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

Leucine has been shown to stimulate adipose tissue protein synthesis in vivo as well as leptin secretion, protein synthesis, hyper-plastic growth, and tissue morphogenesis in in vitro experiments using freshly isolated adipocytes. Recently, others have proposed that leucine oxidation in the mitochondria may be required to activate the mammalian target of rapamycin (mTOR), the cytosolic Ser/Thr protein kinase that appears to mediate some of these effects. The first irreversible and rate-limiting step in leucine oxidation is catalyzed by the branched-chain alpha-keto acid dehydrogenase (BCKD) complex. The activity of this complex is regulated acutely by phosphorylation of the E1alpha-subunit at Ser293 (S293), which inactivates the complex. Because the alpha-keto acid of leucine regulates the activity of BCKD kinase, it has been suggested as a potential target for leucine regulation of mTOR. To study the regulation of BCKD phosphorylation and its potential link to mTOR activation, a phosphopeptide-specific antibody recognizing this site was developed and characterized. Phospho-S293 (pS293) immunoreactivity in liver corresponded closely to diet-induced changes in BCKD activity state. Immunoreactivity was also increased in TREMK-4 cells after the induction of BCKD kinase by a drug-inducible promoter. BCKD S293 phosphorylations in adipose tissue and gastrocnemius (which is mostly inactive in vivo) were similar. This suggests that BCKD complex in epididymal adipose tissue from food-deprived rats is mostly inactive (unable to oxidize leucine), as is the case in muscle. To begin to test the leucine oxidation hypothesis of mTOR activation, the dose-dependent effects of orally administered leucine on acute activation of S6K1 (an mTOR substrate) and BCKD were compared using the pS293 antibodies. Increasing doses of leucine directly correlated with increases in plasma leucine concentration. Phosphorylation of S6K1 (Thr389, the phosphorylation site leading to activation) in adipose tissue was maximal at a dose of leucine that increased plasma leucine approximately threefold. Changes in BCKD phosphorylation state required higher plasma leucine concentrations. The results seem more consistent with a role for BCKD and BCKD kinase in the activation of leucine metabolism/oxidation than in the activation of the leucine signal to mTOR.
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PMID:Potential role of leucine metabolism in the leucine-signaling pathway involving mTOR. 1281 18

The mammalian target of rapamycin (mTOR) is a Ser/Thr protein kinase that plays a crucial role in a nutrient-sensitive signalling pathway that regulates cell growth. TOR signalling is potently inhibited by rapamycin, through the direct binding of a FK506-binding protein 12 (FKBP12)/rapamycin complex to the TOR FRB domain, a segment amino terminal to the kinase catalytic domain. The molecular basis for the inhibitory action of FKBP12/rapamycin remains uncertain. Raptor (regulatory associated protein of mTOR) is a recently identified mTOR binding partner that is essential for mTOR signalling in vivo, and whose binding to mTOR is critical for mTOR-catalysed substrate phosphorylation in vitro. Here we investigated the stability of endogenous mTOR/raptor complex in response to rapamycin in vivo, and to the direct addition of a FKBP12/rapamycin complex in vitro. Rapamycin diminished the recovery of endogenous raptor with endogenous or recombinant mTOR in vivo; this inhibition required the ability of mTOR to bind the FKBP12/rapamycin complex, but was independent of mTOR kinase activity. Rapamycin, in the presence of FKBP12, inhibited the association of raptor with mTOR directly in vitro, and concomitantly reduced the mTOR-catalysed phosphorylation of raptor-dependent, but not raptor-independent substrates; mTOR autophosphorylation was unaltered. These observations indicate that rapamycin inhibits mTOR function, at least in part, by inhibiting the interaction of raptor with mTOR; this action uncouples mTOR from its substrates, and inhibits mTOR signalling without altering mTOR's intrinsic catalytic activity.
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PMID:Dissociation of raptor from mTOR is a mechanism of rapamycin-induced inhibition of mTOR function. 1506 26

The mammalian target of rapamycin (mTOR), a Ser/Thr protein kinase that mediates intracellular signalling related to cell growth, proliferation, and differentiation, has received considerable interest as a possible target for cancer treatment. We evaluated the correlation of mTOR expression with clinicopathological features, outcomes, and the expression of Akt, an upstream regulator of mTOR, in gastric cancer. Tumour samples were obtained from 109 patients with gastric adenocarcinomas who underwent a radical gastrectomy. The expressions of phosphorylated mTOR (p-mTOR) and phosphorylated Akt (p-Akt) in the cytoplasm and in the nucleus were analysed by immunohistochemical staining. Cytoplasmic p-mTOR expression positively correlated with the depth of tumour invasion (T1 vs T2-4, P=0.003), involved lymph nodes (P=0.010), and tumour stage (I vs II-IV, P=0.002). In contrast, nuclear p-mTOR expression negatively correlated with these variables (P<0.001,=0.035, and <0.001). Cytoplasmic p-mTOR expression was associated with significantly poorer relapse-free survival (RFS, P=0.037) and overall survival (OS, P=0.024), whereas nuclear p-mTOR expression was associated with better RFS and OS (P=0.029, 0.059). Neither cytoplasmic nor nuclear p-Akt expression was associated with any clinicopathological factor or with survival. Localisation of p-mTOR may play an important role in tumour progression and outcomes in patients with gastric cancer.
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PMID:Relation between outcomes and localisation of p-mTOR expression in gastric cancer. 1922 2

Ser/Thr protein kinase PKB/Akt is a key regulator of a wide range of cellular processes including growth, proliferation and survival. PKB is clearly a crucial signaling molecule and extensive research efforts aim to understand its regulation and action. Recent studies of the regulation of PKB activity by hydrophobic motif phosphorylation have yielded several exciting findings about members of the PI3-kinase-like family of kinases (PIKKs) acting as PKB regulators. Mammalian target of rapamycin complex 2 (mTORC2) and DNA-dependent protein kinase (DNA-PK) can both phosphorylate Ser473 and activate PKB. This present review concerns PKB regulation by mTORC2 and DNA-PK in a stimulus-dependent and context-dependent manner and the possible implications of this for PKB activity, substrate specificity and therapeutic intervention.
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PMID:PIKKing on PKB: regulation of PKB activity by phosphorylation. 1930 58

The mammalian target of rapamycin (mTOR) is a Ser/Thr protein kinase and a major controller of cell growth. In cells, mTOR forms two distinct multiprotein complexes, mTORC1 and mTORC2. The mTORC1 complex can phosphorylate 4EBP1 and S6K1, two key regulators of translation initiation, whereas mTORC2 phosphorylates AKT1, an event required for AKT1 activation. Here, we expressed and purified human mTORC1 and mTORC2 from HEK-293 cells using FLAG-M2 affinity chromatography. Western blotting analysis using phospho-specific antibodies indicated that recombinant mTORC1 and mTORC2 exhibit distinct substrate preferences in vitro, consistent with their roles in cells. To improve our understanding of the enzymatic properties of mTOR alone and mTOR in its complex form, steady-state kinetic profiles of truncated mTOR containing the kinase domain (residues 1360-2549) and mTORC1 were determined. The results revealed that mTORC1 is catalytically less active than truncated mTOR, as evidenced by 4.7- and 3.1-fold decreases in catalytic efficiency, k(cat)/K(m), for ATP and 4EBP1, respectively. We also found that truncated mTOR undergoes autophosphorylation through an intramolecular mechanism. Mass spectrometric analysis identified two novel mTOR autophosphorylation sites, Ser2454 and either Thr2473 or Thr2474, in addition to the previously reported Ser2481 site. Truncated mTOR and mTORC1 were completely inhibited by ATP competitive inhibitors PI103 and BEZ235 and partially inhibited by rapamycin/FKBP12 in a noncompetitive fashion toward ATP. All inhibitors tested exhibited similar inhibitory potencies between mTORC1 and truncated mTOR containing the kinase domain. Our studies presented here provide the first detailed kinetic studies of a recombinant mTOR complex.
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PMID:Steady-state kinetic and inhibition studies of the mammalian target of rapamycin (mTOR) kinase domain and mTOR complexes. 2080 12

mTOR, an evolutionarily conserved Ser/Thr protein kinase, belongs to the PI3K-related kinase family, which also includes DNA-PKcs, ATM, and ATR. Although other PI3K-related kinase family members have been shown to secure genomic integrity by sensing DNA damage and related stresses, mTOR is known to function as a nutrient and growth factor sensor. mTOR is the catalytic subunit of two distinct multiprotein complexes known as mTOR complex 1 (mTORC1) and mTOR complex 2 (mTORC2). In response to growth factor and nutrient availability, these complexes regulate a variety of cellular processes, such as cell growth, proliferation, and survival by modulating downstream effectors, such as S6K1, 4EBP1, and AKT. Therefore, evaluation of mTOR activity has been a clear readout in order to monitor the physiological status of cells in response to environmental cues. Here, we present the current techniques for the assessment of mTOR kinase activity in different experimental settings.
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PMID:Evaluation of the nutrient-sensing mTOR pathway. 2212 58

Ser/Thr protein kinase CK2 regulates multiple processes that play important roles in the sensitivity of cancer to epidermal growth factor receptor targeting therapeutics, including PI3K-Akt-mTOR signaling, Hsp90 activity, and inhibition of apoptosis. We hypothesized that top-down inhibition of EGFR, combined with lateral suppression of multiple oncogenic pathways by targeting CK2, would create a pharmacologic synthetic lethal event and result in an improved cancer therapy compared to EGFR inhibition alone. This hypothesis was tested by combining CX-4945, a first-in-class clinical stage inhibitor of CK2, with the EGFR tyrosine kinase inhibitor, erlotinib, in vitro and in vivo in models of non-small cell lung carcinoma, NCI-H2170, and squamous cell carcinoma, A431. Our results demonstrate that combination of CX-4945 with erlotinib results in enhanced attenuation of the PI3K-Akt-mTOR pathway. We also observed an increase in apoptosis, synergistic killing of cancer cells in vitro, as well as improved antitumor efficacy in vivo. Taken together, these data position CK2 as a valid pharmacologic target for drug combinations and support further evaluation of CX-4945 in combination with EGFR targeting agents.
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PMID:Combined inhibition of EGFR and CK2 augments the attenuation of PI3K-Akt-mTOR signaling and the killing of cancer cells. 2238 88

Hypoxic or ischemic stress causes serious brain injury via various pathologic mechanisms including suppressed protein synthesis, neuronal apoptosis, and the release of neurotoxic substances. Many neuroprotective treatments of hypoxic or ischemic brain injury rely on these pathologic mechanisms. The mammalian target of rapamycin (mTOR), an atypical Ser/Thr protein kinase, could be a novel therapeutic target. mTOR plays a critical role in regulating many activities such as protein synthesis, cell growth, and cell death. Furthermore, mTOR could promote angiogenesis, neuronal regeneration, and synaptic plasticity, reduce neuronal apoptosis, and remove neurotoxic substances, which are all closely associated with the repair and survival mechanisms of hypoxic or ischemic brain injury. Although there is currently controversy with regard to regulating the activation of mTOR, the effective neuroprotective functions resulting from mTOR activation have been confirmed by various studies. Considering the potential capability for mTOR in regulating the repair and survival mechanisms of hypoxic or ischemic brain injury, mTOR may be a novel target for neuroprotective treatment.
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PMID:Role of mammalian target of rapamycin in hypoxic or ischemic brain injury: potential neuroprotection and limitations. 2275 85

Mammalian target of rapamycin (mTOR), a Ser/Thr protein kinase, is the catalytic component of two distinct signaling complexes, mTOR-raptor complex (mTORC1) and mTOR-rictor complex (mTORC2). Recently, studies have demonstrated mitosis-specific roles for mTORC1, but the functions and expression dynamics of mTOR complexes during meiotic maturation remain unclear. In the present study, to evaluate the roles of respective mTOR complexes in maternal meiosis and compare them with those in mitosis, we sought to elucidate the spatiotemporal immunolocalization of mTOR, the kinase-active Ser2448- and Ser2481-phosphorylated mTOR, and raptor and rictor during cumulus-cell mitosis and oocyte meiotic maturation in mice. mTOR principally accumulated around the chromosomes and on the spindle. Phosphorylated mTOR (Ser2448 and Ser2481) exhibited elevated fluorescence intensities in the cytoplasm and punctate localization adjacent to the chromosomes, on the spindle poles, and on the midbody during mitotic and meiotic maturation, suggesting functional homology of mTOR between the two cell division systems, despite their mechanistically distinctive spindles. Raptor colocalized with mTOR during both types of cell division, indicating that mTORC1 is predominantly associated with these events. Mitotic rictor uniformly distributed through the cytoplasm, and meiotic rictor localized around the spindle poles of metaphase-I oocytes, suggesting functional divergence of mTORC2 between mitosis and female meiosis. Based on the general function of mTORC2 in the organization of the actin cytoskeleton, we propose that mTORC1 controls spindle function during mitosis and meiosis, while mTORC2 contributes to actin-dependent asymmetric division during meiotic maturation in mice.
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PMID:Distribution and association of mTOR with its cofactors, raptor and rictor, in cumulus cells and oocytes during meiotic maturation in mice. 2344 Aug 73

The mammalian target of rapamycin (mTOR) is a Ser/Thr protein kinase conserved in all eukaryotes that plays a key role in cell growth and is a central effector of several pathways regulating essential cell functions. Hyperactivation of the mTORdependent signalling pathway occurs in many human diseases and may be a selective target for their therapy. However, the dual nature of mTOR, existing in two multiprotein complexes mTORC1 and mTORC2 driven by different feedback loops, decreases the therapeutic effects of rapamycin, the specific mTOR inhibitor. In the present study we demonstrate that the mTORC1 signalling pathway is highly activated in human melanoma cells and that up-regulation of this pathway along with the growth and malignity of these cells could be suppressed by disruption of the Src activity. SU6656, the selective inhibitor of the Src kinase activity, decreased up-regulation of the mTORC1 signalling and moreover, unlike rapamycin, it did not induce the activation of Akt/PKB and its downstream targets in HBL melanoma cells. The Src protein was found to be associated with raptor in the mTORC1 complex immunoprecipitated from these cells, suggesting that the Src activity might be a new attractive target for monotherapeutic inhibition of the up-regulated mTORC1 signalling pathway.
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PMID:Inhibition of mTORC1 by SU6656, the selective Src kinase inhibitor, is not accompanied by activation of Akt/PKB signalling in melanoma cells. 2409 74


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