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Query: UNIPROT:P42345 (mTOR)
 26,049 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Elongation factor 2 kinase (eEF2k) phosphorylates and inactivates eEF2. Insulin induces dephosphorylation of eEF2 and inactivation of eEF2 kinase, and these effects are blocked by rapamycin, which inhibits the mammalian target of rapamycin, mTOR. However, the signalling mechanisms underlying these effects are unknown. Regulation of eEF2 phosphorylation and eEF2k activity is lost in cells in which phosphoinositide-dependent kinase 1 (PDK1) has been genetically knocked out. This is not due to loss of mTOR function since phosphorylation of another target of mTOR, initiation factor 4E-binding protein 1, is not defective. PDK1 is required for activation of members of the AGC kinase family; we show that two such kinases, p70 S6 kinase (regulated via mTOR) and p90(RSK1) (activated by Erk), phosphorylate eEF2k at a conserved serine and inhibit its activity. In response to insulin-like growth factor 1, which activates p70 S6 kinase but not Erk, regulation of eEF2 is blocked by rapamycin. In contrast, regulation of eEF2 by stimuli that activate Erk is insensitive to rapamycin, but blocked by inhibitors of MEK/Erk signalling, consistent with the involvement of p90(RSK1).
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PMID:Regulation of elongation factor 2 kinase by p90(RSK1) and p70 S6 kinase. 1150 Mar 64

Amino acids positively regulate signaling through the mammalian target of rapamycin (mTOR). Recent work demonstrated the importance of the tuberous sclerosis protein TSC2 for regulation of mTOR by insulin. TSC2 contains a GTPase-activator domain that promotes hydrolysis of GTP bound to Rheb, which positively regulates mTOR signaling. Some studies have suggested that TSC2 also mediates the control of mTOR by amino acids. In cells lacking TSC2, amino acid withdrawal still results in dephosphorylation of S6K1, ribosomal protein S6, the eukaryotic initiation factor 4E-binding protein, and elongation factor-2 kinase. The effects of amino acid withdrawal are diminished by inhibiting protein synthesis or adding back amino acids. These studies demonstrate that amino acid signaling to mTOR occurs independently of TSC2 and involves additional unidentified inputs. Although TSC2 is not required for amino acid control of mTOR, amino acid withdrawal does decrease the proportion of Rheb in the active GTP-bound state. Here we also show that Rheb and mTOR form stable complexes, which are not, however, disrupted by amino acid withdrawal. Mutants of Rheb that cannot bind GTP or GDP can interact with mTOR complexes. We also show that the effects of hydrogen peroxide and sorbitol, cell stresses that impair mTOR signaling, are independent of TSC2. Finally, we show that the ability of energy depletion (which impairs mTOR signaling in TSC2+/+ cells) to increase the phosphorylation of eukaryotic elongation factor 2 is also independent of TSC2. This likely involves the phosphorylation of the elongation factor-2 kinase by the AMP-activated protein kinase.
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PMID:The tuberous sclerosis protein TSC2 is not required for the regulation of the mammalian target of rapamycin by amino acids and certain cellular stresses. 1577 76

Brief glutamatergic stimulation of neurons from fetal mice, cultured in vitro for 6 days, activates the mTOR-S6 kinase, ERK1/2 and Akt pathways, to an extent approaching that elicited by brain-derived neurotrophic factor. In contrast, sustained glutamatergic stimulation inhibits ERK, Akt, and S6K. Glutamatergic activation of S6K is calcium/calmodulin-dependent and is prevented by inhibitors of calcium/calmodulin-dependent protein kinase 2, phosphatidylinositol 3-OH-kinase and by rapamycin. 2-Amino-5-phosphonovaleric acid, an inhibitor of N'-methyl-D-aspartate receptors, abolishes glutamatergic activation of ERK1/2 but not the activation of mTOR-S6K; the latter is completely abolished by inhibitors of voltage-dependent calcium channels. Added singly, dopamine gives slight, and norepinephrine a more significant, activation of ERK and S6K; both catecholeamines, however, enhance glutamatergic activation of S6K but not ERK. After 12 days in culture, the response to direct glutamatergic activation is attenuated but can be uncovered by suppression of gamma-aminobutyric acid interneurons with bicuculline in the presence of the weak K(+) channel blocker 4-aminopyridine (4-AP). This selective synaptic activation of mTOR-S6K is also resistant to APV and inhibited by Ca(2+) channel blockers and higher concentrations of glutamate. Elongation factor 2 (EF2) is phosphorylated and inhibited by the eEF2 kinase (CaM kinase III); the latter is inhibited by the S6K or Rsk. Bicuculline/4-AP or KCl-induced depolarization reduces, whereas higher concentrations of glutamate increases, EF2 phosphorylation. Thus the mTOR-S6K pathway in neurons, a critical component of the late phase of LTP, is activated by glutamatergic stimulation in a calcium/calmodulin-dependent fashion through a calcium pool controlled by postsynaptic voltage-dependent calcium channels, whereas sustained stimulation of extrasynaptic glutamate receptors is inhibitory.
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PMID:Glutamatergic regulation of the p70S6 kinase in primary mouse neurons. 1618 39

Elongation factor-2 kinase (eEF-2 kinase), also known as Ca(2+)/calmodulin-dependent kinase III, regulates protein synthesis by controlling the rate of peptide chain elongation. The activity of eEF-2 kinase is increased in glioblastoma and other malignancies, yet its role in neoplasia is uncertain. Recent evidence suggests that autophagy plays an important role in oncogenesis and that this can be regulated by mammalian target of rapamycin (mTOR). Because eEF-2 kinase lies downstream of mTOR, we studied the role of eEF-2 kinase in autophagy using human glioblastoma cell lines. Knockdown of eEF-2 kinase by RNA interference inhibited autophagy in glioblastoma cell lines, as measured by light chain 3 (LC3)-II formation, acidic vesicular organelle staining, and electron microscopy. In contrast, overexpression of eEF-2 kinase increased autophagy. Furthermore, inhibition of autophagy markedly decreased the viability of glioblastoma cells grown under conditions of nutrient depletion. Nutrient deprivation increased eEF-2 kinase activity and decreased the activity of S6 kinase, suggesting an involvement of mTOR pathway in the eEF-2 kinase regulation of autophagy. These results suggest that eEF-2 kinase plays a regulatory role in the autophagic process in tumor cells; and eEF-2 kinase is a downstream member of the mTOR signaling; eEF-2 kinase may promote cancer cell survival under conditions of nutrient deprivation through regulating autophagy. Therefore, eEF-2 kinase may be a part of a survival mechanism in glioblastoma and targeting this kinase may represent a novel approach to cancer treatment.
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PMID:Elongation factor-2 kinase regulates autophagy in human glioblastoma cells. 1654 Jun 50

Elongation factor-2 kinase (eEF-2 kinase; Ca(2+)/calmodulin-dependent kinase III) controls the rate of peptide chain elongation. The activity of eEF-2 kinase is increased in many malignancies, yet its precise function in carcinogenesis remains unknown. Autophagy, a well-defined survival pathway in yeast, may also play an important role in oncogenesis. Furthermore, the autophagic response to nutrient deprivation is regulated by the mammalian target of rapamycin (mTOR). eEF-2 kinase lies downstream of mTOR and is regulated by several kinases in this pathway. Therefore, we studied the role of eEF-2 kinase in autophagy. Knockdown of eEF-2 kinase by RNA interference inhibited autophagy in several cell types as measured by light chain 3 (LC3)-II formation, acidic vesicular organelle staining, and electron microscopy. In contrast, overexpression of eEF-2 kinase increased autophagy. Furthermore, inhibition of autophagy markedly decreased the viability of glioblastoma cells grown under conditions of nutrient depletion. These results suggest that eEF-2 kinase plays a regulatory role in the autophagic process in tumor cells and may promote cancer cell survival under conditions of nutrient deprivation. Therefore, eEF-2 kinase activation may be a part of a survival mechanism in glioblastoma, and targeting this kinase may represent a novel approach to cancer treatment.
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PMID:Elongation factor-2 kinase: its role in protein synthesis and autophagy. 1692 Dec 68

In rat epitrochlearis skeletal muscle, contraction inhibited the basal and insulin-stimulated rates of protein synthesis by 75 and 70%, respectively, while increasing adenosine monophosphate-activated protein kinase (AMPK) activity. Insulin, on the other hand, stimulated protein synthesis (by 30%) and increased p70 ribosomal protein S6 kinase (p70S6K) Thr389, 40S ribosomal protein S6 (rpS6) Ser235/236, rpS6 Ser240/244 and eukaryotic initiation factor-4E-binding protein-1 (4E-BP1) Thr37/46 phosphorylation over basal values. Electrical stimulation had no effect on mammalian target of rapamycin complex 1 (mTORC1) signalling, as reflected by the lack of reduction in basal levels of p70S6K, rpS6 Ser235/236, rpS6 Ser240/244 and 4E-BP1 phosphorylation, but did antagonize mTORC1 signalling after stimulation of the pathway by insulin. Eukaryotic elongation factor-2 (eEF2) Thr56 phosphorylation increased rapidly on electrical stimulation reaching a maximum at 1 min, whereas AMPK Thr172 phosphorylation slowly increased to reach threefold after 30 min. Eukaryotic elongation factor-2 kinase (eEF2K) was not activated after 30 min of contraction when AMPK was activated. This could not be explained by the expression of a tissue-specific isoform of eEF2K in skeletal muscle lacking the Ser398 AMPK phosphorylation site. Therefore, in this skeletal muscle system, the contraction-induced inhibition of protein synthesis could not be attributed to a reduction in mTORC1 signalling but could be due to an increase in eEF2 phosphorylation independent of AMPK activation.
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PMID:Effects of contraction and insulin on protein synthesis, AMP-activated protein kinase and phosphorylation state of translation factors in rat skeletal muscle. 1795 82

Cytoplasmic translation is under sophisticated control but how cells adapt its rate to constitutive loss of mitochondrial oxidative phosphorylation is unknown. Here we show that translation is repressed in cells with the pathogenic A3243G mtDNA mutation or in mtDNA-less rho(0) cells by at least two distinct pathways, one transiently targeting elongation factor eEF-2 and the other initiation factor eIF-2alpha constitutively. Under conditions of exponential cell growth and mammalian target of rapamycin (mTOR) activation, eEF-2 becomes transiently phosphorylated by an AMP-activated protein kinase (AMPK)-dependent pathway, especially high in mutant cells. Independent of AMPK and mTOR, eIF-2alpha is constitutively phosphorylated in mutant cells, likely a signature of endoplasmic reticulum (ER)-stress response induced by the loss of oxidative phosphorylation. While the AMPK/eEF-2K/eEF-2 pathway appears to function in adaptation to physiological fluctuations in ATP levels in the mutant cells, the ER stress signified by constitutive protein synthesis inhibition through eIF-2alpha-mediated repression of translation initiation may have pathobiochemical consequences.
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PMID:Transient and constitutive repression of cytoplasmic translation signaling in cells with mtDNA mutation. 1913 59

Eukaryotic elongation factor-2 kinase (eEF2K) relays growth and stress signals to protein synthesis through phosphorylation and inactivation of eukaryotic elongation factor 2 (eEF2). 1-Benzyl-3-cetyl-2-methylimidazolium iodide (NH125) is a widely accepted inhibitor of mammalian eEF2K and an efficacious anti-proliferation agent against different cancer cells. It implied that eEF2K could be an efficacious anticancer target. However, eEF2K siRNA was ineffective against cancer cells including those sensitive to NH125. To test if pharmacological intervention differs from siRNA interference, we identified a highly selective small molecule eEF2K inhibitor A-484954. Like siRNA, A-484954 had little effect on cancer cell growth. We carefully examined the effect of NH125 and A-484954 on phosphorylation of eEF2, the known cellular substrate of eEF2K. Surprisingly, NH125 increased eEF2 phosphorylation, whereas A-484954 inhibited the phosphorylation as expected for an eEF2K inhibitor. Both A-484954 and eEF2K siRNA inhibited eEF2K and reduced eEF2 phosphorylation with little effect on cancer cell growth. These data demonstrated clearly that the anticancer activity of NH125 was more correlated with induction of eEF2 phosphorylation than inhibition of eEF2K. Actually, induction of eEF2 phosphorylation was reported to correlate with inhibition of cancer cell growth. We compared several known inducers of eEF2 phosphorylation including AMPK activators and an mTOR inhibitor. Interestingly, stronger induction of eEF2 phosphorylation correlated with more effective growth inhibition. We also explored signal transduction pathways leading to NH125-induced eEF2 phosphorylation. Preliminary data suggested that NH125-induced eEF2 phosphorylation was likely mediated through multiple pathways. These observations identified an opportunity for a new multipathway approach to anticancer therapies.
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PMID:1-Benzyl-3-cetyl-2-methylimidazolium iodide (NH125) induces phosphorylation of eukaryotic elongation factor-2 (eEF2): a cautionary note on the anticancer mechanism of an eEF2 kinase inhibitor. 2202 Sep 37

2-Deoxy-D-glucose (2-DG), a synthetic glucose analog that acts as a glycolytic inhibitor, is currently under clinical evaluation for targeting tumor cells. Tephrosin (TSN), a plant rotenoid, is known as an anticancer agent. In this study, we describe that the addition of TSN to 2-DG enhanced the cytotoxic activity of 2-DG against various types of cancer cells by accelerating ATP depletion and blunting autophagy. TSN increased the sensitivity of cancer cells to the cytotoxic effect of 2-DG. The combination of TSN and 2-DG induced acceleration of intracellular ATP depletion and the drastic activation of AMP-activated protein kinase (AMPK), which resulted in the inactivation of the mammalian target of rapamycin (mTOR) pathway. Of particular interest, TSN suppressed 2-DG-induced autophagy, a cell survival process in response to nutrient deprivation. We also showed that TSN inhibited 2-DG-induced activation of elongation factor-2 kinase (eEF-2K), which has been known to regulate 2-DG-induced autophagy. Inhibition of eEF-2K by RNA interference blunted 2-DG-induced autophagy and increased the sensitivity of cancer cells to the cytotoxic effect of 2-DG. The addition of TSN to 2-DG, however, did not enhance the cytotoxic activity of 2-DG by knockdown of eEF-2K, suggesting that inhibition of eEF-2K by tephrsoin could be a critical role in the potentiating effect of TSN on the cytotoxicity of 2-DG. Furthermore, we showed that the blunted autophagy and enhanced cytotoxicity of 2-DG was accompanied by the augmentation of apoptosis. These results show that TSN may be valuable for augmenting the therapeutic efficacy of 2-DG.
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PMID:The combination of tephrosin with 2-deoxy-D-glucose enhances the cytotoxicity via accelerating ATP depletion and blunting autophagy in human cancer cells. 2212 75

Recent advances in AMP-activated protein kinase (AMPK) as a target in cancer waxed and waned over the past decade of cancer research. AMPK is a cellular energy sensor, present in almost all eukaryotic cells. An elevated AMP/ATP ratio activates the AMPK, which in turn inhibits energy-consuming processes and induces catabolic events that generate ATP to restore the energy homeostasis inside the cell. Several reports have indicated that AMPK regulates several metabolic pathways and may be a potential therapeutic target for the treatment of cancer. Cancer cells have specific metabolic changes that differ from normal cells, and AMPK prevents the deregulated processes in cancer. AMPK may also act to inhibit tumor formation through modulation of cell growth, cell proliferation, autophagy, stress responses, and cell polarity. AMPK has been shown to inhibit mammalian target of rapamycin (mTOR) through tuberous sclerosis complex 2 (TSC2) phosphorylation and phosphatase and tensin homolog (PTEN), considered as central cell growth controller signals in diseases. In response to glucose deprivation, AMPK phosphorylates and activates p53, which induces cell cycle arrest in the G1/S phase of the cell cycle. AMPK has also been reported to block cyclin-dependent kinases through phosphorylation of p27(kip1) , promoting its stabilization and allowing cells to survive metabolic stress via induction of autophagy. Additionally, AMPK induces autophagy by phosphorylation and activation of eEF-2 kinase, and prevents the formation of new proteins. AMPK activators are also used for the treatment of type II diabetes and cancer. This review focuses on AMPK activation and its possible therapeutic role in the treatment of cancer.
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PMID:Role of AMP-activated protein kinase in cancer therapy. 2467 93


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