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Query: UNIPROT:P42574 (caspase-3)
 45,978 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

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

We prepared a cleavage site-directed antiserum against Caspase-3 (anti-p20/17), which reacts with the p20/17 fragment (p20/17) activated by cleavage but not proCaspase-3 (p32), and examined the relationship between the activation of Caspase-3 and apoptosis. We identified p20/17-positive cells where cell death occurs naturally: interdigits of the forelimbs, small intestine epithelium, thymus, trigeminal ganglia, and dorsal root ganglia of mouse embryos. Withdrawal of nerve growth factor induced the appearance of p20/17-positive cells with DNA fragmentation in the culture of dorsal root ganglia neurons, while DNA fragmentation was detected in both p20/17-positive and -negative neurons in dorsal root ganglia of mouse embryos. These results suggest that not only activation of Caspase-3 but also other molecular mechanism play a role in the naturally occurring dorsal root ganglia apoptosis. Cleavage site-directed antisera against Caspases will be useful for the analysis of the molecular mechanism of naturally occurring apoptosis during development.
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PMID:Detection of activated Caspase-3 by a cleavage site-directed antiserum during naturally occurring DRG neurons apoptosis. 964 70

Caspase-mediated proteolysis is a critical and central element of the apoptotic process; therefore, it is important to identify the downstream molecular targets of caspases. We established a method for cloning the genes of caspase substrates by two major modifications of the yeast two-hybrid system: (i) both large and small subunits of active caspases were expressed in yeast under ADH1 promoters and the small subunit was fused to the LexA DNA-binding domain; and (ii) a point mutation was introduced that substituted serine for the active site cysteine and thereby prevented proteolytic cleavage of the substrates, possibly stabilizing the enzyme-substrate complexes in yeast. After screening a mouse embryo cDNA expression library by using the bait plasmid for caspase-3, we obtained 13 clones that encoded proteins binding to caspase-3, and showed that 10 clones including gelsolin, an actin-regulatory protein implicated in apoptosis, were cleaved by recombinant caspase-3 in vitro. Using the same bait, we also isolated human gelsolin cDNA from a human thymus cDNA expression library. We showed that human gelsolin was cleaved during Fas-mediated apoptosis in vivo and that the caspase-3 cleavage site of human gelsolin was at D352 of DQTD352G, findings consistent with previous observations on murine gelsolin. In addition, we ascribed the antiapoptotic activity of gelsolin (which we previously reported) to prevention of a step leading to cytochrome c release from the mitochondria into the cytosol. Our results indicate that this cloning method is useful for identification of the substrates of caspases and possibly also of other enzymes.
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PMID:A cloning method for caspase substrates that uses the yeast two-hybrid system: cloning of the antiapoptotic gene gelsolin. 967 12

Caspases are a family of cysteine proteases of critical importance in the apoptotic cell death process. They are normally present as zymogens (pro-caspases) in the cytoplasm of vertebrate and other organisms. In this study we have shown that pro-caspase-3 is localized to cytosol and mitochondria of various rat tissues (brain, heart, kidney, liver, spleen and thymus). Although the majority of pro-caspase-3 was localized in the cytosol, the amount of mitochondrial pro-caspase-3 was significant. The ratio of cytosolic and mitochondrial pools of pro-caspase-3 appeared to vary between different tissues. The higher amount of mitochondrial pro-caspase-3 was found in thymus and spleen, i.e. tissues in which spontaneous apoptosis plays an important role. Our findings provide further support for mitochondrial localization of pro-caspase-3 and the critical role of this organelle in apoptosis.
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PMID:Detection of pro-caspase-3 in cytosol and mitochondria of various tissues. 970 95

Retinoids play an important role in the control of lymphocyte function and homeostasis in the thymus. In this study, we show that the induction of growth arrest and apoptosis in a variety of T-cell lymphoma cell lines, including Jurkat and Molt-4 cells, is highly specific for the synthetic retinoid 6-[3-(1-adamantyl)-4-hydroxyphenyl]-2-naphthalene carboxylic acid (AHPN) since all-trans retinoic acid (RA), the RAR-selective retinoid TTAB, the RXR-selective retinoid SR11217 and the retinoid SR11302 exhibiting selective anti-AP1 activity, do not induce apoptosis or cause growth arrest. These findings support the concept that the effects of AHPN on proliferation and induction of apoptosis are mediated by a novel signaling pathway. AHPN-induced apoptosis is associated with an induction of internucleosomal DNA-fragmentation, increased annexin V binding and a 30-fold stimulation of caspase-3-like activity. Overexpression of Bcl-2 in Molt-4 cells greatly inhibits the induction of apoptosis by AHPN as indicated by the inhibition of DNA-fragmentation, annexin V binding and caspase-3-like activity. However, Bcl-2 overexpression does not interfere with the ability of AHPN to cause growth arrest or accumulation of cells in the early S-phase of the cell cycle, indicating that the effects of AHPN on growth arrest can be uncoupled from the effects on apoptosis. The caspase inhibitor Z-VAD-FMK, at concentrations that totally block caspase activity, delays but does not prevent cell death and does not affect the accumulation of cells in the S-phase of the cell cycle. Our results show that induction of caspase-3-like activity plays an important role in the execution of AHPN-induced apoptosis but cells can undergo cell death in the absence of this activity suggesting that AHPN-induced cell death involves caspase-dependent and -independent mechanisms.
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PMID:Induction of apoptosis by the novel retinoid AHPN in human T-cell lymphoma cells involves caspase-dependent and independent pathways. 984 84

The inhibitor of apoptosis (IAP) proteins form a highly conserved gene family that prevents cell death in response to a variety of stimuli. Herein we describe a newly defined murine IAP, designated Tiap, that proved to be a murine homologue of human survivin based on sequence comparison. TIAP has one baculovirus IAP repeat and lacks a C-terminal RING finger motif. TIAP interacted with the processed form of caspase 3 and inhibited caspase-induced cell death. Histological examinations revealed that TIAP is expressed in growing tissues such as thymus, testis, and intestine of adult mice and many tissues of embryos. In in vitro studies, TIAP was induced in splenic T cells activated with anti-CD3 antibody or Con A, and the expression of TIAP was up-regulated in synchronized NIH 3T3 cells at S to G2/M phase of the cell cycle. We propose that during cell proliferation, cellular protective activity may be augmented with inducible IAPs such as TIAP.
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PMID:Expression of a murine homologue of the inhibitor of apoptosis protein is related to cell proliferation. 999 45

Aging is associated with altered immune function. We previously reported that splenocytes and thymocytes undergo apoptosis with aging in rats. In the present study, we examined the expression of genes associated with apoptosis in splenocytes and thymus in aging rats. We evaluated the expression of bax, interleukin 1-beta-converting enzyme (ICE)/ced-3 protease family, caspase-3 and tumor suppressor gene p53. Rats in age groups of 6, 24, 48, and 96 weeks were sacrificed; thymocytes and splenocytes were isolated followed by lysis in a modified RIPA buffer containing protease inhibitors. Western blot analysis of proteins was performed by probing immunoblots with antibodies against p53, bax and PARP (poly ADP-ribose polymerase). Increased aging was associated with enhanced expression of bax, p53 and cleavage of PARP by Caspase-3. The expression of p53 and cleavage of PARP indicates the presence of damaged DNA; nevertheless, the cleavage of PARP or activation of caspase-3 may be playing an important role in the initiation of early events in apoptosis. These results suggest that aging of splenocytes and thymocytes is associated with the expression of cell death genes. The present study provides an insight into age-associated altered immune function.
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PMID:Aging splenocyte and thymocyte apoptosis is associated with enhanced expression of p53, bax, and caspase-3. 1032 82

By using the amino acid sequence motif of tumor necrosis factor (TNF), we searched the expressed sequence tag data base and identified a novel full-length cDNA encoding 285 amino acid residues and named it THANK. THANK is a type II transmembrane protein with 15-20% overall amino acid sequence homology to TNF, LT-alpha, FasL, and LIGHT, all members of the TNF family. The mRNA for THANK was expressed at high levels by peripheral blood leukocytes, lymph node, spleen, and thymus and at low levels by small intestine, pancreas, placenta, and lungs. THANK was also prominently expressed in hematopoietic cell lines. The recombinant purified protein expressed in the baculovirus system had an approximate molecular size 20 kDa with amino-terminal sequence of AVQGP. Treatment of human myeloid U937 cells with purified THANK activated nuclear transcription factor-kappaB (NF-kappaB) consisting of p50 and p65. Activation was time- and dose-dependent, beginning with as little as a 1 pM amount of the cytokines and as early as 15 min. Under the same conditions, THANK also activated c-jun NH2-terminal kinase (JNK) in U937 cells. THANK also strongly suppressed the growth of tumor cell lines and activated caspase-3. Although THANK had all the activities and potency of TNF, it did not bind to the TNF receptors. Thus our results indicate that THANK is a novel cytokine that belongs to the TNF family and activates apoptosis, NF-kappaB, and JNK through a distinct receptor.
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PMID:Identification and characterization of a novel cytokine, THANK, a TNF homologue that activates apoptosis, nuclear factor-kappaB, and c-Jun NH2-terminal kinase. 1034 44

The biological function of CD30 in the thymus has been only partially elucidated, although recent data indicate that it may be involved in negative selection. Because CD30 is expressed only by a small subpopulation of medullary thymocytes, we generated transgenic (Tg) mice overexpressing CD30 in T lymphocytes to further address its role in T cell development. CD30 Tg mice have normal thymic size with a normal number and subset distribution of thymocytes. In vitro, in the absence of CD30 ligation, thymocytes of CD30 Tg mice have normal survival and responses to apoptotic stimuli such as radiation, dexamethasone, and Fas. However, in contrast to controls, CD30 Tg thymocytes are induced to undergo programmed cell death (PCD) upon cross-linking of CD30, and the simultaneous engagement of TCR and CD30 results in a synergistic increase in thymic PCD. CD30-mediated PCD requires caspase 1 and caspase 3, is not associated with the activation of NF-kappaB or c-Jun, but is totally prevented by Bcl-2. Furthermore, CD30 overexpression enhances the deletion of CD4+/CD8+ thymocytes induced by staphylococcal enterotoxin B superantigen and specific peptide. These findings suggest that CD30 may act as a costimulatory molecule in thymic negative selection.
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PMID:CD30 overexpression enhances negative selection in the thymus and mediates programmed cell death via a Bcl-2-sensitive pathway. 1038 16

To date, in vivo apoptosis within the thymus has been assessed using morphological criteria and/or detection of a DNA ladder indicative of oligonucleosomal fragmentation of the DNA. Here, we have used a fluorometric method to investigate activation of the caspase protease family in the thymus following in vivo induction of apoptosis by injection of the synthetic glucocorticoid hydrocortisone. Cleavage of DEVD-MCA by caspase-3 and other group II caspases releases free MCA which can be detected fluorimetrically. We demonstrate a time-dependent increase in DEVD-MCA cleavage activity within this tissue indicating the activation of caspase-3 like enzymes. This activity was inhibited by the specific group II caspase inhibitor DEVD-CHO. The interpretation of increased caspase activity was confirmed by immunoblot analysis to reveal cleavage of the caspase-3 substrate, fodrin. In addition, agarose gel electrophoresis of the DNA yielded a ladder pattern, confirming the occurrence of apoptosis. This study demonstrates that DEVD-MCA cleavage activity may be a useful quantitative method for the analysis of apoptosis in thymus tissue. It is a relatively rapid procedure not requiring thymocyte isolation or gel electrophoresis and detects fairly early biochemical changes occurring during apoptosis. In the present study we have used this method to demonstrate the involvement of caspases in thymocyte apoptotic death induced in vivo by glucocorticoids. Thus, measurement of caspase activity in thymus tissue may have applications for studying the in vivo effects of immunotoxicants.
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PMID:Application of a fluorometric assay to detect caspase activity in thymus tissue undergoing apoptosis in vivo. 1041 Sep 70


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