Effects of single amino acid deficiency on mRNA translation are markedly different for methionine versus leucine (original) (raw)
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Amino acids as regulators of gene expression at the level of mRNA translation
The Journal of nutrition, 2003
Amino acids act through a number of signaling pathways and mechanisms to mediate control of gene expression at the level of mRNA translation. This report reviews recent findings that illustrate the manner through which amino acids act to regulate the initiation phase of mRNA translation. The report focuses on signaling pathways that involve the eukaryotic initiation factor-2 (eIF2) protein kinase, general control non-derepressing kinase-2 and the mammalian target of rapamycin (mTOR) protein kinase. It also describes the mechanisms through which amino acid-induced modulation of eIF2 phosphorylation and mTOR-mediated signaling cause derepression of translation of specific mRNAs and result in an overall change in the pattern of gene expression. Finally, it provides examples of mRNAs whose translation is modulated through these mechanisms.
Journal of Biological Chemistry, 1999
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.
Leucine regulates translation initiation in rat skeletal muscle via enhanced eIF4G phosphorylation
The Journal of nutrition, 2004
The BCAA, leucine, stimulates protein synthesis in skeletal muscle in part through enhanced initiation of mRNA translation. However, understanding how leucine regulates protein synthesis remains elusive. The intent of the present investigation was to examine the effect of leucine, independent of other regulatory agents, on key events in translation initiation in skeletal muscle and to elucidate the extent to which signaling through the mammalian target of rapamycin (mTOR) accounts for the effect of the amino acid on protein synthesis. Hindlimb preparations from postabsorptive rats were perfused with medium containing food-deprived (1X) or superphysiologic (10X) concentrations of leucine with all other amino acids at 1X concentration. Protein synthesis was significantly greater in both gastrocnemius and soleus perfused with 10X compared with 1X leucine. The stimulatory effects of leucine on protein synthesis were unaffected by a specific inhibitor of PI3-kinase (LY 294002). Moreover,...
2000
The objective of the current study was to examine the role of the branched-chain amino acid (BCAA) leucine in the regulation of hepatic protein synthesis and ribosomal protein (rp) mRNA translation in vivo. Food-deprived (18 h) male rats (200 g) were orally administered saline (control) or 270 mg leucine, isoleucine or valine and killed 1 h later. Administration of any BCAA resulted in enhanced phosphorylation of eukaryotic initiation factor (eIF) 4E-binding protein-1 (4E-BP1) compared with controls. However, leucine was the most effective at stimulating phosphorylation of 4E-BP1 as well as the 70-kDa ribosomal protein S6 kinase (S6K1). Despite these effects on components of the translation initiation process, there were no differences in total protein synthesis rates among treatment groups. The distribution of rp (S4, S8, L26) and non-rp (albumin, -actin) mRNAs across sucrose density gradients showed that the preponderance of hepatic rp mRNAs in control rats was unloaded from polysomes. Of the BCAA, only leucine was the most effective in causing a shift in the distribution of rp mRNA to polysomes compared with controls. Non-rp transcripts remained mainly polysome-associated under all conditions. These results suggest that leucine is most effective among the BCAA in its ability to stimulate translation of rp mRNA in liver. Furthermore, the translation of rp mRNA is disjointed from rates of total protein synthesis in liver and related to the degree of S6K1 phosphorylation. . 3 Abbreviations used: BCAA, branched-chain amino acids; Con, food-deprived rats; 4E-BP1, eIF4E-binding protein 1; eIF, eukaryotic initiation factor; Ile, food-deprived rats orally administered 270 mg isoleucine; Leu, food-deprived rats orally administered 270 mg leucine; rp, ribosomal protein; S6K1, 70-kDa ribosomal protein S6 kinase; TOP, terminal oligopyrimidine tract; Val, food-deprived rats orally administered 270 mg valine.
New functions for amino acids: effects on gene transcription and translation
The American journal of clinical nutrition, 2006
Amino acids act to regulate multiple processes related to gene expression, including modulation of the function of the proteins that mediate messenger RNA (mRNA) translation. By modulating the function of translation initiation and elongation factors, amino acids regulate the translation of mRNA on a global scale and also act to cause preferential changes in the translation of mRNAs encoding particular proteins or families of proteins. However, amino acids do not directly regulate the function of translation initiation and elongation factors, but instead modulate signaling through pathways traditionally considered to be solely involved in mediating the action of hormones. The best-characterized example of amino acid-induced regulation of a signal transduction pathway is one involving a protein kinase referred to as the mammalian target of rapamycin (mTOR), through which the branched-chain amino acids, particularly leucine, act to modulate the function of proteins engaged in both glo...
Translational Dysregulation by Pateamine A
2007
Pateamine A inhibits translation by preventing proper translational initiation complex formation. In the December issue of Chemistry & Biology, Bordeleau et al. demonstrated that the effects of Patemine A on translation are mediated through the interaction between the RNA helicase eIF4A and mRNA [1].
Nucleic acids research, 2017
Certain chemical modifications confer increased stability and low immunogenicity to in vitro transcribed mRNAs, thereby facilitating expression of therapeutically important proteins. Here, we demonstrate that N1-methyl-pseudouridine (N1mΨ) outperforms several other nucleoside modifications and their combinations in terms of translation capacity. Through extensive analysis of various modified transcripts in cell-free translation systems, we deconvolute the different components of the effect on protein expression independent of mRNA stability mechanisms. We show that in addition to turning off the immune/eIF2α phosphorylation-dependent inhibition of translation, the incorporated N1mΨ nucleotides dramatically alter the dynamics of the translation process by increasing ribosome pausing and density on the mRNA. Our results indicate that the increased ribosome loading of modified mRNAs renders them more permissive for initiation by favoring either ribosome recycling on the same mRNA or de...
Biochem Biophys Res Commun, 2006
Branched-chain amino acid leucine has been shown to activate the translational regulators through the mammalian target of rapamycin. However, the leucine's effects are self-limiting because leucine promotes its own disposal by an oxidative pathway. The irreversible and rate-limiting step in the leucine oxidation pathway is catalyzed by the branched-chain a-keto acid dehydrogenase (BCKDH) complex. The complex contains E1 (a2b2), E2, and E3 subunits, and its activity is abolished by phosphorylation of the E1a subunit by BCKDH kinase. The relationship between the activity of BCKDH complex and leucine-mediated activation of the protein translation was investigated using the technique of RNA interference. The activity of BCKDH complex in C2C12 cell was modulated by transfection of small interfering RNA (siRNA) for BCKDH E2 subunit or BCKDH kinase. Transfection of siRNAs decreased the mRNA expression and protein amount of corresponding gene. Suppression of either E2 subunit or kinase produced opposite effects on the cell proliferation and the activation of translational regulators by leucine. Suppression of BCKDH kinase for 48 h resulted in decreasing cell proliferation. In contrast, E2 suppression led to increased amount of total cellular protein. The phosphorylation of p70 S6 kinase by leucine was increased in E2-siRNA transfected C2C12 cells, whereas the leucine's effect was diminished in kinase-siRNA transfected cells. These results suggest that the activation of the translational regulators by leucine was partly regulated by the activity of BCKDH complex.
Molecular & Cellular Proteomics, 2011
We describe a method to identify and analyze translationally regulative 5'UTRs (5'TRU) in Saccharomyces cerevisiae. Two-dimensional analyses of (35)S-methionine metabolically labeled cells revealed 13 genes and proteins, whose protein biosynthesis is post-transcriptionally up-regulated on amino acid starvation. The 5'UTRs of the respective mRNAs were further investigated. A plasmid-based reporter-testing system was developed to analyze their capability to influence translation dependent on amino acid availability. Most of the 13 candidate 5'UTRs are able to enhance translation independently of amino acids. Two 5'UTRs generally repressed translation, and the 5'UTRs of ENO1, FBA1, and TPI1 specifically up-regulated translation when cells were starved for amino acids. The TPI1-5'UTR exhibited the strongest effect in the testing system, which is consistent with elevated Tpi1p-levels in amino acid starved cells. Bioinformatical analyses support that an unstructured A-rich 5' leader is beneficial for efficient translation when amino acids are scarce. Accordingly, the TPI1-5'UTR was shown to contain an A-rich tract in proximity to the mRNA-initiation codon, required for its amino acid dependent regulatory function.
The role of mammalian initiation factor eIF-4D and its hypusine modification in translation
Biochimica Et Biophysica Acta Gene Structure and Expression, 1990
Initiation factor eIF-4D functions late in the initiation pathway, apparently daring formation of the first peptide bond. The factor is post-translationally me.lied at a specific lysine residue by reaction with spermidine and subsequent hydroxylation to form hypusine. A precursor form lacking hypusine is inactive in the assay for methionyl-puromycin synthesis, but activity is restored following in vitro modification to deoxyhypusine, thereby suggesting that the modification is essential for function. Since formylated methionyl-tRNA is less dependent on eW-4D in the puromycin assay, we postulate that elF-4D and its hylmsine modification may stabilize charged Met-tRNA binding to the peptldyi transferase center of the 60S ribosomal subunit. Analysis of elF-4D genes in yeast indicate that elF-4D and its hypusine modification are essential for cell growth.