Leucine-Specific Binding of Photoreactive Leu7-MAP to a High Molecular Weight Protein on the Plasma Membrane of the Isolated Rat Hepatocyte

Mortimore, Glenn E.; Wert, Jr Jj; Miotto, Giovanni; Venerando, Rina; Kadowaki, Motoni

doi:10.1006/bbrc.1994.2168

Cited by 35 publications

(12 citation statements)

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“…The correspondence in the time course of the response of synthesis to the change in plasma amino acid concentrations further supports the regulatory role of the plasma, as opposed to intracellular, concentrations. The mechanism whereby changes in plasma concentrations could signal the intracellular synthetic process is unclear, but it has been shown that there is a leucine-specific binding protein on the plasma membrane of isolated hepatocytes that regulates intracellular protein metabolism (17). Perhaps a similar protein exists on muscle cell membranes that transmits signals to the intracellular compartment.…”

Section: Discussionmentioning

confidence: 99%

Reduced amino acid availability inhibits muscle protein synthesis and decreases activity of initiation factor eIF2B

Kobayashi¹,

Børsheim²,

Anthony

et al. 2003

American Journal of Physiology-Endocrinology and Metabolism

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Reduced amino acid availability inhibits muscle protein synthesis and decreases activity of initiation factor eIF2B. Am J Physiol Endocrinol Metab 284: E488-E498, 2003; 10.1152/ ajpendo.00094.2002.-We have examined the effect of a hemodialysis-induced 40% reduction in plasma amino acid concentrations on rates of muscle protein synthesis and breakdown in normal swine. Muscle protein kinetics were measured by tracer methodology using [ 2 H5]phenylalanine and [1-13 C]leucine and analysis of femoral arterial and venous samples and tissue biopsies. Net amino acid release by muscle was accelerated during dialysis. Phenylalanine utilization for muscle protein synthesis was reduced from the basal value of 45 Ϯ 8 to 25 Ϯ 6 nmol ⅐ min Ϫ1 ⅐ 100 ml leg Ϫ1between 30 and 60 min after start of dialysis and was stimulated when amino acids were replaced while dialysis continued. Muscle protein breakdown was unchanged. The signal for changes in synthesis appeared to be changes in plasma amino acid concentrations, as intramuscular concentrations remained constant throughout. The changes in muscle protein synthesis were accompanied by a reduction or stimulation, respectively, in the guanine nucleotide exchange activity of eukaryotic initiation factor (eIF)2B following hypoaminoacidemia vs. amino acid replacement. We conclude that a reduction in plasma amino acid concentrations below the normal basal value signals an inhibition of muscle protein synthesis and that corresponding changes in eIF2B activity suggest a possible role in mediating the response. (2). Therefore, the principal goal of this study was to examine the effect of a reduction in plasma amino acid concentrations below the normal postabsorptive levels on the rates of muscle protein synthesis and breakdown. We used hemodialysis to reduce amino acid concentration. A second goal of our study was to attempt to identify the signal for changes in the rate of synthesis or breakdown when the plasma concentrations of amino acids drop. Whereas it is reasonable to presume that effects of amino acid availability would be mediated by changes in intracellular availability, our previous studies have shown remarkably small changes in intracellular concentrations despite significant changes in plasma concentrations (8,22). However, in previous studies, intracellular samples were obtained in physiologically steady-state circumstances, usually after 2 h or more of continuous infusion of amino acids (e.g., Ref. 8). Thus, in this study, we sampled frequently from the femoral artery and vein, as well as from muscle tissue, as changes in concentration were induced by dialysis. We hoped that a more detailed characterization of the time course of changes in concentrations, synthesis, and breakdown would enable us to gain insight regarding the signal for changes in muscle protein synthesis and breakdown.A third goal of our study was to investigate potential mechanisms by which a reduction in plasma amino acid concentrations might mediate a change in muscle protein synthesis. For this purpose...

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Section: Discussionmentioning

confidence: 99%

Reduced amino acid availability inhibits muscle protein synthesis and decreases activity of initiation factor eIF2B

Kobayashi¹,

Børsheim²,

Anthony

et al. 2003

American Journal of Physiology-Endocrinology and Metabolism

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“…Moreover, we investigated the possible involvement of System L as a possible recognition site of leucine, but this possibility was not feasible. Although a putative leucine-binding protein (s) has not yet been identified, a candidate was detected using a photoreactive derivative of Leu 8 -MAP (15). Recently, it was reported that in yeast, Ssy1p, a homologue of amino acid permease, has the ability to act as an amino acid "sensor," although it has a broad specificity for amino acids (51).…”

Section: Fig 6 Effect Of Leu 8 -Map On Autophagic Proteolysis and P70mentioning

confidence: 99%

“…These findings represented indirect evidence, but strongly suggested the possibility that, for autophagic regulation, amino acids have their own recognition sites at the cell surface and must have a signal transduction mechanism to the site of autophagosome formation inside the cell. A possible candidate for the leucine "receptor/sensor" protein has been reported (15), and its purification has progressed. 1 Concerning the intracellular signaling pathways, possibilities have been raised by several groups.…”

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confidence: 99%

Amino Acids and Insulin Control Autophagic Proteolysis through Different Signaling Pathways in Relation to mTOR in Isolated Rat Hepatocytes

Kanazawa

Taneike

Akaishi

et al. 2004

Journal of Biological Chemistry

195

143

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Autophagy, a major bulk proteolytic pathway, contributes to intracellular protein turnover, together with protein synthesis. Both are subject to dynamic control by amino acids and insulin. The mechanisms of signaling and cross-talk of their physiological anabolic effects remain elusive. Recent studies established that amino acids and insulin induce p70 S6 kinase (p70 S6k ) phosphorylation by mTOR, involved in translational control of protein synthesis. Here, the signaling mechanisms of amino acids and insulin in macroautophagy in relation to mTOR were investigated. In isolated rat hepatocytes, both regulatory amino acids (RegAA) and insulin coordinately activated p70 S6k phosphorylation, which was completely blocked by rapamycin, an mTOR inhibitor. However, rapamycin blocked proteolytic suppression by insulin, but did not block inhibition by RegAA. These contrasting results suggest that insulin controls autophagy through the mTOR pathway, but amino acids do not. Furthermore, micropermeabilization with Saccharomyces aureus ␣-toxin completely deprived hepatocytes of proteolytic responsiveness to RegAA and insulin, but still maintained p70 S6k phosphorylation by RegAA. In contrast, Leu 8 -MAP, a non-transportable leucine analogue, did not mimic the effect of leucine on p70 S6k phosphorylation, but maintained the activity on proteolysis. Finally, BCH, a System L-specific amino acid, did not affect proteolytic suppression or mTOR activation by leucine. All the results indicate that mTOR is not common to the signaling mechanisms of amino acids and insulin in autophagy, and that the amino acid signaling starts extracellularly with their "receptor(s)," probably other than transporters, and is mediated through a novel route distinct from the mTOR pathway employed by insulin.

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“…A nontransportable multiple antigen peptide derivative constructed by attaching eight leucine residues to a lysine core (Leu 8 -MAP) was effective in suppressing deprivation-induced macroautophagy in isolated hepatocytes with an apparent K m equivalent to that of leucine (52). Furthermore, photoaffinity-labeling experiments revealed that the putative Leu 8 -MAP substrate was a protein of approximately 340,000 M r and was enriched within membrane-fractions, suggesting that this factor may be a plasma membrane receptor (53). In light of these data, it is tempting to speculate that leucine-(and/or regulatory amino acid-) specific signals not only regulate hepatic macroautophagic proteolysis, eIF4F assembly, and p70…”

Section: A Determination Of [mentioning

confidence: 99%

Leucine, Glutamine, and Tyrosine Reciprocally Modulate the Translation Initiation Factors eIF4F and eIF2B in Perfused Rat Liver

Shah

Antonetti

Kimball

et al. 1999

Journal of Biological Chemistry

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Leucine, glutamine, and tyrosine, three amino acids playing key modulatory roles in hepatic proteolysis, were evaluated for activation of signaling pathways involved in regulation of liver protein synthesis. Furthermore, because leucine signals to effectors that lie distal to the mammalian target of rapamycin, these downstream factors were selected for study as candidate mediators of amino acid signaling. Using the perfused rat liver as a model system, we observed a 25% stimulation of protein synthesis in response to balanced hyperaminoacidemia, whereas amino acid imbalance due to elevated concentrations of leucine, glutamine, and tyrosine resulted in a protein synthetic depression of roughly 50% compared with normoaminoacidemic controls. The reduction in protein synthesis accompanying amino acid imbalance became manifest at high physiologic concentrations and was dictated by the guanine nucleotide exchange activity of translation initiation factor eIF2B. Paradoxically, this phenomenon occurred concomitantly with assembly of the mRNA cap recognition complex, eIF4F as well as activation of the 70-kDa ribosomal S6 kinase, p70 S6k. Dual and reciprocal modulation of eIF4F and eIF2B was leucine-specific because isoleucine, a structural analog, was ineffective in these regards. Thus, we conclude that amino acid imbalance, heralded by leucine, initiates a liver-specific translational failsafe mechanism that deters protein synthesis under unfavorable circumstances despite promotion of the eIF4F complex.The amino acids represent a class of biologic molecules exerting dynamic and complex influences on highly disparate physiologic processes including pancreatic insulin and glucagon secretion, protein degradation and synthesis, hepatic gluconeogenesis, and sensitization of tissues to the anabolic effects of insulin (1-5). The effects of amino acids are somewhat enigmatic but often involve the interplay of hormones and other factors intrinsic to the cellular environment. Recently, the branched chain amino acid, leucine, has been demonstrated to modulate pathways of signal transduction and may indeed contribute importantly to the cellular interpretation of integrated signals. With regard to protein homeostasis, several reports now exist supporting the hypothesis that leucine impacts protein turnover through mechanisms beyond those of protein synthetic substrates (1-4).In an elegant series of experiments, Mortimore et al. (4,5) demonstrated that the amino acids leucine, glutamine, and tyrosine, individually as well as cooperatively and in a manner that is concentration-dependent, attenuate hepatic macroautophagic proteolysis induced by deprivation of amino acids. Furthermore, insulin functions additively and synergistically with these amino acids, thereby enhancing the efficacy of proteolytic inhibition by leucine, glutamine, and/or tyrosine. Inherent in their potency as modulators of protein homeostasis, amino acids generally exert reciprocal control of hepatic protein degradation and synthesis. The latter process is governed...

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Leucine-Specific Binding of Photoreactive Leu7-MAP to a High Molecular Weight Protein on the Plasma Membrane of the Isolated Rat Hepatocyte

Cited by 35 publications

References 0 publications

Reduced amino acid availability inhibits muscle protein synthesis and decreases activity of initiation factor eIF2B

Reduced amino acid availability inhibits muscle protein synthesis and decreases activity of initiation factor eIF2B

Amino Acids and Insulin Control Autophagic Proteolysis through Different Signaling Pathways in Relation to mTOR in Isolated Rat Hepatocytes

Leucine, Glutamine, and Tyrosine Reciprocally Modulate the Translation Initiation Factors eIF4F and eIF2B in Perfused Rat Liver

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