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)

Apoptotic cell shrinkage, an early hallmark of apoptosis, is regulated by K+ efflux and K+ channel activity. Inhibited apoptosis and downregulated K+ channels in pulmonary artery smooth muscle cells (PASMC) have been implicated in development of pulmonary vascular medial hypertrophy and pulmonary hypertension. The objective of this study was to test the hypothesis that overexpression of KCNA5, which encodes a delayed-rectifier voltage-gated K+ (Kv) channel, increases K+ currents and enhances apoptosis. Transient transfection of KCNA5 caused 25- to 34-fold increase in KCNA5 channel protein level and 24- to 29-fold increase in Kv channel current (I(K(V))) at +60 mV in COS-7 and rat PASMC, respectively. In KCNA5-transfected COS-7 cells, staurosporine (ST)-mediated increases in caspase-3 activity and the percentage of cells undergoing apoptosis were both enhanced, whereas basal apoptosis (without ST stimulation) was unchanged compared with cells transfected with an empty vector. In rat PASMC, however, transfection of KCNA5 alone caused marked increase in basal apoptosis, in addition to enhancing ST-mediated apoptosis. Furthermore, ST-induced apoptotic cell shrinkage was significantly accelerated in COS-7 cells and rat PASMC transfected with KCNA5, and blockade of KCNA5 channels with 4-aminopyridine (4-AP) reduced K+ currents through KCNA5 channels and inhibited ST-induced apoptosis in KCNA5-transfected COS-7 cells. Overexpression of the human KCNA5 gene increases K+ currents (i.e., K+ efflux or loss), accelerates apoptotic volume decrease (AVD), increases caspase-3 activity, and induces apoptosis. Induction of apoptosis in PASMC by KCNA5 gene transfer may serve as an important strategy for preventing the progression of pulmonary vascular wall thickening and for treating patients with idiopathic pulmonary arterial hypertension (IPAH).
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PMID:Overexpression of human KCNA5 increases IK V and enhances apoptosis. 1514 Jul 47

The open reading frame EP153R of African swine fever virus (ASFV) encodes a nonessential protein that has been involved in the hemadsorption process induced in virus-infected cells. By the use of a virus deletion mutant lacking the EP153R gene, we have detected, in several virus-sensitive cells, increased levels of caspase-3 and cell death as compared with those obtained after infection with the parental BA71V strain. Both transient and stable expression of the EP153R gene in Vero or COS cells resulted in a partial protection of the transfected lines from the apoptosis induced in response to virus infection or external stimuli. The presence of gene EP153R resulted in a reduction of the transactivating activity of the cellular protein p53 in Vero cell cultures in which apoptosis was induced by virus infection or staurosporine treatment. This is to our knowledge the first description of a viral C-type lectin with anti-apoptotic properties.
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PMID:The C-type lectin homologue gene (EP153R) of African swine fever virus inhibits apoptosis both in virus infection and in heterologous expression. 1526 4

Human parvovirus B19 has been found in various tissues in addition to erythroid lineage cells, and non-structural protein (NS1) is reported to induce cytotoxicity and apoptosis in erythroid lineage cells, but the mechanism in non-permissive cells is still unclear. To address this issue, we have constructed the NS1 gene in a cytomegalovirus episomal vector, pEGFP-C1 and transfected it into monkey epithelial cells, COS-7. EGFP-NS1 expression in transfected cells was monitored and assessed by fluorescence microscopy, RT-PCR and Western blot. The flow cytometric analysis showed that the NS1-transfected cells were arrested at G1 phase by paclitaxel treatment and there was increased apoptosis. The expression of p53, an important molecule in apoptosis and cell cycle regulation, and its downstream cell cycle kinase inhibitors p16(INK4) and p21(WAF1/CIP1) were up-regulated in the NS1-transfected cells. Also, increased expression of the pro-apoptotic Bcl-2 members Bax, Bad and activation of caspase 3 and caspase 9, but not the activation of caspase 8 or Fas were detected in the NS1-transfected cells. p53-induced Bax expression and subsequent activation of caspase 9 is probably the apoptotic pathway in NS1-transfected cells since activation of the caspase 9 was suppressed by the p53 inhibitor and apoptosis was significantly inhibited by the caspase 9 inhibitor. Our results suggest that the cell death of the NS1-transfected cells is associated with mitochondria related apoptosis. These findings might provide alternative information for further study and characterization of B19 NS1 protein in B19 non-permissive cells.
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PMID:Human parvovirus B19 non-structural protein (NS1) induces apoptosis through mitochondria cell death pathway in COS-7 cells. 1537 Jun 68

PDE4A11 is a novel cAMP-specific phosphodiesterase that is conserved in humans, mouse, rat, pig, and bat. Exon-1(4A11) encodes its unique, 81 amino acid N-terminal region. Reverse-transcriptase polymerase chain reaction performed across the splice junction, plus identification of expressed sequence tags, identifies PDE4A11 as a long isoform possessing UCR1 and UCR2 regulatory domains. Transcript analysis shows that PDE4A11 is widely expressed compared with PDE4A10 and PDE4A4B long isoforms. Truncation analysis identifies a putative promoter in a 250-base pair region located immediately upstream of the start site in Exon-1(4A11). Recombinant PDE4A11, expressed in COS-7 cells, is a 126-kDa protein localized predominantly around the nucleus and in membrane ruffles. PDE4A11 exhibits a K(m) for cAMP hydrolysis of 4 microM, with relative V(max) similar to that of PDE4A10 and PDE4A4B. PDE4A11 is dose-dependently inhibited by rolipram, 4-[(3-butoxy-4-methoxyphenyl)-methyl]-2-imidazolidinone (Ro 20-1724), cilomilast, roflumilast, and denbufylline, with IC(50) values of 0.7, 0.9, 0.03, 0.004, and 0.3 microM, respectively. Soluble and particulate PDE4A11 exhibit distinct rates of thermal inactivation (55 degrees C; T((0.5)) = 2.5 and 4.4 min, respectively). Elevating cAMP levels in COS-7 cells activates PDE4A11 concomitant with its phosphorylation at Ser119 by protein kinase A (PKA). PDE4A11 differs from PDE4A4 in sensitivity to cleavage by caspase-3, interaction with LYN SH3 domain, redistribution upon long-term rolipram challenge, and sensitivity to certain PDE4 inhibitors. PDE4A11, PDE4A10, and PDE4A4 all can interact with betaarrestin. PDE4A11 is a novel, widely expressed long isoform that is activated by PKA phosphorylation and shows a distinct intracellular localization, indicating that it may contribute to compartmentalized cAMP signaling in cells in which it is expressed.
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PMID:Identification and characterization of PDE4A11, a novel, widely expressed long isoform encoded by the human PDE4A cAMP phosphodiesterase gene. 1573 10

Many human proteins have homopolymeric amino acid (HPAA) tracts, although the physiological significance or cellular effects of their presence is poorly understood. We previously reported that 20 kinds of HPAAs show characteristic intracellular localization and that among those, hydrophobic HPAAs aggregate strongly and form high molecular weight proteins when expressed in cultured cells. In this study, we investigated the cytotoxicity of 20 kinds of HPAAs. HPAA tracts of approximately 30 residues fused to the C-terminus of YFP were expressed in COS-7 cells. Cells expressing homopolymeric-Cys, -Ile, -Leu, and -Val showed low viability in Trypan Blue assay. Caspase-3 activity, which is usually upregulated in dying cells, was determined by measuring the cleavage of the peptide substrate Ac-DEVD-MCA and by detecting the cleaved active form of the caspase-3 by Western blotting. The activity of caspase-3 was drastically elevated in cells expressing those HPAAs which showed low viability in Trypan Blue assay. Interestingly, it was found that there is a correlation between the hydrophobicity of a single amino acid and the cytotoxicity of the corresponding HPAA as a homopolymer. These results indicate that the hydrophobicity of HPAAs may cause cytotoxicity.
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PMID:Comparative analysis of the cytotoxicity of homopolymeric amino acids. 1576 94

Recent evidence has implicated the protein phosphatase PP5 in a variety of signaling pathways. Whereas several proteins have been identified that interact with PP5 and regulate its activity, a possibility of its regulation by second messengers remains speculative. Activation of PP5 in vitro by polyunsaturated fatty acids (e.g. arachidonic acid) and fatty acyl-CoA esters (e.g. arachidonoyl-CoA) has been reported. We report here that PP5 is strongly inhibited by micromolar concentrations of a natural polyamine spermine. This inhibition was observed both in assays with a low molecular weight substrate p-nitrophenyl phosphate as well as phosphocasein and apoptosis signal-regulating kinase 1 (ASK1), thought to be a physiological substrate of PP5. Furthermore, a decrease in polyamine levels in COS-7 cells induced by alpha-difluoromethylornithine (DFMO), an inhibitor of ornithine decarboxylase, led to accelerated dephosphorylation of oxidative stress-activated ASK1. This effect was suppressed by okadaic acid and by siRNA-mediated PP5 depletion, indicating that the effect of polyamine levels on ASK1 dephosphorylation was mediated by PP5. In line with the decreased ASK1 activation, polyamine depletion in COS-7 cells abrogated oxidative stress-induced activation of caspase-3, which executes ASK1-induced apoptosis, as well as caspase-3 activation induced by ASK1 overexpression, but had no effect on basal caspase-3 activity. These results implicate polyamines, emerging intracellular signaling molecules, as potential physiological regulators of PP5. Our findings also suggest a novel mechanism of the anti-apoptotic action of a decrease in polyamine levels via de-inhibition of PP5 and accelerated dephosphorylation and deactivation of ASK1.
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PMID:Regulation of apoptosis signal-regulating kinase 1 (ASK1) by polyamine levels via protein phosphatase 5. 1589 Jun 60

The calmodulin-activated transporter hPMCA4 (human plasma membrane Ca2+-ATPase isoform 4) is a target for cleavage by caspase-3 during apoptosis. We have demonstrated that caspase-3 generates a 120 kDa fragment of this pump which lacks the complete autoinhibitory sequence [Paszty, Verma, Padanyi, Filoteo, Penniston and Enyedi (2002) J. Biol. Chem. 277, 6822-6829]. In the present study we analysed further the characteristics of the fragment of hPMCA4b produced by caspase-3. We did this by overexpressing the caspase-3 cleavage product of hPMCA4b in COS-7 and MDCKII (Madin-Darby canine kidney II) cells. This technique made it possible to clearly define the properties of this fragment, and we showed that it is constitutively active, as it forms a phosphoenzyme intermediate and has high Ca2+ transport activity in the absence of calmodulin. When this fragment of hPMCA4b was stably expressed in MDCKII cell clones, it was targeted without degradation to the basolateral plasma membrane. In summary, our studies emphasize that the caspase-3 cleavage product of hPMCA4b is constitutively active, and that the C-terminus is not required for proper targeting of hPMCA4b to the plasma membrane. Also, for the first time, we have generated cell clones that stably express a constitutively active PMCA.
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PMID:The caspase-3 cleavage product of the plasma membrane Ca2+-ATPase 4b is activated and appropriately targeted. 1608 Jul 82

Fluorescence resonance energy transfer (FRET)-based protein biosensors allow the spatial and temporal imaging of signaling events in living cells. However, the simultaneous correlation of multiple events of a signaling pathway is hindered by the spectral cross-talk between fluorescent proteins. Here, we show, for signaling pathways that progress synchronously, multiple events can be correlated by using co-cultures expressing different FRET-based protein biosensors. As a demonstration, we investigated the simultaneous caspase-3 and Ca2+ signaling events involved in cell death of COS-7 cells induced by 10 mM H2O2. Interestingly, this H2O2 stimulus induced synchronous caspase-3 activation and Ca2+ signaling. In parallel to caspase-3 activation, cytosolic Ca2+ concentration, [Ca2+]c, gradually rises to its peak and then slowly drops. As cell shrinkage and rounding ensues, [Ca2+]c again gradually rises to its peak and then reaches a plateau. These observations reveal the relative timing and location of these signaling events in cell death induced by this stimulus of H2O2. Finally, our approach offers an exciting opportunity for spatial and temporal imaging of multiple events in a signaling pathway in living cells.
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PMID:Using co-cultures expressing fluorescence resonance energy transfer based protein biosensors to simultaneously image caspase-3 and Ca2+ signaling. 1615 67

Nitric oxide (NO) may block apoptosis by inhibiting caspases via S-nitrosylation of cysteines. Here, we investigated whether effector caspases might cleave and thereby inhibit endothelial nitric oxide synthase (eNOS). Exposure of eNOS-transfected COS-7 cells and bovine aortic endothelial cells to staurosporine resulted in significant loss of 135-kDa eNOS protein and activity, and appearance of a 60-kDa eNOS fragment; effects were inhibited by the general caspase inhibitor, benzyloxycarbonyl-Val-Ala-Asp[OMe]-fluoromethyl ketone (zVAD-fmk). In eNOS-transfected COS-7 cells, staurosporine-induced activation of caspase-3 and poly(ADP-ribose) polymerase (PARP) cleavage coincided with increased eNOS degradation and decreased activity. Loss of eNOS activity was greater than the degree of proteolysis. Incubation of immunoprecipitated eNOS with caspase-3, caspase-6 or caspase-7 resulted in eNOS cleavage. Staurosporine, a general protein kinase inhibitor, also reduced phosphorylation and decreased calmodulin binding, an effect that may explain the reduction in activity. eNOS, therefore, is both an inhibitor of apoptosis and a target of apoptosis-associated proteolysis.
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PMID:Effect of staurosporine-induced apoptosis on endothelial nitric oxide synthase in transfected COS-7 cells and primary endothelial cells. 1619 40

Klotho-mutated mice manifest multiple age-related disorders that are observed in humans. A recent study suggested that Klotho protein might function as an anti-aging hormone in mammals. Because it has been reported that apoptosis and senescence in vascular endothelial cells are closely related to the progression of atherosclerosis, we investigated Klotho's ability to interfere with apoptosis and cellular senescence in human umbilical vascular endothelial cells (HUVEC). Klotho overexpression decreased H(2)O(2)-induced apoptosis in COS-1 cells and Jurkat cells. Klotho protein also reduced H(2)O(2)- and etoposide-induced apoptosis in HUVEC. Caspase-3 and caspase-9 activity was lower in Klotho-treated HUVEC than in control cells. Senescence-associated beta-gal staining showed that Klotho protein interferes with H(2)O(2)-induced premature cellular senescence. The expression of p53 and p21 was lower in Klotho-treated cells. Our study suggests that Klotho acts as a humoral factor to reduce H(2)O(2)-induced apoptosis and cellular senescence in vascular cells.
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PMID:Anti-apoptotic and anti-senescence effects of Klotho on vascular endothelial cells. 1632 73


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