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:2.7.11.2 (PDK1)
2,238 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The RET/PTC3 oncogene is a genetically rearranged and constitutively activated tyrosine kinase receptor that is common in papillary thyroid cancer. Because RET/PTC3 is chronically overexpressed in these thyroid cancer cells, and RET/PTC3-expressing tumors are associated with overactivity of tyrosine kinase signaling pathways and a more aggressive clinical course, we questioned whether chronic RET/PTC3 expression enhances cellular responses to thyroid mitogens in vitro. We stably transfected FRTL-5 cells with the RET/PTC3 gene; transfected and control cell lines were cultured without insulin, TSH, or serum. Thymidine incorporation into DNA was enhanced in the RET/PTC3 cells, but transformation was not observed. RET/PTC3 cells demonstrated higher basal and insulin-stimulated levels of activated Akt, both of which were reduced by LY294002, a PI3 kinase inhibitor, but not PD98059, a MEK inhibitor. By contrast, mitogen activated protein kinase (MAP kinase) was only minimally activated in RET/PTC3 cells before and after stimulation. Consistent with preferential activation of PI3 kinase, increased levels of total and phosphorylated IRS2 protein, relative activation of PDK-1, and enhanced IRS2-p85 interactions were identified in RET/PTC3-expressing cells. RET/PTC3 cells were also sensitized to insulin-induced thymidine incorporation; this effect was blocked by PI3 kinase (LY294002) rather than MEK 1/2 (PD98059) inhibitors. In summary, we have demonstrated that RET/PTC3 expression enhances basal and insulin-stimulated DNA synthesis through PI3 kinase, cooperatively activates Akt with insulin via PI3 kinase, and preferentially activates the Akt rather than MAP kinase pathway in FRTL-5 cells.
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PMID:Chronic expression of RET/PTC 3 enhances basal and insulin-stimulated PI3 kinase/AKT signaling and increases IRS-2 expression in FRTL-5 thyroid cells. 1537 48

The RAS-activated RAF-->MEK-->extracellular signal-regulated kinase (ERK) and phosphatidylinositol 3'-kinase (PI3'-kinase)-->PDK1-->AKT signaling pathways are believed to cooperate to promote the proliferation of normal cells and the aberrant proliferation of cancer cells. To explore the mechanisms that underlie such cooperation, we have derived cells harboring conditionally active, steroid hormone-regulated forms of RAF and AKT. These cells permit the assessment of the biological and biochemical effects of activation of these protein kinases either alone or in combination with one another. Under conditions where activation of neither RAF nor AKT alone promoted S-phase progression, coactivation of both kinases elicited a robust proliferative response. Moreover, under conditions where high-level activation of RAF induced G(1) cell cycle arrest, activation of AKT bypassed the arrest and promoted S-phase progression. At the level of the cell cycle machinery, RAF and AKT cooperated to induce cyclin D1 and repress p27(Kip1) expression. Repression of p27(Kip1) was accompanied by a dramatic reduction in KIP1 mRNA and was observed in primary mouse embryo fibroblasts derived from mice either lacking SKP2 or expressing a T187A mutated form of p27(Kip1). Consistent with these observations, pharmacological inhibition of MEK or PI3'-kinase inhibited the effects of activated RAS on the expression of p27(Kip1) in NIH 3T3 fibroblasts and in a panel of bona fide human pancreatic cancer cell lines. Furthermore, we demonstrated that AKT activation led to sustained activation of cyclin/cdk2 complexes that occurred concomitantly with the removal of RAF-induced p21(Cip1) from cyclin E/cdk2 complexes. Cumulatively, these data strongly suggest that the RAF-->MEK-->ERK and PI3'K-->PDK-->AKT signaling pathways can cooperate to promote G(0)-->G(1)-->S-phase cell cycle progression in both normal and cancer cells.
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PMID:Cooperative regulation of the cell division cycle by the protein kinases RAF and AKT. 1557 89

Cerebellar granule neurons undergo apoptosis when switched from culture medium containing high potassium (HK) to medium that contains low potassium (LK). HK treatment leads to an activation of p21-activated kinase-1 (PAK-1). Overexpression of a constitutively active form of PAK-1 protects against apoptosis in LK medium. Overexpression of a dominant-negative form of PAK-1 blocks survival in HK. Although PAK-1 is usually considered to be a downstream effector of Rac and Cdc42, we were unable to detect association between PAK-1 and either Rac1 or Cdc42 in cerebellar granule neurons. Interaction between PAK-1 and PDK1 is detected in granule neurons, although there is no change in the extent of interaction in neurons primed to die. Neuronal survival by PAK-1 overexpression is not inhibited by PD98059 or LY294002, which inhibit the activity of MEK and PI-3 kinase, respectively. The ability of PAK-1 to maintain neuronal survival is, however, blocked by ML-9, a compound known to inhibit Akt. Our results show that that PAK-1 is necessary for neuronal survival in HK and suggest that its neuroprotective action may be mediated by a GTPase-independent, but Akt-dependent, mechanism.
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PMID:p21-Activated kinase-1 is necessary for depolarization-mediated neuronal survival. 1569 23

Epidermal growth factor (EGF) is a potent mitogen for mesangial cells. The mechanism by which EGF induces DNA synthesis is not precisely understood. We investigated the role of phosphatidylinositol (PI)3-kinase in regulating mitogenesis. EGF increased PI3-kinase activity resulting in stimulation of PDK-1 and Akt kinase activities. Blocking of PI3-kinase activity using LY-294002 or adenoviral expression of PTEN, which dephosphorylates PI3,4,5-tris-phosphate and thus inactivates PI3-kinase signaling, significantly inhibits EGF-induced DNA synthesis. Expression of dominant-negative Akt kinase, however, had no effect on DNA synthesis. But it inhibited EGF-induced phosphorylation of FoxO3a transcription factor, thus demonstrating its functional consequences. These data indicate that EGF increases the DNA synthesis in a PI3-kinase-dependent but Akt-independent manner. In addition to activating PI3-kinase signaling, EGF increased Erk1/2 MAPK activity, leading to transcriptional activation of its nuclear target Elk-1 and resulting in c-fos expression. Inhibition of MAPK activity by MEK inhibitor U-0126 abolished EGF-induced DNA synthesis. Because EGF activates PI3-kinase, which also regulates DNA synthesis, the effect of PI3-kinase on MAPK activity was also examined. Inhibition of PI3-kinase signaling blocked EGF-induced MAPK activity as well as Elk-1-dependent reporter transcription and c-fos gene transcription. To further determine the mechanism of EGF-induced DNA synthesis, we investigated the effect of EGF on the cyclin-dependent kinase inhibitor p27(Kip1). EGF reduced the expression of p27(Kip1). Inhibition of PI3-kinase action or MAPK activity abolished the reduction in p27(Kip1) expression induced by EGF. These data provide the evidence that a linear signal transduction pathway involving PI3-kinase-dependent MAPK regulates EGF-induced DNA synthesis in mesangial cells by regulating c-fos and p27(Kip1) expression.
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PMID:EGF stimulates mesangial cell mitogenesis via PI3-kinase-mediated MAPK-dependent and AKT kinase-independent manner: involvement of c-fos and p27Kip1. 1570 16

We showed previously [K. Moissoglu, I.H. Gelman, J. Biol. Chem. 278 (2003) 47946-47959] that oncogenic v-Src could induce 7- to 10-fold greater anchorage-independent growth (AIG) in FAK-null mouse embryo fibroblasts (MEF) compared to those expressing FAK. Here, we demonstrate that the enhanced AIG (eAIG) correlates with increased activation levels of phosphatidylinositol 3-kinase (PI3K) and not with changes in the protein levels of the p85 regulatory subunit of PI3K, PDK1 or PTEN- modulators, and/or mediators of PI3K activity. eAIG could be blunted selectively by treatment with the PI3K inhibitor, LY294002, or by overexpression of either the PI3K antagonist, PTEN, dominant-interfering alleles of PI3K or a downstream PI3K mediator, AKT, but not by the MEK inhibitor, PD98059, dominant-interfering alleles of MEK or the signal transducer and activator of transcription (STAT)-3. In contrast, RNAi-mediated knockdown of FAK resulted in increased v-Src-induced AIG. Expression of a constitutively active PI3K allele was sufficient to induce higher levels of AIG, whereas overexpression of v-Src produced only larger-sized colonies in soft agar. Interestingly, FAK was required for full activation of PI3K by PDGF whereas the activation of PI3K by insulin was significantly increased in FAK-/- cells. Thus, although FAK is dispensable for v-Src-induced oncogenic transformation in vitro, it may exert either positive or negative effects on signaling or motility depending on which pathways are activated in cancer cells.
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PMID:Enhanced v-Src-induced oncogenic transformation in the absence of focal adhesion kinase is mediated by phosphatidylinositol 3-kinase. 1580 50

Characteristics of hVSMC apoptosis and its inhibition by insulin-like growth factor-1 (IGF-1) remain unclear. Also unclear is whether a balance in hVSMCs exists whereby c-Jun N-terminal stress kinases (JNK) promote apoptosis while extracellular signal-regulated (ERK1/2) MAP kinases inhibit cell death. In this study, we examined the involvement of Akt/PKB and its upstream kinase, PDK1 and whether JNK activation correlated with human and rat VSMC apoptosis induced by staurosporine and by c-myc, respectively. We observed a strong, sustained JNK activation (and c-Jun phosphorylation), which correlated with VSMC apoptosis. IGF-1 (13.3 nM), during apoptosis inhibition, transiently inhibited JNK activity at 1 h in a phosphatidylinositol 3-kinase (PI3-K)- and MEK-ERK-dependent manner, as wortmannin (100 nM) or PD98059 (30 muM) partially attenuated the IGF-1 effect. PKC down-regulation had no effect on JNK inhibition by IGF-1. While IGF-1 alone produced a strong phosphorylation of Akt/PKB in hVSMCs up to 6 h, it was notably stronger and more sustained during ratmyc and hVSMCs apoptosis inhibition. Further, whereas transient expression of phosphorylated Akt protected VSMCs from apoptosis by nearly 50%, expression of dominant interfering alleles of Akt or PDK1 strongly inhibited IGF-1-mediated VSMC survival. These results demonstrate for the first time that transient inhibition of a pro-apoptotic stimulus in VSMCs may be sufficient to inhibit a programmed cell death and that sustained anti-apoptotic signals (Akt) elicited by IGF-1 are augmented during a death stimulus. Furthermore, PI3-K and ERK-MAPK pathways may cooperate to protect VSMCs from cell death.
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PMID:Sustained Akt/PKB activation and transient attenuation of c-jun N-terminal kinase in the inhibition of apoptosis by IGF-1 in vascular smooth muscle cells. 1590 15

Nonsteroidal anti-inflammatory drugs (NSAIDs) are frequently used as analgesics. They inhibit cyclooxygenases (COX), preventing the formation of prostaglandins, including prostacyclin and thromboxane. A serious side effect of COX-1 and COX-2 inhibitors is renal damage. To investigate the molecular basis of the renal injury, we evaluated the expression of the stress marker, heme oxygenase-1 (HO-1), in celecoxib-stimulated mesangial cells. We report here that a COX-2 selective NSAID, celecoxib, induced a concentration- and time-dependent increase of HO-1 expression in glomerular mesangial cells. Celecoxib-induced HO-1 protein expression was inhibited by actinomycin D and cycloheximide, suggesting that de novo transcription and translation are required in this process. N-acetylcysteine, a free radical scavenger, strongly decreased HO-1 expression, suggesting the involvement of reactive oxygen species (ROS). Celecoxib-induced HO-1 expression was attenuated by pretreatment of the cells with SP 600125 (a specific JNK inhibitor), but not SB 203580 (a specific p38 MAPK inhibitor), or PD 98059 (a specific MEK inhibitor). Consistently, celecoxib activated c-Jun N-terminal kinase (JNK) as demonstrated by kinase assays and by increasing phosphorylation of this kinase. N-acetylcysteine reduced the stimulatory effect of celecoxib on stress kinase activities, suggesting an involvement of JNK in HO-1 expression. On the other hand, LY 294002, a phosphatidylinositol 3-kinase (PI-3K)-specific inhibitor, prevented the enhancement of HO-1 expression. This effect was correlated with inhibition of the phosphorylation of the PDK-1 downstream substrate Akt/protein kinase B (PKB). In conclusion, our data suggest that celecoxib-induced HO-1 expression in glomerular mesangial cells may be mediated by ROS via the JNK-PI-3K cascade.
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PMID:Celecoxib induces heme-oxygenase expression in glomerular mesangial cells. 1596 68

Little is known about the regulatory mechanisms of endothelial cell (EC) proliferation by retinal pericytes and vice versa. In a model of coculture with bovine retinal pericytes lasting for 24 h, rat brain ECs showed an increase in arachidonic acid (AA) release, whereas Western blot and RT-PCR analyses revealed that ECs activated the protein expression of cytosolic phospholipase A(2) (cPLA(2)) and its phosphorylated form and calcium-independent intracellular phospholipase A(2) (iPLA(2)). No activation of the same enzymes was seen in companion pericytes. In ECs, the protein level of phosphorylated extracellular signal-regulated kinase (ERK) 1/2 was also enhanced significantly, a finding not observed in cocultured pericytes. The expression of protein kinase C-alpha (PKCalpha) and its phosphorylated form was also enhanced in ECs. Wortmannin, LY294002, and PD98059, used as inhibitors of upstream kinases (the PI3-kinase/Akt/PDK1 or MEK-1 pathway) in cultures, markedly attenuated AA release and the expression of phosphorylated forms of endothelial cPLA(2), PKCalpha, and ERK1/2. By confocal microscopy, activation of PKCalpha in perinuclear regions of ECs grown in coculture as well as strong activation of cPLA(2) in ECs taken from a model of mixed culture were clearly observed. However, no increased expression of both enzymes was found in cocultured pericytes. Our findings indicate that a sequential activation of PKCalpha contributes to endothelial ERK1/2 and cPLA(2) phosphorylation induced by either soluble factors or direct cell-to-cell contact, and that the PKCalpha-cPLA(2) pathway appears to play a key role in the early phase of EC-pericyte interactions regulating blood retina or blood-brain barrier maturation.
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PMID:Endothelial cell-pericyte cocultures induce PLA2 protein expression through activation of PKCalpha and the MAPK/ERK cascade. 1726 47

p90 ribosomal S6 kinase (RSK1) is an effector of both Ras/MEK/MAPK and PI3K/PDK1 pathways. We present evidence that RSK1 drives p27 phosphorylation at T198 to increase RhoA-p27 binding and cell motility. RSK1 activation and p27pT198 both increase in early G(1). As for many kinase-substrate pairs, cellular RSK1 coprecipitates with p27. siRNA to RSK1 and RSK1 inhibition both rapidly reduce cellular p27pT198. RSK1 overexpression increases p27pT198, p27-cyclin D1-Cdk4 complexes, and p27 stability. Moreover, RSK1 transfectants show mislocalization of p27 to cytoplasm, increased motility, and reduced RhoA-GTP, phospho-cofilin, and actin stress fibers, all of which were reversed by shRNA to p27. Phosphorylation by RSK1 increased p27pT198 binding to RhoA in vitro, whereas p27T157A/T198A bound poorly to RhoA compared with WTp27 in cells. Coprecipitation of cellular p27-RhoA was increased in cells with constitutive PI3K activation and increased in early G(1). Thus T198 phosphorylation not only stabilizes p27 and mislocalizes p27 to the cytoplasm but also promotes RhoA-p27 interaction and RhoA pathway inhibition. These data link p27 phosphorylation at T198 and cell motility. As for other PI3K effectors, RSK1 phosphorylates p27 at T198. Because RSK1 is also activated by MAPK, the increased cell motility and metastatic potential of cancer cells with PI3K and/or MAPK pathway activation may result in part from RSK1 activation, leading to accumulation of p27T198 in the cytoplasm, p27:RhoA binding, inhibition of RhoA/Rock pathway activation, and loss of actomyosin stability.
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PMID:RSK1 drives p27Kip1 phosphorylation at T198 to promote RhoA inhibition and increase cell motility. 1947 Apr 70

We explored the crosstalk between cell survival (phosphatidylinositol 3-kinase (PI3K)/Akt) and mitogenic (Ras/Raf/MEK/extracellular signal-regulated kinase (ERK)) signaling pathways activated by an epidermal growth factor (EGF) and analyzed their sensitivity to small molecule inhibitors in the PI3K-mutant estrogen receptor (ER)-positive MCF7 and T47D breast cancer cells. In contrast to MCF7 cells, ERK phosphorylation in T47D cells displayed resistance to MEK inhibition by several structurally different compounds, such as U0126, PD 098059 and PD 198306, MEK suppression by small interfering RNA (siRNA) and was also less sensitive to PI3K inhibition by wortmannin. Similar effect was observed in PI3K-wild type ER-positive BT-474 cells, albeit to a much lesser extent. MEK-independent ERK activation was induced only by ErbB receptor ligands and was resistant to inhibition of several kinases and phosphatases that are known to participate in the regulation of Ras/mitogen-activated protein kinase (MAPK) cascade. Although single agents against PDK1 or Akt did not affect EGF-induced ERK phosphorylation, a combination of PI3K/Akt and MEK inhibitors synergistically suppressed ERK activation and cellular growth. siRNA-mediated silencing of class I PI3K or Akt1/2 genes also significantly decreased U0126-resistant ERK phosphorylation. Our data suggest that in T47D cells ErbB family ligands induce a dynamic, PI3K/Akt-sensitive and MEK-independent compensatory ERK activation circuit that is absent in MCF7 cells. We discuss candidate proteins that can be involved in this activation circuitry and suggest that PDZ-Binding Kinase/T-LAK Cell-Originated Protein Kinase (PBK/TOPK) may play a role in mediating MEK-independent ERK activation.
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PMID:PI3K/Akt-sensitive MEK-independent compensatory circuit of ERK activation in ER-positive PI3K-mutant T47D breast cancer cells. 2047 74


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