scholarly journals Elevated cJUN expression and an ATF/CRE site within the ATF3 promoter contribute to activation of ATF3 transcription by the amino acid response

2013 ◽  
Vol 45 (4) ◽  
pp. 127-137 ◽  
Author(s):  
Lingchen Fu ◽  
Michael S. Kilberg
Keyword(s):  
Amino Acid ◽  
Transcriptional Activation ◽  
Translational Control ◽  
Mammalian Cells ◽  
Transcriptional Control ◽  
Present Report ◽  
Binding Activity ◽  
Maximal Response ◽  
Protein Levels ◽  
Amino Acid Response

Mammalian cells respond to amino acid deprivation through multiple signaling pathways referred to as the amino acid response (AAR). Transcription factors mediate the AAR after their activation by several mechanisms; examples include translational control (activating transcription factor 4, ATF4), phosphorylation (p-cJUN), and transcriptional control (ATF3). ATF4 induces ATF3 transcription through a promoter-localized C/EBP-ATF response element (CARE). The present report characterizes an ATF/CRE site upstream of the CARE that also contributes to AAR-induced ATF3 transcription. ATF4 binds to the ATF/CRE and CARE sequences and both are required for a maximal response to ATF4 induction. ATF3, which antagonizes ATF4 and represses its own gene, also exhibited binding activity to the ATF/CRE and CARE sequences. The AAR resulted in elevated total cJUN and p-cJUN protein levels and both forms exhibited binding activity to the ATF/CRE and CARE ATF3 sequences. Knockdown of AAR-enhanced cJUN expression blocked induction of the ATF3 gene and mutation of either the ATF/CRE or the CARE site prevented the cJUN-dependent increase in ATF3-driven luciferase activity. The results indicate that both increased cJUN and the cis-acting ATF/CRE sequence within the ATF3 promoter contribute to the transcriptional activation of the gene during the AAR.

scholarly journals Hypoxia-inducible factor 1 levels vary exponentially over a physiologically relevant range of O2 tension

1996 ◽  
Vol 271 (4) ◽  
pp. C1172-C1180 ◽  
Author(s):  
B. H. Jiang ◽  
G. L. Semenza ◽  
C. Bauer ◽  
H. H. Marti
Keyword(s):  
Dna Binding ◽  
Transcriptional Activation ◽  
Mammalian Cells ◽  
Functional Characterization ◽  
Hypoxia Inducible Factor ◽  
Binding Activity ◽  
Maximal Response ◽  
Hypoxia Inducible Factor 1 ◽  
Dna Binding Activity

Hypoxia-inducible factor 1 (HIF-1) is a heterodimeric basic helix-loop-helix protein implicated in the transcriptional activation of genes encoding erythropoietin, glycolytic enzymes, and vascular endothelial growth factor in hypoxic mammalian cells. In this study, we have quantitated HIF-1 DNA-binding activity and protein levels of the HIF-1 alpha and HIF-1 beta subunits in human HeLa cells exposed to O2 concentrations ranging from 0 to 20% in the absence or presence of 1 mM KCN to inhibit oxidative phosphorylation and cellular O2 consumption. HIF-1 DNA-binding activity, HIF-1 alpha protein and HIF-1 beta protein each increased exponentially as cells were subjected to decreasing O2 concentrations, with a half maximal response between 1.5 and 2% O2 and a maximal response at 0.5% O2, both in the presence and absence of KCN. The HIF-1 response was greatest over O2 concentrations associated with ischemic/hypoxic events in vivo. These results provide evidence for the involvement of HIF-1 in O2 homeostasis and represent a functional characterization of the putative O2 sensor that initiates hypoxia signal transduction leading to HIF-1 expression.

scholarly journals Aminoacyl-tRNA synthetase inhibition activates a pathway that branches from the canonical amino acid response in mammalian cells

2020 ◽  
Vol 117 (16) ◽  
pp. 8900-8911 ◽  
Author(s):  
Yeonjin Kim ◽  
Mark S. Sundrud ◽  
Changqian Zhou ◽  
Maja Edenius ◽  
Davide Zocco ◽  
...  
Keyword(s):  
Amino Acid ◽  
Mammalian Cells ◽  
Inflammatory Responses ◽  
Mammalian Target Of Rapamycin ◽  
Cell Types ◽  
Trna Synthetase ◽  
Aminoacyl Trna Synthetase ◽  
Therapeutic Benefits ◽  
Fibroblast Like Synoviocytes ◽  
Amino Acid Response

Signaling pathways that sense amino acid abundance are integral to tissue homeostasis and cellular defense. Our laboratory has previously shown that halofuginone (HF) inhibits the prolyl-tRNA synthetase catalytic activity of glutamyl-prolyl-tRNA synthetase (EPRS), thereby activating the amino acid response (AAR). We now show that HF treatment selectively inhibits inflammatory responses in diverse cell types and that these therapeutic benefits occur in cells that lack GCN2, the signature effector of the AAR. Depletion of arginine, histidine, or lysine from cultured fibroblast-like synoviocytes recapitulates key aspects of HF treatment, without utilizing GCN2 or mammalian target of rapamycin complex 1 pathway signaling. Like HF, the threonyl-tRNA synthetase inhibitor borrelidin suppresses the induction of tissue remodeling and inflammatory mediators in cytokine-stimulated fibroblast-like synoviocytes without GCN2, but both aminoacyl-tRNA synthetase (aaRS) inhibitors are sensitive to the removal of GCN1. GCN1, an upstream component of the AAR pathway, binds to ribosomes and is required for GCN2 activation. These observations indicate that aaRS inhibitors, like HF, can modulate inflammatory response without the AAR/GCN2 signaling cassette, and that GCN1 has a role that is distinct from its activation of GCN2. We propose that GCN1 participates in a previously unrecognized amino acid sensor pathway that branches from the canonical AAR.

scholarly journals Characterization of the amino acid response element within the human sodium-coupled neutral amino acid transporter 2 (SNAT2) System A transporter gene

2006 ◽  
Vol 395 (3) ◽  
pp. 517-527 ◽  
Author(s):  
Stela S. Palii ◽  
Michelle M. Thiaville ◽  
Yuan-Xiang Pan ◽  
Can Zhong ◽  
Michael S. Kilberg
Keyword(s):  
Amino Acid ◽  
Rna Polymerase Ii ◽  
Mammalian Cells ◽  
Amino Acid Transporter ◽  
Neutral Amino Acid ◽  
System A ◽  
Neutral Amino Acid Transporter ◽  
Acid Transporter ◽  
Amino Acid Response

The neutral amino acid transport activity, System A, is enhanced by amino acid limitation of mammalian cells. Of the three gene products that encode System A activity, the one that exhibits this regulation is SNAT2 (sodium-coupled neutral amino acid transporter 2). Fibroblasts that are deficient in the amino acid response pathway exhibited little or no induction of SNAT2 mRNA. Synthesis of SNAT2 mRNA increased within 1–2 h after amino acid removal from HepG2 human hepatoma cells. The amino acid responsive SNAT2 genomic element that mediates the regulation has been localized to the first intron. Increased binding of selected members of the ATF (activating transcription factor) and C/EBP (CCAAT/enhancer-binding protein) families to the intronic enhancer was established both in vitro and in vivo. In contrast, there was no significant association of these factors with the SNAT2 promoter. Expression of exogenous individual ATF and C/EBP proteins documented that specific family members are associated with either activation or repression of SNAT2 transcription. Chromatin immunoprecipitation analysis established in vivo that amino acid deprivation led to increased RNA polymerase II recruitment to the SNAT2 promoter.

scholarly journals Role of hydrophobic amino acid clusters in the transactivation activity of the human glucocorticoid receptor.

1997 ◽  
Vol 17 (2) ◽  
pp. 934-945 ◽  
Author(s):  
T Almlöf ◽  
J A Gustafsson ◽  
A P Wright
Keyword(s):  
Amino Acid ◽  
Glucocorticoid Receptor ◽  
Transcriptional Activation ◽  
Mammalian Cells ◽  
Alpha Helix ◽  
Transactivation Domain ◽  
Hydrophobic Residues ◽  
Transactivation Activity ◽  
Reduced Activity ◽  
Helical Region

We have performed a mutagenesis analysis of the 58-amino-acid tau1-core peptide, which represents the core transactivation activity of the tau1 transactivation domain from the glucocorticoid receptor. Mutants with altered activity were identified by phenotypic screening in the yeast Saccharomyces cerevisiae. Most mutants with reduced activity had substitutions of hydrophobic amino acids. Most single-substitution mutants with reduced activity were localized near the N terminus of the tau1-core within a segment that has been shown previously to have a propensity for alpha-helix conformation, suggesting that this helical region is of predominant importance. The particular importance of hydrophobic residues within this region was confirmed by comparing the activities of alanine substitutions of the hydrophobic residues in this and two other helical regions. The hydrophobic residues were shown to be important for the transactivation activity of both the isolated tau1-core and the intact glucocorticoid receptor in mammalian cells. Rare mutations in helical regions I and II gave rise to increased transcriptional activation activity. These mutations increase the hydrophobicity of hydrophobic patches on each of these helices, suggesting a relationship between the hydrophobicity of the patches and transactivation activity. However, certain nonhydrophobic residues are also important for activity. Interestingly, helical region I partially matches a consensus motif found in the retinoic acid receptor, VP16, and several other activator proteins.

scholarly journals A feedback transcriptional mechanism controls the level of the arginine/lysine transporter cat-1 during amino acid starvation

2007 ◽  
Vol 402 (1) ◽  
pp. 163-173 ◽  
Author(s):  
Alex B. Lopez ◽  
Chuanping Wang ◽  
Charlie C. Huang ◽  
Ibrahim Yaman ◽  
Yi Li ◽  
...  
Keyword(s):  
Amino Acid ◽  
Transcriptional Activation ◽  
Transcriptional Repression ◽  
Mammalian Cells ◽  
Leucine Zipper ◽  
Amino Acid Starvation ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Basic Leucine Zipper

The adaptive response to amino acid limitation in mammalian cells inhibits global protein synthesis and promotes the expression of proteins that protect cells from stress. The arginine/lysine transporter, cat-1, is induced during amino acid starvation by transcriptional and post-transcriptional mechanisms. It is shown in the present study that the transient induction of cat-1 transcription is regulated by the stress response pathway that involves phosphorylation of the translation initiation factor, eIF2 (eukaryotic initiation factor-2). This phosphorylation induces expression of the bZIP (basic leucine zipper protein) transcription factors C/EBP (CCAAT/enhancer-binding protein)-β and ATF (activating transcription factor) 4, which in turn induces ATF3. Transfection experiments in control and mutant cells, and chromatin immunoprecipitations showed that ATF4 activates, whereas ATF3 represses cat-1 transcription, via an AARE (amino acid response element), TGATGAAAC, in the first exon of the cat-1 gene, which functions both in the endogenous and in a heterologous promoter. ATF4 and C/EBPβ activated transcription when expressed in transfected cells and they bound as heterodimers to the AARE in vitro. The induction of transcription by ATF4 was inhibited by ATF3, which also bound to the AARE as a heterodimer with C/EBPβ. These results suggest that the transient increase in cat-1 transcription is due to transcriptional activation caused by ATF4 followed by transcriptional repression by ATF3 via a feedback mechanism.

scholarly journals Marked Variation in Response of Consensus Binding Elements for the Rta Protein of Epstein-Barr Virus

2005 ◽  
Vol 79 (15) ◽  
pp. 9635-9650 ◽  
Author(s):  
Lee-Wen Chen ◽  
Pey-Jium Chang ◽  
Henri-Jacques Delecluse ◽  
George Miller
Keyword(s):  
Dna Sequences ◽  
Transcriptional Activation ◽  
Mammalian Cells ◽  
Epstein Barr Virus ◽  
Lytic Cycle ◽  
Maximal Response ◽  
Binding Affinities ◽  
Barr Virus ◽  
Epstein Barr

ABSTRACT The R transactivator (Rta) protein activates Epstein-Barr virus (EBV) lytic-cycle genes by several distinct mechanisms that include direct binding to viral promoters, synergy with BamHI Z EBV replication activator (ZEBRA), and activation of cellular signaling pathways. In the direct and synergistic mechanisms of action, Rta binds to specific DNA sequences that are present in the promoters of responsive genes. It has been difficult to demonstrate the capacity of Rta expressed in mammalian cells to bind DNA in vitro in order to study the relative affinities of Rta binding elements. We discovered that a short C-terminal region of Rta inhibits the ability of Rta to bind DNA in vitro. C-terminally truncated versions of Rta bind DNA efficiently and thus facilitate a comparison of consensus Rta binding elements (CRBEs) found in promoters of five Rta-responsive genes: BMLF1, BHLF1, BMRF1, BaRF1, and BLRF2. All CRBEs in the promoters of the five genes conform to the proposed recognition sequence GNCCN9GGNG, where N is any nucleotide and N9 represents a sequence of nine nucleotides. Nonetheless, CRBEs varied markedly in their abilities to bind Rta in electrophoretic mobility shift assays. Not all CRBEs bound or responded to Rta. Binding affinities of the CRBEs and the capacity to be activated by Rta in reporter assays were strongly correlated. The CRBEs from the BMLF1 and BHLF1 promoters conferred the greatest response. The response of the BMRF1, BaRF1, and BLRF2 CRBEs was less robust. By creation of chimeras, inversions, and point mutations, differences in binding affinities and transcriptional activation levels could be attributed to N9 sequence variation. The length of N9 was also critical for a maximal response. In Raji and BZLF1-knockout cells, the mRNAs of the five Rta-responsive lytic-cycle genes differed dramatically in kinetics of expression, abundance, and synergistic responses to ZEBRA and Rta. Affinities of Rta response elements for Rta are likely to play an important role in temporal regulation and the level of lytic-cycle EBV gene expression.

scholarly journals The Transcription Factor Network Associated With the Amino Acid Response in Mammalian Cells

2012 ◽  
Vol 3 (3) ◽  
pp. 295-306 ◽  
Author(s):  
Michael S. Kilberg ◽  
Mukundh Balasubramanian ◽  
Lingchen Fu ◽  
Jixiu Shan
Keyword(s):  
Transcription Factor ◽  
Amino Acid ◽  
Mammalian Cells ◽  
Transcription Factor Network ◽  
Amino Acid Response

scholarly journals Total 4EBP1 Is Elevated in Liver of Rats in Response to Low Sulfur Amino Acid Intake

2013 ◽  
Vol 2013 ◽  
pp. 1-11 ◽  
Author(s):  
Angelos K. Sikalidis ◽  
Kevin M. Mazor ◽  
Minji Kang ◽  
Hongyun Liu ◽  
Martha H. Stipanuk
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Dietary Treatment ◽  
Binding Activity ◽  
Initiation Factor ◽  
Sulfur Amino Acid ◽  
Sulfur Amino Acids ◽  
Protein Levels ◽  
Cellular Stresses

Translation initiation is known to be regulated by the binding of eukaryotic initiation factor 4E (eIF4E) by binding proteins (4EBPs), and there is evidence that amino acid deprivation and other cellular stresses upregulate 4EBP1 expression. To pursue the question of whether diets limited in an essential amino acid lead to induction of 4EBP1 expression in vivo, diets that varied in methionine and cystine content were fed to rats for 7 days, and 4EBP1 mRNA and protein levels and 4EBP1 phosphorylation state were determined. Total 4EBP1 mRNA and protein abundance increased in liver of rats with severely deficient intakes of sulfur amino acids (0.23% or 0.11% methionine without cystine) but not in animals with a less restricted intake of sulfur amino acids (0.11% methionine plus 0.35% cystine) but a similarly restricted intake of total diet (53 to 62% of control). The amount of 4EBP1 binding activity (α + β forms) was elevated in liver of rats fed sulfur amino acid-deficient diets, whereas the hyperphosphorylation of 4EBP1 was not affected by dietary treatment. Results suggest that changes in total 4EBP1 expression should be considered when examining mechanisms that attenuate protein synthesis during amino acid deficiency states.

Sign in / Sign up

Export Citation Format

Share Document