mechanism. Upon caspase cleavage at Asp-87, the 10- and 25-kDa cleavage products of P35 remain tightly associated with the inhibited caspase. Mutations in the alpha-helix of the reactive site loop preceding the cleavage site abrogate caspase inhibition and antiapoptotic activity. Substitution of Pro for Val-71, which is located in the middle of this alpha-helix, produces a protein that is cleaved at the requisite Asp-87 but does not remain bound to the caspase. This loss-of-function mutation provided the opportunity to structurally analyze the conformational changes of the P35 reactive site loop after caspase cleavage. We report here the 2.7 A resolution crystal structure of V71P-mutated P35 after cleavage by human caspase-3. The structure reveals a large movement in the carboxyl-terminal side of the reactive site loop that swings down and forms a new beta-strand that augments an existing beta-sheet. Additionally, the hydrophobic amino terminus releases and extends away from the protein core. Similar movements occur when P35 forms an inhibitory complex with human caspase-8. These findings suggest that the alpha-helix mutation may alter the sequential steps or kinetics of the conformational changes required for inhibition, thereby causing P35 loss of function.
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PMID:Crystal structure of baculovirus P35 reveals a novel conformational change in the reactive site loop after caspase cleavage. 1140 50

Langat (LGT) flavivirus, derived from infectious full-length cDNA clone 636, was investigated for its apoptotic activities in mouse neuroblastoma (Neuro-2a) and simian kidney (Vero and LLC-MK(2)) cells. The hallmark of apoptosis, cleavage of cellular DNA, was observed 48 h after infection of Vero, LLC-MK(2), and Neuro-2a cells by electrophoresis analysis. Apoptosis in infected cells was also confirmed by TUNEL assay. LGT-infected Neuro-2a cells showed an increase in caspase-3-like protease (DEVDase) activity. Expression of the major envelope glycoprotein (E) alone reduced cell viability in both Vero and Neuro-2a cells, and the baculovirus P35 protein, which inhibits multiple caspases, completely blocked this effect. Cleavage of cellular DNA was observed in E gene-transfected Vero cells by TUNEL assay. Expression of E protein or caspase-9 resulted in activation of caspase-3-like proteases in Neuro-2a cells. The caspase-3-like protease specific inhibitor, Ac-DEVD-CHO peptide, partially inhibited E protein- or caspase-9-induced apoptosis in Neuro-2a cells. These observations indicate that infection of cells with LGT virus or expression of LGT virus E protein induces apoptosis through a caspase-3-like protease pathway.
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PMID:Infection with Langat Flavivirus or expression of the envelope protein induces apoptotic cell death. 1148

We have investigated the effects of expression of the viral proteins CrmA, P35 and IAP, and the three mammalian IAP homologues (MIHA, MIHB and MIHC), on the regulation of apoptosis induced by either the overexpression of caspases (ICE, CPP32 and Nedd2), by serum-deprivation, or by gamma-irradiation in NIH3T3 fibroblasts. As previously shown, CrmA strongly inhibited ICE-induced apoptosis but was ineffective against Nedd2- or CPP32-mediated apoptosis. P35, IAP and MIHA protected cells from apoptosis induced by the three caspases to varying extents but MIHB and MIHC were largely ineffective. NIH3T3 cells expressing P35 and MIHA, but not IAP, CrmA, MIHB and MIHC, showed enhanced cell survival under serum-deprived conditions. In addition, P35, CrmA and MIHA could provide substantial protection against death induced by gamma-irradiation. These results suggest the presence of multiple apoptotic pathways with differential sensitivity to various naturally occurring apoptosis inhibitors.
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PMID:Differential inhibitory effects of CrmA, P35, IAP and three mammalian IAP homologues on apoptosis in NIH3T3 cells following various death stimuli. 1455 70

Cowpox virus (CPV) expresses the serpin (serine proteinase inhibitor) CrmA, an anti-inflammatory, anti-apoptotic protein required for production of red pocks on chicken chorioallantoic membranes (CAMs). In vitro, CrmA inhibits several caspases and granzyme B. Altering the critical P1-aspartate in the CrmA reactive centre loop to alanine resulted in a virus (CPV-CrmA-D303A) that resembled CPV deleted for CrmA (CPVDeltaCrmA : : lacZ); on CAMs it produced white, inflammatory pocks with activated caspase-3 and reduced virus yields, suggesting that CrmA activities are mediated via proteinase inhibition. CrmA in CPV was replaced with SERP2 from Myxoma virus (MYX) or baculovirus P35, which inhibit similar proteinases in vitro. SERP2 and P35 each blocked caspase-3-mediated apoptosis but were unable to control inflammation of CAMs. However, SERP2 and P35 restored virus yields, indicating that the decreased virus titres seen with CPVDeltaCrmA : : lacZ resulted from apoptosis rather than inflammation. To compare the activities of CrmA and SERP2 further, rabbits were infected with MYX recombinant viruses. Intradermal infection of rabbits with MYX was uniformly lethal, generating raised primary lesions and many secondary lesions. In contrast, deletion of SERP2 from MYX (MYXDeltaSERP2 : : lacZ) caused little mortality and produced flat primary lesions with few secondary lesions. Replacement of SERP2 with CrmA (MYXDeltaSERP2 : : CrmA) resulted in partial complementation with flat primary lesions, many secondary lesions and death in 70 % of the rabbits. Therefore, CrmA and SERP2 were not functionally interchangeable during infection of CAMs or rabbits, implying that these serpins have activities that are not evident from biochemical studies with human caspases.
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PMID:Cowpox virus CrmA, Myxoma virus SERP2 and baculovirus P35 are not functionally interchangeable caspase inhibitors in poxvirus infections. 1510 44

The baculovirus protein P35 inhibits apoptosis in a diverse range of animals such as insects, nematodes and mammals. Evidence suggests that P35 can inhibit members of caspase family proteases that are key mediators of mammalian apoptosis. We demonstrate that p35 inhibits activation-induced nitric oxide (NO)-mediated apoptosis in the RAW 264.7 mouse macrophages. Parent or vector-transfected RAW 264.7 cells underwent apoptosis when treated with a combination of cisplatin and interferon-gamma (IFN-gamma) or LPS and IFN-gamma in a NO-dependent manner. By contrast, RAW 264.7 cells stably expressing P35 did not undergo apoptosis when treated with a combination of cisplatin and IFN-gamma or LPS and IFN-gamma. Activation of parent, vector- or p35-transfected cells with cisplatin and IFN-gamma or LPS and IFN-gamma caused equivalent levels of inducible nitric oxide synthase (iNOS) expression and produced equal amounts of nitrite, which ruled out attenuated iNOS activity during P35-mediated protection. Rather, expression of P35 inhibited translocation of mitochondrial cytochrome c into cytosol, mitochondrial depolarization, activation of caspase-9 and caspase-3, and cleavage of poly (ADP-ribose) polymerase (PARP). These findings indicate that P35 inhibits NO-induced apoptotic cell death of activated macrophages by inhibiting mitochondrial cytochrome c release, which suggests that P35 has targets upstream of the caspase cascade in apoptosis.
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PMID:Baculovirus P35 inhibits NO-induced apoptosis in activated macrophages by inhibiting cytochrome c release. 1517 17

In most retinal degenerations in humans and in animal models, photoreceptor cells die by apoptosis. Although the biochemical features are similar in all apoptotic cells, different molecular events lead the cell to death. In the present study we used a rat model of inherited retinal degeneration, the RCS rats, to investigate the involvement of the proteases, caspases and/or calpains, in photoreceptor apoptosis. In the first experiments, rats were untreated or injected intravitreally at post natal day 27 (P27) with the large broad spectrum caspase inhibitor, ZVAD, the calpain inhibitor, MuhPhe, or with the vehicle, DMSO. Retinal status was evaluated at P35 and P42 by electroretinography, morphometry and apoptotic nuclei detection. DMSO and MuhPhe had no effect on RCS retinas as evidenced by equivalent loss of function and equivalent number of apoptotic cells than in untreated group. ZVAD transiently reduced apoptotic cells and preserved photoreceptor function at P35 but not at P42. These results suggest that caspases but not calpains are involved in retinal degeneration in the RCS. In the second experiments, RCS rats were injected twice at P27 and P35 with ZVAD or DMSO. Although ZVAD-treated retinas were preserved at P35 compared to the DMSO controls, the second injection of ZVAD did not extend the preserving effect to P42. Moreover, a single injection of ZVAD at P35 had no preserving effect at P42. All these data taken together suggest that caspases do not play a pivotal role after P35. In a fourth set of experiments, we used specific caspase inhibitors to elucidate which caspase was activated. The caspase-1/4 inhibitor (YVAD) or the caspase-3/7 inhibitor (DEVD) were injected intravitreally at P27 and retinal status was evaluated at P35 and P42. Electroretinograms and apoptotic nuclei detection demonstrated that YVAD and DEVD preserved photoreceptors at P35 but not at P42. These results suggest that both caspase-1/4 and caspase-3/7 play a major role in the apoptotic pathway between P27 and P35 in retinal degeneration of RCS rats. In this study, we show that 1/ the photoreceptor apoptotic process in the RCS rat involves caspases but not calpains, and 2/ the retinal degeneration seems to be composed of different phases involving different molecular players. Indeed, we have demonstrated that caspases are playing a major role at P35, but not at P42.
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PMID:Transient protective effect of caspase inhibitors in RCS rat. 1827 51

Birdsongs and the regions of their brain that control song exhibit obvious sexual differences. However, the mechanisms underlying these sexual dimorphisms remain unknown. To address this issue, we first examined apoptotic cells labeled with caspase-3 or TUNEL in Bengalese finch song control nuclei - the robust nucleus of the archopallium (RA), the lateral magnocellular nucleus of the anterior nidopallium (LMAN), the high vocal center (HVC) and Area X from post-hatch day (P) 15 to 120. Next, we investigated the expression dynamics of pro-apoptotic (Bid, Bad and Bax) and anti-apoptotic (Bcl-2 and Bcl-xL) genes in the aforementioned nuclei. Our results revealed that the female RA at P45 exhibited marked cell apoptosis, confirmed by low densities of Bcl-xL and Bcl-2. Both the male and female LMAN exhibited apoptotic peaks at P35 and P45, respectively, and the observed cell loss was more extensive in males. A corresponding sharp decrease in the density of Bcl-2 after P35 was observed in both sexes, and a greater density of Bid was noted at P45 in males. In addition, we observed that RA volume and the total number of BDNF-expressing cells decreased significantly after unilateral lesion of the LMAN or HVC (two areas that innervate the RA) and that greater numbers of RA-projecting cells were immunoreactive for BDNF in the LMAN than in the HVC. We reasoned that a decrease in the amount of BDNF transported via HVC afferent fibers might result in an increase in cell apoptosis in the female RA. Our data indicate that cell apoptosis resulting from different pro- and anti-apoptotic agents is involved in generating the differences between male and female song control nuclei.
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PMID:Sexual Differences in Cell Loss during the Post-Hatch Development of Song Control Nuclei in the Bengalese Finch. 2593 74


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