Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P42345 (mTOR)
 26,049 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The immunosuppressant rapamycin, in complex with its cellular receptor FKBP12, targets the cellular protein FKBP12-rapamycin-associated protein/mammalian target of rapamycin/rapamycin and FKBP12 target 1 (FRAP/mTOR/RAFT1) and inhibits/delays G1 cell cycle progression in mammalian cells. As a member of the novel phosphatidylinositol kinase-related kinase family, FRAP's kinase activity is essential for its signaling function. The FKBP12-rapamycin binding (FRB) domain in FRAP is also speculated to play an important role in FRAP function and signaling. However, the biochemical and physiological functions of FRB, as well as the mechanism for rapamycin inhibition, have been unclear. The present study focuses on investigation of FRB's role and the functional relationship between FRB domain and kinase domain in FRAP. Microinjection of purified FRB protein into human osteosarcoma MG63 cells results in a drastic blockage of the G1 to S cell cycle progression; such a dominant negative effect is reversed by a point mutation (Trp2027 --> Phe). The same mutation also abolishes kinase activity of FRAP without affecting ATP binding, and truncation studies suggest that upstream sequences including FRB are required for kinase activity in vitro. Given these data, we propose a model for FRAP function, in which the FRB domain is required for activation of the kinase domain, possibly through the interaction with an upstream activator. In addition, our observations provide direct evidence linking FRAP function to G1 cell cycle progression.
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PMID:The FKBP12-rapamycin-binding domain is required for FKBP12-rapamycin-associated protein kinase activity and G1 progression. 993 27

Aging is characterized by a decrease of muscle mass associated with a decrease in postprandial anabolism. This study was performed to gain a better understanding of the intracellular mechanisms involved in the stimulation of muscle protein synthesis by amino acids and their role in the decrease of muscle sensitivity to food intake during aging. The effects of amino acids or leucine alone were assessed in vitro on epitrochlearis muscle from young, adult and old rats. Protein synthesis was assessed by incorporation of radiolabeled phenylalanine into protein and p70 S6 kinase activity by incorporation of (32)P into a synthetic substrate. Amino acids, at physiologic concentrations, stimulated muscle protein synthesis (P < 0.05) and leucine reproduced this effect. The intracellular targets of amino acids were phosphatidylinositol 3' kinase and the rapamycin-sensitive pathways mammalian target of rapamycin (mTOR)/p70 S6 kinase. In old rats, the sensitivity of muscle protein synthesis to leucine was lower than in adults (P < 0.05) and this paralleled the lesser ability of leucine to stimulate the rapamycin-sensitive pathways (P < 0.05). We demonstrated that amino acids and leucine stimulate muscle protein synthesis and that aging is associated with a decrease in this effect. However, because aged rats are still able to respond normally to high leucine concentrations, we hypothesize that a nutritional manipulation increasing the availability of this amino acid to muscle could be beneficial in maintaining the postprandial stimulation of protein synthesis.
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PMID:Stimulation of in vitro rat muscle protein synthesis by leucine decreases with age. 1105 98

Mammalian target of rapamycin (mTOR) is the central element of a signaling pathway involved in the control of mRNA translation and cell growth. The actions of mTOR are mediated in part through the phosphorylation of the eukaryotic initiation factor 4E-binding protein, PHAS-I. In vitro mTOR phosphorylates PHAS-I in sites that control PHAS-I binding to eukaryotic initiation factor 4E; however, whether mTOR directly phosphorylates PHAS-I in cells has been a point of debate. The Arg-Ala-Ile-Pro (RAIP motif) and Phe-Glu-Met-Asp-Ile (tor signaling motif) sequences found in the NH2- and COOH-terminal regions of PHAS-I, respectively, are required for the efficient phosphorylation of PHAS-I in cells. Here we show that mutations in either motif markedly decreased the phosphorylation of recombinant PHAS-I by mTOR in vitro. Wild-type PHAS-I, but none of the mutant proteins, was coimmunoprecipitated with hemagglutinin-tagged raptor, an mTOR-associated protein, after extracts of cells overexpressing raptor had been supplemented with recombinant PHAS-I proteins. Moreover, raptor overexpression enhanced the phosphorylation of wild-type PHAS-I by mTOR but not the phosphorylation of the mutant proteins. The results not only provide direct evidence that both the RAIP and tor signaling motifs are important for the phosphorylation by mTOR, possibly by allowing PHAS-I binding to raptor, but also support the view that mTOR phosphorylates PHAS-I in cells.
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PMID:Two motifs in the translational repressor PHAS-I required for efficient phosphorylation by mammalian target of rapamycin and for recognition by raptor. 1266 11

Proteolysis, as well as protein synthesis, is a major process that contributes to the body protein turnover. Despite the huge variety of proteases in the body, there are very few proteolytic systems contributing to the complete hydrolysis of proteins to amino acids. The autophagic-lysosomal pathway is responsible for bulk proteolysis, whereas the ubiquitin-proteasome pathway plays a significant role in the fine control of the degradation of specific proteins. Both systems can produce free amino acids as a final product, but only the autophagy system is physiologically controlled by plasma amino acids. Recently, the study of amino acids as regulators of macromolecular turnover has been focused on for their signal transduction mechanism. In autophagic proteolysis, several amino acids have a direct regulatory potential: Leu, Gln, Tyr, Phe, Pro, Met, Trp and His in the liver, and Leu in the skeletal muscle. These amino acids are recognized at the plasma membrane, indicating the possible existence of an amino acid receptor/sensor for their recognition and subsequent intracellular signaling. Another line of evidence has emerged that protein kinase cascades such as mTOR, Erk, eIF2alpha etc. may be involved in the regulation of autophagy, and that amino acids, in combination with insulin, may exert their effects through these pathways. From the viewpoint of amino acid safety, the contribution of proteolysis to possible adverse effects caused by excessive amino acid intake is not clear. At present, there is one report that excess glutamine at 10-fold the plasma level has an abnormal inhibitory effect on hepatic proteolysis, due to a lysosomotropic toxicity of ammonia derived from glutamine degradation. Whether this may lead to an adverse effect in humans remains to be clarified.
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PMID:Amino acids as regulators of proteolysis. 1277 64

Although mTOR is a member of the PI-kinase-related kinase family, mTOR possesses serine-threonine protein kinase activities, which phosphorylate itself and exogenous substrates. mTOR autophosphorylates in vitro and is phosphorylated in vivo on serine residues. Ser2481, which is located in a His-Ser-Phe motif near the conserved carboxyl-terminal mTOR tail, has been reported as an autophosphorylation site in vivo and in vitro. The significance of the autophosphorylation remains unclear. Another phosphorylation site on mTOR in vivo is Ser2448. This site appears not to be an autophosphorylation site but a site potentially phosphorylated by protein kinase B (PKB). mTOR immunopurified from culture cells or tissues phosphorylates in vitro p70 S6 kinase (p70) alpha and p70beta, mainly on Thr412 or Thr401, respectively, located in a Phe-Thr-Tyr motif. Another exogenous substrate phosphorylated by immunopurified mTOR in vitro is eIF4E-binding protein 1 (4E-BP1) at sites corresponding to those phosphorylated in vivo during insulin stimulation in a Ser/Thr-Pro motif. Recently, raptor, a 150-kDa TOR-binding protein that contains a carboxyl-terminal WD-repeat domain, was discovered as a scaffold for the mTOR-catalyzed phosphorylation of 4E-BP1 and for the mTOR-mediated phosphorylation and activation of p70alpha. Other potential substrates phosphorylated by mTOR are nPKCdelta, nPKCepsilon, STAT3, and p53. The requirement of raptor for binding to and phosphorylation by mTOR of these potential substrates would clarify their physiological importance in the mTOR signaling pathway.
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PMID:Kinase activities associated with mTOR. 1456 Sep 63

The protein kinase B (PKB)/mammalian target of rapamycin (mTOR) signaling pathway is thought to play a critical role in the regulation of protein synthesis and skeletal muscle mass. The purpose of the present study was to determine the effects of voluntary wheel running on the PKB/mTOR signaling pathway in muscles from aged mice (20-22 months). The total levels of mTOR were 65% higher in gastrocnemius muscles from aged mice subjected to wheel running compared to aged sedentary mice (p-0.002) PKB phosphorlation on Ser473 was 45% higher in gastrocnemius muscles from aged mice subjected to wheel running compared to aged sedentary mice (p=0.01) The total abundance of PKB was 50% higher in gastrocnemius muscles from wheel running mice compared to aged controls (p=0.005). Three months of wheel running did not alter the total amount of p70 S6K in gastrocnemius muscle. Protein synthesis, as assessed by [(14)C]phenylalanine incorporation into protein was significantly higher in soleus muscles from aged mice subjected to wheel running compared to aged sedentary mice (p-0.001) These findings indicate the aerobic exercise training may attenuate the age-related decline in protein synthesis, a process that appears to be due, in part, to increases in mTOR and PKB.
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PMID:Effects of aerobic exercise training on the protein kinase B (PKB)/mammalian target of rapamycin (mTOR) signaling pathway in aged skeletal muscle. 1503 97

To determine the mechanism of meal-regulated synthesis of pancreatic digestive enzymes, we studied the effect of fasting and refeeding on pancreatic protein synthesis, relative mRNA levels of digestive enzymes, and activation of the translational machinery. With the use of the flooding dose technique with L-[3H]phenylalanine, morning protein synthesis in the pancreas of Institute for Cancer Research mice fed ad libitum was 7.9 +/- 0.3 nmol phenylalanine.10 min(-1).mg protein(-1). Prior fasting for 18 h reduced total protein synthesis to 70 +/- 1.4% of this value. Refeeding for 2 h, during which the mice consumed 29% of their daily food intake, increased protein synthesis to 117.3 +/- 4.9% of the control level. Pancreatic mRNA levels of amylase, lipases, trypsins, chymotrypsin, elastases, as well as those for several housekeeping genes tested were not significantly changed after refeeding compared with fasted mice. By contrast, the major translational control pathway involving Akt, mTOR, and S6K was strongly regulated by fasting and refeeding. Fasting for 18 h decreased phosphorylation of ribosomal protein S6 to almost undetectable levels, and refeeding highly increased it. The most highly phosphorylated form of the eIF4E binding protein (4E-BP1) made up the 14.6% of total 4E-BP1 in normally fed animals, was only 2.8% after fasting, and was increased to 21.4% after refeeding. This was correlated with an increase in the formation of the eIF4E-eIF4G complex after refeeding. By contrast, feeding did not affect eIF2B activity. Thus food intake stimulates pancreatic protein synthesis and translational effectors without increasing digestive enzyme mRNA levels.
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PMID:Feeding activates protein synthesis in mouse pancreas at the translational level without increase in mRNA. 1511 79

Hyperthyroidism increases metabolic rate, mitochondrial ATP production, and protein synthesis, but it remains to be determined whether all tissues and synthesis of specific protein pools are equally affected by hyperthyroidism. Previous studies showed that mitochondrial function was less responsive to elevated triiodothyronine (T(3)) levels in the low-oxidative plantaris muscle compared with other tissues in rats. We tested the hypothesis that in T(3)-treated animals mitochondrial protein synthesis would increase in oxidative but not glycolytic tissues. Male rats received either T(3) (200 mug/day, n = 10) or saline (controls, n = 9) by subcutaneous pump for 14 days, and then in vivo protein synthesis rates were measured using [(15)N]phenylalanine in liver, heart, plantaris, and red gastrocnemius (Red Gast). Mitochondrial protein synthesis rate in T(3)-treated rats was higher than in controls by 62% in Red Gast and plantaris and 89 and 115% in liver and heart, respectively (P < 0.01). Cytoplasmic protein synthesis rates in the T(3) group were 107-176% higher than control values (P < 0.01). There was also indirect evidence that protein breakdown was increased in all tissues of the T(3)-treated rats. Phosphorylation of selected regulators of protein synthesis in plantaris and Red Gast (mTOR, p70 S6 kinase, 4E-BP1), however, were not significantly affected by T(3). We conclude that T(3) infusion stimulates a general increase in mitochondrial and cytoplasmic protein synthesis rate among tissues and that this does not appear to explain the tissue-specific responses in mitochondrial oxidative capacity.
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PMID:Effect of T(3)-induced hyperthyroidism on mitochondrial and cytoplasmic protein synthesis rates in oxidative and glycolytic tissues in rats. 1704 59

Recent identification of the mammalian target of rapamycin (mTOR) pathway as an amino acid-sensing mechanism that regulates protein synthesis led us to investigate its role in rotavirus diarrhea. We hypothesized that malnutrition would reduce the jejunal protein synthetic rate and mTOR signaling via its target, ribosomal p70 S6 kinase (p70(S6K)). Newborn piglets were artificially fed from birth and infected with porcine rotavirus on day 5 of life. Study groups included infected (fully fed and 50% protein calorie malnourished) and noninfected fully fed controls. Initially, in "worst-case scenario studies," malnourished infected piglets were killed on days 1, 3, 5, and 11 postinoculation, and jejunal samples were compared with controls to determine the time course of injury and p70(S6K) activation. Using a 2 x 2 factorial design, we subsequently determined if infection and/or malnutrition affected mTOR activation on day 3. Western blot analysis and immunohistochemistry were used to measure total and phosphorylated p70(S6K); [(3)H]phenylalanine incorporation was used to measure protein synthesis; and lactase specific activity and villus-crypt dimensions were used to quantify injury. At the peak of diarrhea, the in vitro jejunal protein synthetic rate increased twofold (compared with the rate in the uninfected pig jejunum), concomitant with increased jejunal p70(S6K) phosphorylation (4-fold) and an increased p70(S6K) level (3-fold, P < 0.05). Malnutrition did not alter the magnitude of p70(S6K) activation. Immunolocalization revealed that infection produced a major induction of cytoplasmic p70(S6K) and nuclear phospho-p70(S6K), mainly in the crypt. A downregulation of semitendinosus muscle p70(S6K) phosphorylation was seen at days 1-3 postinoculation. In conclusion, intestinal activation of p70(S6K) was not inhibited by malnutrition but was strongly activated during an active state of mucosal regeneration.
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PMID:Intestinal ribosomal p70(S6K) signaling is increased in piglet rotavirus enteritis. 1713 69

The ability of the skeletal musculature to use amino acids to build or renew constitutive proteins is gradually lost with age and this is partly due to a decline in skeletal muscle insulin sensitivity. Since long-chain omega-3 polyunsaturated fatty acids (LCn-3PUFA) from fish oil are known to improve insulin-mediated glucose metabolism in insulin-resistant states, their potential role in regulating insulin-mediated protein metabolism was investigated in this study. Experimental data are based on a switchback design composed of three 5 week experimental periods using six growing steers to compare the effect of a continuous abomasal infusion of LCn-3PUFA-rich menhaden oil with an iso-energetic control oil mixture. Clamp and insulin signalling observations were combined with additional data from a second cohort of six steers. We found that enteral LCn-3PUFA potentiate insulin action by increasing the insulin-stimulated whole-body disposal of amino acids from 152 to 308 micromol kg(-1) h(-1) (P=0.006). The study further showed that in the fed steady-state, chronic adaptation to LCn-3PUFA induces greater activation (P<0.05) of the Akt-mTOR-S6K1 signalling pathway. Simultaneously, whole-body total flux of phenylalanine was reduced from 87 to 67 micromol kg(-1) h(-1) (P=0.04) and oxidative metabolism was decreased (P=0.05). We conclude that chronic feeding of menhaden oil provides a novel nutritional mean to enhance insulin-sensitive aspects of protein metabolism.
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PMID:Long-chain omega-3 fatty acids regulate bovine whole-body protein metabolism by promoting muscle insulin signalling to the Akt-mTOR-S6K1 pathway and insulin sensitivity. 1715 67


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