Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.22.62 (caspase-9)
 7,507 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Caspases are universal effectors of apoptosis. The mitochondrial and death receptor pathways activate distinct apical caspases (caspase-9 and -8, respectively) that converge on the proteolytic activation of the downstream executioner caspase-3. Caspase-9 and -8 cleave procaspase-3 to produce a p24 processing intermediate (composed of its prodomain and large subunit), which then undergoes autoproteolytic cleavage to remove the prodomain from the active protease. Recently, several heat shock proteins have been shown to selectively inhibit the mitochondrial apoptotic pathway by disrupting the activation of caspase-9 downstream of cytochrome c release. We report here that the small heat shock protein alphaB-crystallin inhibits both the mitochondrial and death receptor pathways. In S-100 cytosolic extracts treated with cytochrome c/dATP or caspase-8, alphaB-crystallin inhibits the autoproteolytic maturation of the p24 partially processed caspase-3 intermediate. In contrast, neither the closely related small heat shock protein family member Hsp27 nor Hsp70 inhibited the maturation of the p24 intermediate. We also demonstrate that alphaB-crystallin co-immunoprecipitates with the p24 partially processed caspase-3 in vivo. Taken together, our results demonstrate that alphaB-crystallin is a novel negative regulator of apoptosis that acts distally in the conserved cell death machinery by inhibiting the autocatalytic maturation of caspase-3.
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PMID:The small heat shock protein alpha B-crystallin negatively regulates cytochrome c- and caspase-8-dependent activation of caspase-3 by inhibiting its autoproteolytic maturation. 1127 39

We determined whether the cytoprotective heat shock protein HSP72 protects against the injurious effects of Clostridium difficile toxin A (TxA) on intestinal epithelial cells. Colonic epithelial Caco-2/bbe (C2) cells were stably transfected with HSP72 antisense (C2AS) or vector only (C2VC), resulting in low and high HSP72 expression, respectively. Measurements of epithelial barrier integrity, mitochondrial function, and apoptosis activation were assessed after TxA exposure. HSP72 and RhoA interactions were evaluated with immunoprecipitations. In C2AS cells, TxA was associated with a greater decrease in transepithelial resistance (TER), an increase in [(3)H]mannitol flux, and increased dissociation of perijunctional actin. Although HSP72 binds RhoA, it failed to prevent RhoA glucosylation. TxA caused a more rapid decrease in ATP, release of cytochrome c, and activation of caspase-9 in C2AS cells. To determine whether ATP depletion decreases TER, we treated cells with antimycin A, which caused a decline in TER. We conclude that HSP72 may protect intestinal epithelial cells from TxA-mediated damage through several mechanisms, including actin stabilization, mitochondrial protection, and inhibition of apoptosis activation, but not by prevention of RhoA glucosylation.
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PMID:Protective role of HSP72 against Clostridium difficile toxin A-induced intestinal epithelial cell dysfunction. 1249 Apr 34

17-allylamino-demethoxy geldanamycin (17-AAG) inhibits the chaperone function of heat shock protein-90 (Hsp-90) and promotes the proteasomal degradation of its misfolded client proteins. Here, we demonstrate that treatment of the human acute myeloid leukemia HL-60 cells with 17-AAG attenuates the intracellular levels of a number of Hsp-90 client proteins, including Akt, c-Raf-1, and c-Src. Also, 17-AAG induced the mitochondrial release and cytosolic accumulation of cytochrome c (cyt c) and second mitochondria-derived activator of caspases (Smac)/DIABLO, resulting in the activation of caspase-9 and caspase-3 and apoptosis. Treatment with 17-AAG triggered the B-cell lymphoma-2 (Bcl-2)-associated X protein (Bax) conformational change associated with apoptosis, while Bax-deficient cells were resistant to 17-AAG-induced apoptosis. In addition, in HL-60/Bcl-2 and HL-60/Bcl-xL cells, which ectopically express Bcl-2 and Bcl-xL respectively, 17-AAG-induced Bax conformational change, cytosolic accumulation of cyt c and Smac/DIABLO, and apoptosis were markedly inhibited. Although the rate of 17-AAG-mediated decline in Akt, c-Raf-1, and c-Src levels was blunted, the total decline was not compromised in HL-60/Bcl-2 and HL-60/Bcl-xL cells. Cotreatment with HA14-1, a nonpeptidic ligand that can bind and inhibit the antiapoptotic activity of Bcl-2, significantly overcame the resistance to 17-AAG-induced apoptosis in HL-60/Bcl-2 cells. Together, these findings indicate that although 17-AAG treatment causes the levels of a number of survival-signaling protein kinases to decline, the downstream engagement of the mitochondrial pathway of apoptosis is regulated by the activity of the Bcl-2 family of proteins. Also, neutralizing the antiapoptotic effect of Bcl-2 would further enhance the antileukemia activity of 17-AAG.
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PMID:Regulation of 17-AAG-induced apoptosis: role of Bcl-2, Bcl-XL, and Bax downstream of 17-AAG-mediated down-regulation of Akt, Raf-1, and Src kinases. 1262 37

The effect of exercise on apoptosis in postmitotic tissues is not known. In this study, we investigated the effect of regular moderate physical activity (i.e., exercise training) on the extent of apoptosis in rat skeletal and cardiac muscles. Adult Sprague Dawley rats were trained (TR) 5 days weekly for 8 wk on treadmill. Sedentary rats served as controls (CON). An ELISA was used to detect mono- and oligonucleosome fragmentation as an indicator of apoptosis. Bcl-2, Bax, Apaf-1, AIF, cleaved PARP, cleaved caspase-3, cleaved/active caspase-9, heat shock protein (HSP)70, Cu/Zn-SOD, and Mn-SOD protein levels were determined by Western analyses. Bcl-2 and Bax transcript contents were estimated by RT-PCR. A spectrofluorometric assay was used to determine caspase-3 activity. DNA fragmentation in ventricles of the TR group decreased by 15% whereas that in soleus of the TR group tended to decrease (P=0.058) when compared with CON group. Protein contents of Bcl-2, HSP70, and Mn-SOD increased in both soleus and ventricle muscles of TR animals when compared with CON animals. Apaf-1 protein content in the soleus of TR animals was lower than that of CON animals. Bcl-2 mRNA levels increased in both ventricle and soleus muscles of TR animals, and Bax mRNA levels decreased in the soleus of TR animals when compared with CON animals. Furthermore, HSP70 protein content was negatively correlated to Bax mRNA content and was positively correlated to Bcl-2 protein and mRNA contents. Mn-SOD protein content was negatively correlated to the apoptotic index, and caspase-3 activity and was positively correlated to Bcl-2 transcript content and HSP70 protein content. These data suggest that exercise training attenuates the extent of apoptosis in cardiac and skeletal muscles.
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PMID:Apoptotic adaptations from exercise training in skeletal and cardiac muscles. 1513 82

Bcr-Abl-expressing primary or cultured leukemia cells display high levels of the antiapoptotic heat shock protein (hsp) 70 and are resistant to cytarabine (Ara-C), etoposide, or Apo-2L/TRAIL (TNF-related apoptosis-inducing ligand)-induced apoptosis. Conversely, a stable expression of the cDNA of hsp70 in the reverse orientation attenuated not only hsp70 but also signal transducers and activators of transcription 5 (STAT5) and Bcl-x(L) levels. This increased apoptosis induced by cytarabine, etoposide, or Apo-2L/TRAIL. Ectopic expression of hsp70 in HL-60 cells (HL-60/hsp70) inhibited Ara-C and etoposide-induced Bax conformation change and translocation to the mitochondria; attenuated the accumulation of cytochrome c, Smac, and Omi/HtrA2 in the cytosol; and inhibited the processing and activity of caspase-9 and caspase-3. Hsp70 was bound to death receptors 4 and 5 (DR4 and DR5) and inhibited Apo-2L/TRAIL-induced assembly and activity of the death-inducing signaling complex (DISC). HL-60/hsp70 cells exhibited increased levels and DNA binding activity of STAT5, which was associated with high levels of Pim-2 and Bcl-x(L) and resistance to apoptosis. Expression of the dominant negative (DN) STAT5 resensitized HL-60/hsp70 cells to cytarabine, etoposide, and Apo-2L/TRAIL-induced apoptosis. Collectively, these findings suggest that hsp70 inhibits apoptosis upstream and downstream of the mitochondria and is a promising therapeutic target for reversing drug-resistance in chronic myeloid leukemia-blast crisis and acute myeloid leukemia cells.
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PMID:Mechanistic role of heat shock protein 70 in Bcr-Abl-mediated resistance to apoptosis in human acute leukemia cells. 1538 81

Previously, we showed that gene expression of the rheumatoid arthritis-related antigen RA-A47, which is identical to human heat shock protein (HSP)47, was downregulated in chondrocytes by inflammatory cytokines such as TNFalpha. Associated with this phenomenon, RA-A47 appeared on the cell surface concomitant with upregulation of metabolic factors related to cartilage destruction. The upregulation of the metabolic factors could be achieved by downregulation of RA-A47 expression with ra-a47-specific anti-sense oligonucleotide. Here, we show that the enhanced surface expression of RA-A47 on a chondrocytic cell line, HCS-2/8 was also a direct result of RA-A47 downregulation by ra-a47 anti-sense oligonucleotide, independent of the cytokine effects. Moreover, cell-surface expression of CD9, a beta1 integrin-associated transmembrane protein that is involved in cell adhesion and cell motility events, was enhanced in the ra-a47 anti-sense oligonucleotide-treated cells. The CD9 was colocalized with RA-A47 on the cell surface, where it may have affected integrin signaling. Furthermore, Annexin-V binding to the cell surface and the level of a number of apoptosis-related genes including caspase-9 were increased after ra-a47 anti-sense oligonucleotide treatment, suggesting that enhanced surface expression of RA-A47 and CD9 may be initiating apoptosis. Differential screening using a cDNA gene array showed induction of metallothionein-III and chemokine receptor CXCR4 and of factors of the Notch signaling pathway by the anti-sense treatment, but not by TNFalpha. Thus, here we show for the first time an alternative mechanism of inducing apoptosis by downregulating molecular chaperones, independent of the action of TNFalpha. The surface-exposed RA-A47 may induce autoantibodies and inflammatory reactions in autoimmune disease situations such as rheumatoid arthritis.
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PMID:Downregulation of rheumatoid arthritis-related antigen RA-A47 (HSP47/colligin-2) in chondrocytic cell lines induces apoptosis and cell-surface expression of RA-A47 in association with CD9. 1538 25

We have previously observed that near-infrared (IR) pre-irradiation protects normal human dermal fibroblasts from ultraviolet (UV) cytotoxicity in vitro. Here, we show that IR pre-irradiation of human fibroblasts inhibited UVB activation of caspase-9 and -3, leading us to study early events in the mitochondrial apoptotic pathway after IR irradiation. IR irradiation led to a partial release of cytochrome c and Smac/Diablo but not apoptosis-inducing factor (AIF). This was accompanied by a slight but transient decrease in the mitochondrial membrane potential (Deltapsim) and by the insertion of Bax into mitochondrial membrane. Early apoptotic events in the mitochondrial pathway thus occurred after IR irradiation despite a lack of caspase-9 and -3 activation. This could be explained by the induction by IR of the expression of heat shock protein Hsp27, which is known to prevent apoptosome assembly. Furthermore, the balance between pro-apoptotic (i.e., Bax) and anti-apoptotic (i.e., Bcl-2 or Bcl-xL) proteins, which was rather pro-apoptotic after IR exposure, became anti-apoptotic 24 h later, suggesting a protective effect. Together, these actions could also contribute to prepare the cell to resist UVB-triggered apoptosis. Finally, isolated rat liver mitochondria-released cytochrome c in response to IR, demonstrating that mitochondria were a primary target of IR radiation.
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PMID:Infrared radiation affects the mitochondrial pathway of apoptosis in human fibroblasts. 1548 67

Viruses have evolved different strategies to interfere with apoptotic pathways in order to halt cellular responses to infection. One previous study showed that transient transfection of bovine herpesvirus type-1 (BHV-1) UL14 protein is efficient in protecting Madin Darby kidney (MDBK) and human chronic myelogenous leukemia (K562) cells from sorbitol-induced apoptosis. This protein corresponds to a putative protein of BHV-1, which shares aminoacid sequence with a part of the peptide-binding domain conserved in human heat shock protein (HSP70) family. The pBK-CMV-UL14 plasmid transfected MDBK cells treated with sorbitol did not show caspase-3 and caspase-9 activation with respect to non-transfected MDBK cells (UL14 negative). Furthermore, we report that the expression of the full length sequence of BHV-1 UL14 is evident after 7 h of infection of BHV-1 on MDBK cells which were then treated with sorbitol. These results indicate that UL14 gene product has important implications to enhance cell survival in response to apoptotic stimuli.
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PMID:Antiapoptotic activity of bovine herpesvirus type-1 (BHV-1) UL14 protein. 1740 88

We observed that photodynamic therapy (PDT) induces the expression and phosphorylation of the inhibitor of apoptosis (IAP) protein survivin in murine and human cancer cells and tumors. Survivin inhibits caspase-9, blocks apoptosis, and is associated with resistance to chemotherapy and radiation. Survivin is a client protein for the 90-kDa heat shock protein (Hsp-90), and the binding of survivin to Hsp-90 assists in the maturation, proper folding, assembly, and transport of this IAP protein. A derivative of the antibiotic geldanamycin, 17-allylamino-17-demethoxygeldanamycin (17-AAG), interferes with proper binding of client proteins, such as survivin, to Hsp-90 and leads to misfolding of client proteins, ubiquination, and proteasome degradation. We hypothesized that PDT efficacy may be reduced by treatment-mediated expression and phosphorylation of survivin, and therefore, targeting the survivin pathway could increase PDT responsiveness. To address this hypothesis, we examined cellular and molecular responses following exposure to PDT, 17-AAG, and the combination of PDT plus 17-AAG in human BT-474 breast cancer cells using Photofrin and NPe6 as photosensitizers. Cells treated with the combination of PDT and 17-AAG exhibited decreased expression of the Hsp-90 client proteins phosphorylated survivin, phosphorylated Akt, and Bcl-2. The decreased expression of these client proteins was accompanied by higher apoptotic indexes and increased cytotoxicity. To confirm a specific role for survivin in modulating PDT, we used a human melanoma cell line, YUSAC2/T34A-C4, stably transfected with an inducible dominant-negative survivin gene under the control of a tetracycline-regulated (tet-off) promoter. PDT treatment of melanoma cells expressing the dominant-negative survivin resulted in increased cleavage of the caspase substrate poly(ADP-ribose) polymerase, apoptosis, and cytotoxicity when compared with results following PDT of the same melanoma cell line expressing wild-type survivin. These results show for the first time that targeting survivin and possibly other Hsp-90 client proteins improves in vitro PDT responsiveness and suggest that manipulation of the antiapoptotic pathway maintained by survivin may enhance PDT-mediated cancer therapy.
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PMID:Survivin, a member of the inhibitor of apoptosis family, is induced by photodynamic therapy and is a target for improving treatment response. 1751 Apr 30

Oncolytic measles virus strains have activity against multiple tumor types and are currently in phase I clinical testing. Induction of the heat shock protein 70 (HSP70) constitutes one of the earliest changes in cellular gene expression following infection with RNA viruses including measles virus, and HSP70 upregulation induced by heat shock has been shown to result in increased measles virus cytotoxicity. HSP90 inhibitors such as geldanamycin (GA) or 17-allylaminogeldanamycin result in pharmacologic upregulation of HSP70 and they are currently in clinical testing as cancer therapeutics. We therefore investigated the hypothesis that heat shock protein inhibitors could augment the measles virus-induced cytopathic effect. We tested the combination of a measles virus derivative expressing soluble human carcinoembryonic antigen (MV-CEA) and GA in MDA-MB-231 (breast), SKOV3.IP (ovarian) and TE671 (rhabdomyosarcoma) cancer cell lines. Optimal synergy was accomplished when GA treatment was initiated 6-24 h following MV infection. Western immunoblotting confirmed HSP70 upregulation in combination-treated cells. Combination treatment resulted in statistically significant increase in syncytia formation as compared to MV-CEA infection alone. Clonogenic assays demonstrated significant decrease in tumor colony formation in MV-CEA/GA combination-treated cells. In addition there was increase in apoptosis by 4,6-diamidino-2-phenylindole staining. Western immunoblotting for caspase-9, caspase-8, caspase-3 and poly(ADP-ribose) polymerase (PARP) demonstrated increase in cleaved caspase-8 and PARP. The pan-caspase inhibitor Z-VAD-FMK and caspase-8 inhibitor Z-IETD-FMK, but not the caspase-9 inhibitor Z-IEHD-FMK, protected tumor cells from MV-CEA/GA-induced PARP activation, indicating that apoptosis in combination-treated cells occurs mainly via the extrinsic caspase pathway. Treatment of normal cells, such as normal human fibroblasts, however, with the MV-CEA/GA combination, did not result in cytopathic effect, indicating that GA did not alter the MV-CEA specificity for tumor cells. One-step viral growth curves, western immunoblotting for MV-N protein expression, QRT-PCR quantitation of MV-genome copy number and CEA levels showed comparable proliferation of MV-CEA in GA-treated vs -untreated tumor cells. Rho activation assays and western blot for total RhoA, a GTPase associated with the actin cytoskeleton, demonstrated decrease in RhoA activation in combination-treated cells, a change previously shown to be associated with increase in paramyxovirus-induced cell-cell fusion. The enhanced cytopathic effect resulting from measles virus/GA combination supports the translational potential of this approach in the treatment of cancer.
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PMID:Heat shock protein inhibitors increase the efficacy of measles virotherapy. 1835 18


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