Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: 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)

Intact fibronectin (FN) protects cells from apoptosis. When FN is fragmented, specific domains induce proteinase expression in fibroblasts. However, it is not known whether specific domains of FN can also regulate apoptosis. We exposed fibroblasts to four recombinant FN fragments and then assayed for apoptosis using criteria of cellular shape change, condensed nuclear morphology, and DNA fragmentation. The fragments extended from the RGD-containing repeat III10 to III15; they included (V(+)) or excluded (V(-)) the alternatively spliced V region and contained either a mutated (H(-)) or an unmutated (H(+)) heparin binding domain. Only the V(+)H(-) fragment triggered decreases in pp125(FAK) levels and apoptosis, which was rescued by intact FN and inhibitors of caspase-1 and caspase-3. This apoptotic mechanism was mediated by a chondroitin sulfate proteoglycan, since treating cells with chondroitin sulfate or chondroitinase reversed the apoptotic cell shape changes. The alpha4 integrin receptor may also be involved, since using a blocking antibody to alpha4 alone induced apoptotic cell shape changes, whereas co-treatment with this antibody plus V(+)H(+) reversed these effects. These results demonstrate that the V and heparin binding domains of FN modulate pp125(FAK) levels and regulate apoptosis through a chondroitin sulfate proteoglycan- and possibly alpha4 integrin-mediated pathway, which triggers a caspase cascade.
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PMID:Mutations in the heparin binding domain of fibronectin in cooperation with the V region induce decreases in pp125(FAK) levels plus proteoglycan-mediated apoptosis via caspases. 1052 84

Programmed cell death or apoptosis leads to the activation of the caspase-activated DNase (CAD), which degrades chromosomal DNA into nucleosomal fragments. Biochemical studies revealed that CAD forms an inactive heterodimer with the inhibitor of caspase-activated DNase (ICAD), or its alternatively spliced variant, ICAD-S, in the cytoplasm. It was initially proposed that proteolytic cleavage of ICAD by activated caspases causes the dissociation of the ICAD/CAD heterodimer and the translocation of active CAD into the nucleus in apoptotic cells. Here, we show that endogenous and heterologously expressed ICAD and CAD reside predominantly in the nucleus in nonapoptotic cells. Deletional mutagenesis and GFP fusion proteins identified a bipartite nuclear localization signal (NLS) in ICAD and verified the function of the NLS in CAD. The two NLSs have an additive effect on the nuclear targeting of the CAD-ICAD complex, whereas ICAD-S, lacking its NLS, appears to have a modulatory role in the nuclear localization of CAD. Staurosporine-induced apoptosis evoked the proteolysis and disappearance of endogenous and exogenous ICAD from the nuclei of HeLa cells, as monitored by immunoblotting and immunofluorescence microscopy. Similar phenomenon was observed in the caspase-3-deficient MCF7 cells upon expressing procaspase-3 transiently. We conclude that a complex mechanism, involving the recognition of the NLSs of both ICAD and CAD, accounts for the constitutive accumulation of CAD/ICAD in the nucleus, where caspase-3-dependent regulation of CAD activity takes place.
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PMID:Determinants of the nuclear localization of the heterodimeric DNA fragmentation factor (ICAD/CAD). 1090 75

In this study, two alternatively spliced forms of the mouse death-associated protein kinase (DAPK) have been identified and their roles in apoptosis examined. The mouse DAPK-alpha sequence is 95% identical to the previously described human DAPK, and it has a kinase domain and calmodulin-binding region closely related to the 130-150 kDa myosin light chain kinases. A 12-residue extension of the carboxyl terminus of DAPK-beta distinguishes it from the human and mouse DAPK-alpha. DAPK phosphorylates at least one substrate in vitro and in vivo, the myosin II regulatory light chain. This phosphorylation occurs preferentially at Ser-19 and is stimulated by calcium and calmodulin. The mRNA encoding DAPK is widely distributed and detected in mouse embryos and most adult tissues, although the expression of the encoded 160-kDa DAPK protein is more restricted. Overexpression of DAPK-alpha, the mouse homolog of human DAPK has a negligible effect on tumor necrosis factor (TNF)-induced apoptosis. Overexpression of DAPK-beta has a strong cytoprotective effect on TNF-treated cells. Biochemical analysis of TNF-treated cell lines expressing mouse DAPK-beta suggests that the cytoprotective effect of DAPK is mediated through both intrinsic and extrinsic apoptotic signaling pathways and results in the inhibition of cytochrome c release from the mitochondria as well as inhibition of caspase-3 and caspase-9 activity. These results suggest that the mouse DAPK-beta is a negative regulator of TNF-induced apoptosis.
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PMID:Identification of a new form of death-associated protein kinase that promotes cell survival. 1148 96

Pancreatic adenocarcinoma continues to be a devastating tumor (28,000 new cases per year in the United States; 10% 2-year survival). Pancreatic adenocarcinoma frequently (90% of the time) overexpresses fibroblast growth factor ligands (FGF-1 and FGF-2) and alternatively spliced high-affinity receptors (FGFR-1beta) (FGFR-1alpha was previously found in normal pancreatic tissue). To study the significance of this observation in vitro, PANC-1 cells were stably transfected via the pMEXneo vector containing FGFR-1alpha (PANC-1alpha) or FGFR-1beta (PANC-1beta) isoforms. Cells were treated with 1 mg/ml of 5-fluorouracil. Cells were evaluated for growth inhibition, apoptosis (propidium iodide staining and flow cytometry, caspase 3 activation) and for Bcl-x(L)/BAX expression (by Western blot analysis). In vivo, 7 x 10(6) cells of each isoform were injected into nude Balb/c mice for xenograft formation (N = 10). Compared to PANC-1beta (9%) in vitro, 5-fluorouracil-induced death was significantly (P < 0.05) increased in PANC-1alpha (20%) at 24 hours. Increased cell death in PANC-1alpha was mediated by activated caspase 3 and was correlated with decreased expression of Bcl-x(L)/BAX. In vivo, PANC-1beta readily demonstrated formation of tumor xenograft at 2 weeks, whereas PANC-1alpha did not form tumors. Alternative splicing of FGFR-1 to the beta isoform appears to correlate with pancreatic adenocarcinoma cell growth in vivo and resistance to chemotherapy. Inhibition of FGFR-1 splicing or overexpression of FGFR-1alpha inhibits pancreatic adenocarcinoma cell growth in vivo and restores cytotoxic responses to chemotherapy, thereby suggesting the basis of rational interventional strategies for this devastating tumor.
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PMID:Ligand activation of alternatively spliced fibroblast growth factor receptor-1 modulates pancreatic adenocarcinoma cell malignancy. 1212 20

Survivin is a relatively unique member of the inhibitor of apoptosis protein (IAP) family in that it contains a single baculovirus IAP repeat (BIR) domain combined with a COOH-terminal alpha-helix coiled-coil domain instead of the more common zinc-binding RING finger. Results from a variety of transformed or continuous mammalian cell lines suggest that, due to the combination of these features, Survivin is capable of regulating both cell proliferation and apoptotic cell death. However, to date there is essentially no information regarding Survivin expression, regulation or function within the ovary, or in any nonmammalian vertebrate species. In the present studies, cDNAs for chicken (ch) Survivin-142 (homologous to human Survivin-142) plus three alternatively spliced variants (ch Survivin-short, -gamma, and -delta) are described, and of these, transcripts for ch Survivin-142 and -short are expressed in granulosa cells from the hen ovary. Highest levels of Survivin mRNA during follicle development occur in mitotically active granulosa cells from undifferentiated, prehierarchal follicles. Cell cycle analysis determined that Survivin mRNA expression is elevated specifically during the G2/M phase of mitosis. Significantly, transient transfection with ch Survivin-142 in primary cultures of hen granulosa cells attenuates taxol- and N-octanoylsphingosine- (C8-ceramide-) induced caspase-3 activity, whereas overexpression of ch Survivin-short (a truncated variant that lacks much of the functional BIR domain plus the entire alpha-helix coil domain) lacks this antiapoptotic activity. Taken together, these data provide evidence for Survivin in granulosa cells acting as a bifunctional protein associated with regulation of the cell cycle and the inhibition of apoptosis.
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PMID:Survivin as a cell cycle-related and antiapoptotic protein in granulosa cells. 1219 53

The stability of the p53 tumor suppressor protein is critically regulated by the Hdm2 and Hdmx proteins. Hdm2 protein levels are auto-regulated by the self-ubiquitination activity of Hdm2 and on the transcriptional level by p53-activated transcription of the hdm2 gene. Little is known about the regulation of Hdmx expression levels, apart from the observation that the Mdmx protein can be cleaved by caspase-3 in a p53-inducible manner. In the functional analysis of two mutant Hdmx proteins, products of two alternatively spliced mRNAs, it was found that Hdmx proteins are targets for ubiquitination by Mdm2. The stability of the Hdmx protein is partly dependent on the presence of its internal acidic domain. Mdm2 appears only to require an intact RING domain to be able to ubiquitinate Hdmx and target it for proteasomal degradation. These findings highlight the intricate functional relationships between p53, Mdm2, and Hdmx.
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PMID:Hdmx protein stability is regulated by the ubiquitin ligase activity of Mdm2. 1287 96

Functional inducible NOS (iNOS) may be involved in the prolonged lifespan of chronic lymphocytic leukemia cells (B-CLL), although the exact mechanisms implicated remain elusive as yet. In this work, we have examined iNOS expression in normal B lymphocytes and B-CLL cells in pro- and antiapoptotic conditions. Our results demonstrate: (1) The existence of a new splice variant characterized by a complete deletion of exon 14 (iNOS 13-16(14del)), which was preferentially detected in normal B lymphocytes and may represent an isoform that could play a role in the regulation of enzyme activity. (2) The existence of another alternatively spliced iNOS mRNA transcript involving a partial deletion of the flavodoxin region (iNOS 13-16(neg)) was correlated to a decreased B-CLL cell viability. The 9-beta-D-arabinofuranosyl-2-fluoradenine or fludarabine (F-ara) treatment induced iNOS 13-16(neg) transcript variants, whereas IL-4 enhanced both the transcription of variants, including these exons (iNOS 13-16(pos)), and the expression of a 122 kDa iNOS protein. These results suggest that in B-CLL, a regulation process involving nitric oxide (.- NO) levels could occur by a post-transcriptional mechanism mediated by soluble factors. Our results also provide an insight into a new complementary proapoptotic action of F-ara in B-CLL by the induction of particular iNOS splice variants, leading to the activation of a caspase-3-dependent apoptotic pathway.
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PMID:Post-transcriptional regulation of inducible nitric oxide synthase in chronic lymphocytic leukemia B cells in pro- and antiapoptotic culture conditions. 1457 28

Alternative RNA splicing is now known to be pervasive throughout the genome and a target of human disease. We evaluated if targeting intronic splicing regulatory sequences with antisense oligonucleotides could be used to correct aberrant exon skipping. As a model, we targeted the intronic silencing sequence (ISS) elements flanking the alternatively spliced alpha-exon of the endogenous fibroblast growth factor receptor 1 (FGFR1) gene, which is aberrantly skipped in human glioblastoma. Antisense morpholino oligonucleotides targeting either upstream or downstream ISS elements increased alpha-exon inclusion from 10% up to 70% in vivo. The effect was dose dependent, sequence specific and reproducible in several human cell lines, but did not necessarily correlate with blocking of protein association in vitro. Simultaneous targeting of the ISS elements had no additive effect, suggesting that splicing regulation occurred through a shared mechanism. Broad applicability of this approach was demonstrated by similar targeting of the ISS elements of the human hnRNPA1 gene. The correction of FGFR1 gene splicing to >90% alpha-exon inclusion in glioblastoma cells had no discernable effect on cell growth in culture, but was associated with an increase in unstimulated caspase-3 and -7 activity. The ability to manipulate endogenously expressed mRNA variants allows exploration of their functional relevance under normal and diseased physiological states.
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PMID:Correction of aberrant FGFR1 alternative RNA splicing through targeting of intronic regulatory elements. 1533 83

The WT1 gene is overexpressed in human primary leukemia and a wide variety of solid cancers. The WT1 gene is alternatively spliced at two sites, yielding four isoforms: 17AA(+)KTS(+), 17AA(+)KTS(-), 17AA(-)KTS(+), and 17AA(-)KTS(-). Here, we showed that 17AA(+)WT1-specific siRNA induced apoptosis in three WT1-expressing leukemia cell lines (K562, HL-60, and Kasumi-1), but not in WT1-non-expressing lymphoma cell line (Daudi). 17AA(+)WT1-specific siRNA activated caspase-3 and -9 in the intrinsic apoptosis pathway but not caspase-8 in the extrinsic one. On the other hand, 17AA(-)WT1-specific siRNA did not induce apoptosis in the three WT1-expressing cell lines. The apoptosis was associated with activation of proapoptotic Bax, which was activated upstream of the mitochondria. Constitutive expression of 17AA(+)WT1 isoforms inhibited apoptosis of K562 leukemia cells induced by apoptosis-inducing agents, etoposide and doxorubicin, through the protection of mitochondrial membrane damages, and DNA-binding zinc-finger region of 17AA(+)WT1 isoform was essential for the antiapoptotic functions. We further studied the gene(s) whose expression was altered by the expression of 17AA(+)WT1 isoforms and showed that the expression of proapoptotic Bak was decreased by the expression of 17AA(+)KTS(-)WT1 isoform. Taken together, these results indicated that 17AA(+)WT1 isoforms played antiapoptotic roles at some points upstream of the mitochondria in the intrinsic apoptosis pathway.
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PMID:Antiapoptotic function of 17AA(+)WT1 (Wilms' tumor gene) isoforms on the intrinsic apoptosis pathway. 1651 14

NT2 cells are a human teratocarcinoma cell line that, upon treatment with retinoic acid (RA), begin differentiating into a neuronal phenotype. The transformation of undifferentiated NT2 cells into hNT neurons presents an opportunity to investigate the mechanisms involved in neurogenesis because a key component is cell apoptosis, which is essential for building neural networks. Protein kinase Cdelta (PKCdelta) plays an important role as a mediator of cellular apoptosis in response to various stimuli. PKCdelta (deltaI) is proteolytically cleaved at its hinge region (V3) by caspase 3 and the catalytic fragment is sufficient to induce apoptosis in various cell types. Mouse PKCdeltaII is rendered caspase resistant due to an insertion of 78 bp within the caspase recognition site in its V3 domain. No functional role has been attributed to these alternatively spliced variants of PKCdelta. We sought to find a correlation between the onset of apoptosis, neurogenesis, and the expression of PKCdelta isoforms. Our results indicate that RA regulates the expression of PKCdelta alternative splicing variants in NT2 cells. Further, overexpression of PKCdeltaI promotes apoptosis while PKCdeltaII overexpression shields the cells from apoptosis. This is the first report to attribute physiological function to PKCdeltaI and -deltaII isoforms. Next we demonstrated that mouse embryonic stem cells differentiate in vitro into dopaminergic neurons upon stimulation with RA and ciliary neurotrophic factor. These cells showed a simultaneous increase in tyrosine hydroxylase and PKCdeltaII expression. We suggest that the molecular mechanisms regulating differentiation and apoptosis could be understood by alternative expression of PKCdelta isoforms.
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PMID:PKCdelta alternatively spliced isoforms modulate cellular apoptosis in retinoic acid-induced differentiation of human NT2 cells and mouse embryonic stem cells. 1701 22


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