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: EC:2.7.10.1 (ERK)
95,504 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have demonstrated that ginsenoside Rh2 (G-Rh2), a ginseng saponin with a dammarane skeleton, induces apoptosis of human hepatoma SK-HEP-1 cells as evidenced by analyses of DNA fragmentation, flow cytometry and changes in cell morphology. Ac-YVAD-CMK or Ac-DEVD-CHO effectively prevented G-Rh2-induced DNA fragmentation, indicating the involvement of caspase-like proteases in the process of apoptosis. In addition, G-Rh2 induced the processing of caspase-3 to an active form, p17. In stable Bcl-2 transfectants, G-Rh2 also induced DNA fragmentation, while staurosporine-induced DNA fragmentation was totally blocked. As it did in wild-type cells, G-Rh2 induced the proteolytic activation of caspase-3 protease and subsequent cleavage of PARP in the bcl-2 transfectants. In summary, G-Rh2 contains an apoptotic inducing activity in SK-HEP-1 cells which functions via Bcl-2-insensitive activation of caspase-3, followed by proteolytic cleavage of PARP.
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PMID:Activation of caspase-3 protease via a Bcl-2-insensitive pathway during the process of ginsenoside Rh2-induced apoptosis. 945 77

Focal adhesion kinase (Fak) is a non-receptor protein-tyrosine kinase that stimulates cell spreading and motility by promoting the formation of contact sites between the cell and the extracellular matrix (focal adhesions). It suppresses apoptosis by transducing survival signals that emanate from focal adhesions via the clustering of transmembrane integrins by components of the extracellular matrix. We demonstrate that Fak is cleaved by caspases at two distinct sites during apoptosis. The sites were mapped to DQTD772, which was preferentially cleaved by caspase-3, and VSWD704, which was preferentially cleaved by caspase-6 and cytotoxic T lymphocyte-derived granzyme B. The cleavage of Fak during apoptosis separates the tyrosine kinase domain from the focal adhesion targeting (FAT) domain. The carboxyl-terminal fragments that are generated suppress phosphorylation of endogenous Fak and thus resemble a natural variant of Fak, FRNK, that inhibits Fak activity by preventing the localization of Fak to focal adhesions. The cleavage of Fak by caspases may thus play an important role in the execution of the suicide program by disabling the anti-apoptotic function of Fak. Interestingly, rodent Fak lacks an optimal caspase-3 consensus cleavage site although it is cleaved in murine cells undergoing apoptosis at an upstream site. This appears to be the first example of a caspase substrate where the cleavage sites are not conserved between species.
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PMID:Caspases cleave focal adhesion kinase during apoptosis to generate a FRNK-like polypeptide. 964 76

We report here that p21WAF1/CIP1, an inhibitor of cyclin kinases, underwent proteolytic processing into a smaller fragment, p14, in the early stage of apoptosis in SK-HEP-1 cells. Apoptosis was induced by either staurosporine or ginsenoside Rh2, a ginseng saponin with a dammarane skeleton. Proteolytic processing was the result of caspase-3 activity, which accompanied the early changes in cell morphology and DNA fragmentation. p21WAF1/CIP1 translated in vitro was cleaved into a p14 fragment when incubated with cell extracts obtained from either ginsenoside Rh2-treated or staurosporine-treated cells. Cleavage was equally inhibited in both cases by adding Ac-DEVD-CHO, a specific caspase-3 inhibitor, but not by Ac-YVAD-CHO, a specific caspase-1 inhibitor. Similarly, p21WAF1/CIP1 was efficiently cleaved by recombinant caspase-3, overexpressed in Escherichia coli. Moreover, the endogenous p21WAF1/CIP1 of untreated cell extracts was also cleaved by recombinant caspase 3, as measured by immunoblotting. Mutation analysis allowed identification of two caspase-3 cleavage sites, DHVD112/L and SMTD149/F, which are located within or near the interaction domains for cyclins, Cdks, and proliferating cell nuclear antigen (PCNA). Taken together, these results show that ginsenoside Rh2 and staurosporine increase caspase-3 activity, which in turn directly cleaves p21WAF1/CIP1 during the early stages of apoptosis. We propose that proteolytic cleavage of p21WAF1/CIP1 is a functionally relevant event that allows release of the cyclin/Cdk complex from the p21WAF1/CIP1 inhibitor, resulting in the elevated levels of cyclin/Cdk kinase activity seen in the earlier stage of apoptosis.
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PMID:Caspase 3 specifically cleaves p21WAF1/CIP1 in the earlier stage of apoptosis in SK-HEP-1 human hepatoma cells. 979 25

Tat proteins (trans-activating proteins) are present in all known lentiviruses and are early RNA binding proteins that regulate transcription. Tat from the human immunodeficiency virus type-1 is a protein comprising 86 amino acids and encoded by 2 exons. The first 72 amino acids are encoded by exon 1 and exhibit full trans-activating activity. The second exon encodes a 14-amino-acid C-terminal sequence that is not required for trans-activation but does contain an RGD motif, which is important in binding to alphavbeta3 and alpha5beta1 integrins. Tat has an unusual property for a transcription factor; it can be released and enter cells freely, yet still retain its activity, enabling it to up-regulate a number of genes. Tat also has an angiogenic effect; it is a potent growth factor for Kaposi sarcoma-derived spindle cells, and, separately, it has been shown to bind to a specific receptor, Flk-1/KDR, on vascular smooth muscle cells, as well as to integrin-like receptors present on rat skeletal muscle cells and the lymphocyte cell line H9. It appears that the basic domain of tat is important, not only for translocation but also for nuclear localisation and trans-activation of cellular genes. As such, targeting of tat protein or, more simply, the basic domain provides great scope for therapeutic intervention in HIV-1 infection. There is also opportunity for tat to be used as a molecular tool; the protein can be manipulated to deliver non-permeable compounds into cells, an approach that already has been employed using ovalbumin, beta-galactosidase, horseradish peroxidase, and caspase-3.
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PMID:HIV-1-trans-activating (Tat) protein: both a target and a tool in therapeutic approaches. 1053 42

CHO cells expressing the human insulin receptors (IR) were used to evaluate the effect of the potent farnesyltransferase inhibitor, manumycin, on insulin antiapoptotic function. Cell treatment with manumycin blocked insulin's ability to suppress pro-apoptotic caspase-3 activity which led to time-dependent proteolytic cleavage of two nuclear target proteins. The Raf-1/MEK/ERK cascade and the serine/threonine protein kinase Akt are two survival pathways that may be activated in response to insulin. We tested the hypothesis that inhibition of farnesylated Ras was causally related to manumycin-induced apoptosis and showed that the response to manumycin was found to be independent of K-Ras function because membrane association and activation of endogenous K-Ras proteins in terms of GTP loading and ERK activation were unabated following treatment with manumycin. Moreover, blocking p21Ras/Raf-1/MEK/ERK cascade by the expression of a transdominant inhibitory mSOS1 mutant in CHO-IR cells kept cells sensitive to the antiapoptotic action of insulin. Insulin-dependent activation of Akt was blocked by 4 h treatment with manumycin (P < 0.01), a kinetic too rapid to be explained by Ras inhibition. This study suggests that the depletion of short-lived farnesylated proteins by manumycin suppresses the antiapoptotic action of insulin at least in part by disrupting Akt activation but not that of the K-Ras/Raf-1/ERK-dependent cascade.
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PMID:Akt-dependent antiapoptotic action of insulin is sensitive to farnesyltransferase inhibitor. 1102 30

Low oxygen and nutrient depletion play critical roles in tumorigenesis, but little is known about how they interact to produce tumor survival and tumor malignancy. In the present study, we investigated the mechanism underlying hypoxia-modulated apoptosis of serum-deprived HepG2 cells. Our results showed that hypoxia blocked the apoptosis, which was accompanied with decreased Bax/Bcl-2 ratio, inhibited cytochrome c release, and reduced caspase-3 activity. More importantly, increased expressions of VEGF and its receptor-2 (KDR) under hypoxic/serum-deprived condition suggest that VEGF may act as a survival factor in a self-promoting manner. Data were further supported by results that recombinant human VEGF (rhVEGF) suppressed the serum deprivation-induced apoptosis, and anti-VEGF neutralizing antibody block anti-apoptotic activity of hypoxia. In addition, inhibitors of receptor tyrosine kinase blocked antiapoptosis of hypoxia. Our study further showed that rhVEGF or hypoxia induced ERK phosphorylation in serum-deprived cells, and that a specific inhibitor of MAPK/ERK, PD98059 eliminated the anti-apoptotic activity of rhVEGF or hypoxia by increasing Bax/Bcl-2 ratio and caspase-3 activity. Our data led us to conclude that induction of ERK phosphorylation and decrease of Bax/Bcl-2 ratio by rhVEGF implies that hypoxia-induced VEGF prevents apoptosis of serum-deprived cells by activating the MAPK/ERK pathway. Taken together, we propose that hypoxia enhances survival of nutrient-depleted tumor cells by reducing susceptibility to apoptosis, which consequently leads to tumor malignancy.
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PMID:Hypoxia-induced VEGF enhances tumor survivability via suppression of serum deprivation-induced apoptosis. 1103 Jan 51

When PC12 cells are deprived of trophic support they undergo apoptosis. We have previously shown that survival of trophic factor-deprived PC12M1 cells can be promoted by activation of the G protein-coupled muscarinic receptors. The mechanism whereby muscarinic receptors inhibit apoptosis is poorly understood. In the present study we investigated this mechanism by examining the effect of muscarinic receptor activation on the serum deprivation-induced activity of key players in apoptosis, the caspases, in PC12M1 cells. The results showed that m1 muscarinic activation inhibits caspase activity induced by serum deprivation. This effect appeared to be caused by the prevention of activation of caspases such as caspase-2 and caspase-3, and not by the inhibition of existing activity. Muscarinic receptor activation also stimulated the mitogen-activated protein kinase/extracellular signaling-regulated kinase (MAPK/ERK) and phosphoinositide (PI) 3-kinase signaling pathways. The PI 3-kinase pathway inhibitors wortmannin and LY294002, as well as the MAPK/ERK pathway PD98059 inhibitor, did not however suppress the inhibitory effect of the muscarinic receptors on caspase activity. The results therefore suggested that the muscarinic survival effect is mediated by a pathway that leads to caspase inhibition by MAPK/ERK- and PI 3-kinase-independent signaling cascades.
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PMID:M1 muscarinic receptors block caspase activation by phosphoinositide 3-kinase- and MAPK/ERK-independent pathways. 1104 77

In the present study, we investigated the effects of geranylgeraniol (GGO), a potent inducer of apoptosis in various lines of human tumor cells, on signal transduction cascades involved in apoptosis in human leukemia cells. GGO strongly induced the activation of c-Jun N-terminal kinase (JNK/SAPK) within 2 h in U937 and K562 cells, while neither ERK nor p38 was activated to any considerable extent during GGO-induced apoptosis. Transient expression of a constitutively active mutant form of mitogen-activated protein kinase kinase 1 (MEKK1), deltaMEKK1, or of deltaMEKK1-green fluorescent protein (GFP) in K562 cells activated JNK, but not a caspase-3-like protease, and was insufficient to induce cell death but rendered cells susceptible to GGO-induced cell death. Stable expressions of deltaMEKK1-GFP in U937 cells gave similar results. In contrast to VP-16-induced apoptosis, GGO-induced activation of JNK was almost completely inhibited by benzyloxycarbonyl-Asp-Glu-Val-Asp-fluoromethylketone (Z-DEVD) and by benzyloxycarbonyl-Asp-CH2OC[O]-2,6,-dichlorobenzene (Z-Asp), indicating that the JNK-activation step is located downstream of the caspase signaling pathway in GGO-induced apoptosis. Moreover, apoptosis induced by GGO was significantly inhibited in two lines of cells with a dominant-negative deletion mutation in c-Jun, indicating a requirement for JNK signaling. In addition, unlike the effects on other inducers of apoptosis, the activation of JNK and of the caspase-3-like protease by GGO was significantly delayed by 12-O-tetradecanoylphorbol-13-acetate (TPA), suggesting that the site of inhibition by TPA might be located upstream of the protease and JNK in the GGO-induced apoptotic signaling pathway.
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PMID:The mechanism of geranylgeraniol-induced apoptosis involves activation, by a caspase-3-like protease, of a c-jun N-terminal kinase signaling cascade and differs from mechanisms of apoptosis induced by conventional chemotherapeutic drugs. 1108 77

Caspase-3 knockout mice exhibit thickening of the internal granule cell layer of the cerebellum. Concurrently, it has been shown that intracerebral injection of pituitary adenylate cyclase-activating polypeptide (PACAP) induces a transient increase of the thickness of the cerebellar cortex. In the present study, we have investigated the possible effect of PACAP on caspase activity in cultured cerebellar granule cells from 8-day-old rat. Incubation of granule neurons with PACAP for 24 h promoted cell survival and prevented DNA fragmentation. Exposure of cerebellar granule cells to the specific caspase-3 inhibitor N-benzyloxycarbonyl-Asp-Glu-Val-Asp fluoromethylketone (Z-DEVD-FMK) for 24 h markedly enhanced cell survival and inhibited apoptotic cell death. Time-course studies revealed that PACAP causes a prolonged inhibition of caspase-3 activity without affecting caspase-1. Administration of graded concentrations of PACAP for 3 h induced a dose-dependent inhibition of caspase-3 activity. Incubation of granule cells with both dibutyryl-cAMP (dbcAMP) and phorbol 12-myristate 13-acetate (PMA) mimicked the inhibitory effect of PACAP on caspase-3. Cotreatment of cultured neurons with the protein kinase A inhibitor H89 and the protein kinase C inhibitor chelerythrine abrogated the effect of PACAP on caspase-3 activity. In contrast, the ERK kinase inhibitor U0126 did not affect the action of PACAP on caspase-3 activity. These data demonstrate that PACAP prevents cerebellar granule neurons from apoptotic cell death through a protein kinase A- and protein kinase C-dependent inhibition of caspase-3 activity.
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PMID:The neuroprotective effect of pituitary adenylate cyclase-activating polypeptide on cerebellar granule cells is mediated through inhibition of the CED3-related cysteine protease caspase-3/CPP32. 1108 78

Adipocyte number, a determinant of adipose tissue mass, reflects the balance between the rates of proliferation/differentiation vs. apoptosis of preadipocytes. The percentage of 3T3-L1 preadipocytes undergoing cell death following serum deprivation was reduced by 10 nM insulin-like growth factor (IGF)-1 (from 50.0 +/- 0.7% for control starved cells to 27.5 +/- 3.1%). TUNEL staining confirmed the apoptotic nature of the cell death. The protective effect of IGF-1 was blocked by phosphoinositide 3-kinase (PI3K) inhibitors, wortmannin, and LY294002, but was unaffected by rapamycin, PD98059, or SB203580, which inhibit mammalian target of rapamycin (mTOR), ERK kinase (MEK1), and p38 MAPK respectively. Exogenous PI(3,4,5)P3 (10 microM), the principal product of IGF-1-stimulated PI3K in 3T3-L1 preadipocytes, had a modest survival effect on its own, reducing cell death from 47.9 +/- 3.4% to 35.6 +/- 3.5%. When added to the combination of IGF-1 and LY294002, PI(3,4,5)P3 reversed most of the inhibitory effect of LY294002 on IGF-1-dependent cell survival, protein kinase B/Akt phosphorylation, and caspase-3 activity. Taken together, these results implicate PI(3,4,5)P3 as a necessary signal for the anti-apoptotic action of IGF-1 on 3T3-L1 preadipocytes.
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PMID:Phosphatidylinositol-3,4,5-trisphosphate is required for insulin-like growth factor 1-mediated survival of 3T3-L1 preadipocytes. 1114 83


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