Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:6.1.1.4 (leucyl-tRNA synthetase)
 297 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Exposure of the temperature-sensitive leucyl-tRNA synthetase mutant of Chinese hamster ovary cells, tsH1, to the non-permissive temperature of 39.5 degrees C results in a rapid inhibition of polypeptide chain initiation. This inhibition is caused by a reduced ability of the eukaryotic initiation factor eIF-2 to participate in the formation of eIF-2.GTP.Met-tRNAf ternary complexes and thus in the formation of 43S ribosomal pre-initiation complexes. Associated with this decreased eIF-2 activity is an increased phosphorylation of the eIF-2 alpha subunit. It has previously been shown in other systems that phosphorylation of eIF-2 alpha slows the rate of recycling of eIF-2.GDP to eIF-2.GTP catalysed by the guanine nucleotide exchange factor eIF-2B. We show here that phosphorylation of eIF-2 alpha by the reticulocyte haem-controlled repressor also inhibits eIF-2B activity in cell-free extracts derived from tsH1 cells. Thus the observed increased phosphorylation of eIF-2 alpha at the non-permissive temperature in this system is consistent with impaired recycling of eIF-2 in vivo. Using a single-step temperature revertant of tsH1 cells, TR-3 (which has normal leucyl-tRNA synthetase activity at 39.5 degrees C), we demonstrate here that all inhibition of eIF-2 function reverts together with the synthetase mutation. This establishes the close link between synthetase function and eIF-2 activity. In contrast, recharging tRNALeu in vivo in tsH1 cells at 39.5 degrees C by treatment with a low concentration of cycloheximide failed to reverse the inhibition of eIF-2 function. This indicates that tRNA charging per se is not involved in the regulatory mechanism. Our data indicate a novel role for aminoacyl-tRNA synthetases in the regulation of eIF-2 function mediated through phosphorylation of the alpha subunit of this factor. However, in spite of the fact that cell-free extracts from Chinese hamster ovary cells contain protein kinase and phosphatase activities active against either exogenous or endogenous eIF-2 alpha, we have been unable to show any activation of kinase or inactivation of phosphatase following incubation of the cells at 39.5 degrees C.
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PMID:A novel role for aminoacyl-tRNA synthetases in the regulation of polypeptide chain initiation. 254 69

The effects of varying concentrations of ethanol on reactions involved in protein biosynthesis have been examined using a cell-free system from Chinese hamster ovary cells that actively translates natural mRNAs in order to detect those components most sensitive to alcohol. Ethanol, at relatively low concentrations (0.2 M or lower) inhibited the translation of endogenous polysomal mRNAs and, in mRNA-depleted extracts, of exogenous natural mRNA. Ethanol markedly inhibited leucyl-tRNA synthetase, and it inhibited Phe- and Glu-tRNA synthetases to some extent, but had only a small effect on several other aminoacyl-tRNA synthetases, elongation factors 1 and 2, ribosomes, or the formation of eukaryotic initiation factor 2 . GTP . Met-tRNAr ternary complex. Methanol inhibited slightly the translation of mRNA and Leu-tRNA synthetase, but isobutyl alcohol and isopropyl alcohol strongly depressed these activities. Ethanol inhibited the interaction of leucine with Leu-tRNA synthetase competitively, whereas isobutyl alcohol and acetaldehyde inhibited the leucine interaction in a noncompetitive manner. Leu-tRNA synthetase from Chinese hamster ovary cells was more sensitive to ethanol than that from yeast.
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PMID:Studies on the effect of ethanol on eukaryotic protein synthesis in vitro. 655 51

Signaling through mammalian target of rapamycin complex 1 (mTORC1) is stimulated by amino acids and insulin. Insulin inactivates TSC1/2, the GTPase-activator complex for Rheb, and Rheb.GTP activates mTORC1. It is not clear how amino acids regulate mTORC1. FKBP38 (immunophilin FK506-binding protein, 38 kDa), was recently reported to exert a negative effect on mTORC1 function that is relieved by its binding to Rheb.GTP. We confirm that Rheb binds wild type FKBP38, but inactive Rheb mutants showed contrasting abilities to bind FKBP38. We were unable to observe any regulation of FKBP38/mTOR binding by amino acids or insulin. Furthermore, FKBP38 did not inhibit mTORC1 signaling. The translationally controlled tumor protein (TCTP) in Drosophila was recently reported to act as the guanine nucleotide-exchange factor for Rheb. We have studied the role of TCTP in mammalian TORC1 signaling and its control by amino acids. Reducing TCTP levels did not reproducibly affect mTORC1 signaling in amino acid-replete/insulin-stimulated cells. Moreover, overexpressing TCTP did not rescue mTORC1 signaling in amino acid-starved cells. In addition, we were unable to see any stable interaction between TCTP and Rheb or mTORC1. Accumulation of uncharged tRNA has been previously proposed to be involved in the inhibition of mTORC1 signaling during amino acid starvation. To test this hypothesis, we used a Chinese hamster ovary cell line containing a temperature-sensitive mutation in leucyl-tRNA synthetase. Leucine deprivation markedly inhibited mTORC1 signaling in these cells, but shifting the cells to the nonpermissive temperature for the synthetase did not. These data indicate that uncharged tRNA(Leu) does not switch off mTORC1 signaling and suggest that mTORC1 is controlled by a distinct pathway that senses the availability of amino acids. Our data also indicate that, in the mammalian cell lines tested here, neither TCTP nor FKBP38 regulates mTORC1 signaling.
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PMID:Re-evaluating the roles of proposed modulators of mammalian target of rapamycin complex 1 (mTORC1) signaling. 1867 70

The GTP-bound form of elongation factor Tu (EF-Tu) brings aminoacylated tRNAs (aa-tRNA) to the A-site of the ribosome. EF-Tu binds all cognate elongator aa-tRNAs with highly similar affinities, and its weaker or tighter binding of misacylated tRNAs may discourage their participation in translation. Norvaline (Nva) is a non-proteinogenic amino acid that is activated and transferred to tRNALeu by leucyl-tRNA synthetase (LeuRS). No notable accumulation of Nva-tRNALeu has been observed in vitro, because of the efficient post-transfer hydrolytic editing activity of LeuRS. However, incorporation of norvaline into proteins in place of leucine does occur under certain conditions in vivo. Here we show that EF-Tu binds Nva-tRNALeu and Leu-tRNALeu with similar affinities, and that Nva-tRNALeu and Leu-tRNALeu dissociate from EF-Tu at comparable rates. The inability of EF-Tu to discriminate against norvaline may have driven evolution of highly efficient LeuRS editing as the main quality control mechanism against misincorporation of norvaline into proteins.
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PMID:Lack of discrimination against non-proteinogenic amino acid norvaline by elongation factor Tu from Escherichia coli. 2375 44

A protein synthesis enzyme, leucyl-tRNA synthetase (LRS), serves as a leucine sensor for the mechanistic target of rapamycin complex 1 (mTORC1), which is a central effector for protein synthesis, metabolism, autophagy, and cell growth. However, its significance in mTORC1 signaling and cancer growth and its functional relationship with other suggested leucine signal mediators are not well-understood. Here we show the kinetics of the Rag GTPase cycle during leucine signaling and that LRS serves as an initiating "ON" switch via GTP hydrolysis of RagD that drives the entire Rag GTPase cycle, whereas Sestrin2 functions as an "OFF" switch by controlling GTP hydrolysis of RagB in the Rag GTPase-mTORC1 axis. The LRS-RagD axis showed a positive correlation with mTORC1 activity in cancer tissues and cells. The GTP-GDP cycle of the RagD-RagB pair, rather than the RagC-RagA pair, is critical for leucine-induced mTORC1 activation. The active RagD-RagB pair can overcome the absence of the RagC-RagA pair, but the opposite is not the case. This work suggests that the GTPase cycle of RagD-RagB coordinated by LRS and Sestrin2 is critical for controlling mTORC1 activation, and thus will extend the current understanding of the amino acid-sensing mechanism.
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PMID:Coordination of the leucine-sensing Rag GTPase cycle by leucyl-tRNA synthetase in the mTORC1 signaling pathway. 2978 13