A MYC/GCN2/eIF2alpha negative feedback loop limits protein synthesis to prevent MYC-dependent apoptosis in colorectal cancer
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA418238 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR6288001.fastq.gz | Fastqsanger.gz | |||
| SRR6288002.fastq.gz | Fastqsanger.gz | |||
| SRR6288003.fastq.gz | Fastqsanger.gz | |||
| SRR6288004.fastq.gz | Fastqsanger.gz | |||
| SRR6288005.fastq.gz | Fastqsanger.gz |
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Project description:Tumours depend on altered rates of protein synthesis for growth and survival, suggesting that mechanisms controlling mRNA translation may be exploitable for therapy. Here, we show that loss of APC, which occurs almost universally in colorectal tumours, strongly enhances the dependence on the translation initiation factor eIF2B5. Depletion of eIF2B5 induces an integrated stress response and enhances translation of MYC via an internal ribosomal entry site. This perturbs cellular amino acid and nucleotide pools and strains energy resources and causes MYC-dependent apoptosis. eIF2B5 limits MYC expression and prevents apoptosis in APC-deficient murine and patient-derived organoids and in APC-deficient murine intestinal epithelia in vivo. Conversely, the high MYC levels present in APC-deficient cells induce phosphorylation of eIF2alpha via the GCN2 and PKR kinases. Pharmacological inhibition of GCN2 phenocopies eIF2B5 depletion and has therapeutic efficacy in tumour organoids, demonstrating that a negative MYC/eIF2alpha feedback loop constitutes a targetable vulnerability of colorectal tumours.
2019-11-04 | GSE106858 | GEO
Project description:Tumours depend on altered rates of protein synthesis for growth and survival, which suggests that mechanisms controlling mRNA translation may be exploitable for therapy. Here, we show that loss of APC, which occurs almost universally in colorectal tumours, strongly enhances the dependence on the translation initiation factor eIF2B5. Depletion of eIF2B5 induces an integrated stress response and enhances translation of MYC via an internal ribosomal entry site. This perturbs cellular amino acid and nucleotide pools, strains energy resources and causes MYC-dependent apoptosis. eIF2B5 limits MYC expression and prevents apoptosis in APC-deficient murine and patient-derived organoids and in APC-deficient murine intestinal epithelia in vivo. Conversely, the high MYC levels present in APC-deficient cells induce phosphorylation of eIF2α via the kinases GCN2 and PKR. Pharmacological inhibition of GCN2 phenocopies eIF2B5 depletion and has therapeutic efficacy in tumour organoids, which demonstrates that a negative MYC-eIF2α feedback loop constitutes a targetable vulnerability of colorectal tumours.
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Project description:To identify new Gcn2 target proteins, we performed a set of Stable Isotope Labeling by Amino Acids in Cell Culture (SILAC)-based quantitative phosphoproteomics experiments, in which we compared WT cells to gcn2∆ and sui2S52A cells (n=3), all treated, or not, with rapamycin, which has previously been shown to stimulate eIF2alpha phosphorylation by Gcn2.
2021-03-22 | PXD021109 | Pride
Project description:Ribosome stalling during translation has recently been shown to cause neurodegeneration, yet the signaling pathways triggered by stalled elongation complexes are unknown. To investigate these pathways we analyzed the brain of B6J-nmf205-/- mice in which neuronal elongation complexes are stalled at AGA codons due to deficiencies in a tRNA Arg(UCU) tRNA and GTPBP2, a mammalian ribosome rescue factor. Increased levels of phosphorylation of eIF2α (Ser51) were detected prior to neurodegeneration in these mice and transcriptome analysis demonstrated activation of ATF4, a key transcription factor in the integrated stress response (ISR) pathway. Genetic experiments showed that this pathway was activated by the eIF2alpha kinase, GCN2, in an apparent deacylated tRNA-independent fashion. Further we found that the ISR attenuates neurodegeneration in B6J-nmf205-/- mice, underscoring the importance of cellular and stress context on the outcome of activation of this pathway. These results demonstrate the critical interplay between translation elongation and initiation in regulating neuron survival during cellular stress. Examination of gene expression in cerebellum and hippocampus for 4 mice strains derived from C57BL/6J (B6J) strain. Microarray data was performed for 3 week and 5 week old mice in both cerebellum and hippocampus for B6J and B6J-nmf205-/- three replicates each. RNA-Seq data was perform on cerebellum of mice 3 weeks old, three replicates for each genotype: B6J, B6J-nmf205-/-, B6J-Gcn2-/- and B6J-nmf205-/-;Gcn2-/-.
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Project description:The p53 transcription factor is a master regulator of cellular stress responses restrained by repressors such as MDM2 and the phosphatase PPM1D. Activation of p53 with pharmacological inhibitors of its repressors is being tested in clinical trials for cancer therapy, but efficacy has been limited by poor induction of tumor cell death. We demonstrate that dual inhibition of MDM2 and PPM1D induces cell death in multiple cancer cell types via amplification of the p53 transcriptional program through the eIF2alpha -ATF4 pathway. PPM1D inhibition induces phosphorylation of eIF2alpha, ATF4 accumulation, and ATF4-dependent enhancement of p53-dependent transactivation upon MDM2 inhibition. Dual inhibition of p53 repressors depletes heme and induces HRI-dependent eIF2alpha phosphorylation. Pharmacological induction of eIF2alpha phosphorylation synergizes with MDM2 inhibition to induce cell death and halt tumor growth in mice. These results demonstrate that PPM1D inhibits both the p53 network and the integrated stress response controlled by eIF2alpha, with clear therapeutic implications.
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Project description:The p53 transcription factor is a master regulator of cellular stress responses restrained by repressors such as MDM2 and the phosphatase PPM1D. Activation of p53 with pharmacological inhibitors of its repressors is being tested in clinical trials for cancer therapy, but efficacy has been limited by poor induction of tumor cell death. We demonstrate that dual inhibition of MDM2 and PPM1D induces cell death in multiple cancer cell types via amplification of the p53 transcriptional program through the eIF2alpha -ATF4 pathway. PPM1D inhibition induces phosphorylation of eIF2alpha, ATF4 accumulation, and ATF4-dependent enhancement of p53-dependent transactivation upon MDM2 inhibition. Dual inhibition of p53 repressors depletes heme and induces HRI-dependent eIF2alpha phosphorylation. Pharmacological induction of eIF2alpha phosphorylation synergizes with MDM2 inhibition to induce cell death and halt tumor growth in mice. These results demonstrate that PPM1D inhibits both the p53 network and the integrated stress response controlled by eIF2alpha, with clear therapeutic implications.
2022-09-01 | GSE191149 | GEO
Project description:Aging is a highly regulated process in eukaryotes. Many biochemical pathways are affected by aging and, among them, protein homeostasis is one key process. Our hypothesis is that protein translation may be important to maintain a correct balance of active, properly folded proteins in the cell and, therefore, help supporting a normal longevity. Phosphorylation of eIF2alpha is one of the most important regulatory steps in protein translation in eukaryotes, reducing total protein translation and promoting the specific production of some proteins (Gcn4, Atf4, etc) We decided to test whether eIF2alpha phosphorylation has specific roles in protein translation during aging in Schizosaccharomyces pombe
2021-04-19 | GSE155256 | GEO
Project description:In eukaryotes, regulation of mRNA translation enables a fast, localized and finely tuned expression of gene products. Within the translation process, the first stage of translation initiation is most rigorously modulated by the actions of eukaryotic initiation factors (eIFs) and their associated proteins. These 11 eIFs catalyze the joining of the tRNA, mRNA and rRNA into a functional translation complex. Their activity is influenced by a wide variety of extra- and intracellular signals, ranging from global, such as hormone signaling and unfolded proteins, to specific, such as single amino acid imbalance and iron deficiency. Their action is correspondingly comprehensive, in increasing or decreasing recruitment and translation of most cellular mRNAs, and specialized, in targeting translation of mRNAs with regulatory features such as a 5’ terminal oligopyrimidine tract (TOP), upstream open reading frames (uORFs), or an internal ribosomal entry site (IRES). In mammals, two major pathways are linked to targeted mRNA translation. The target of rapamycin (TOR) kinase induces translation of TOP and perhaps other subsets of mRNAs, whereas a family of eIF2 kinases does so with mRNAs containing uORFs or an IRES. TOR targets translation of mRNAs that code for proteins involved in translation, an action compatible with its widely accepted role in regulating cellular growth. The four members of the eIF2 kinase family increase translation of mRNAs coding for stress response proteins such as transcription factors and chaperones. Though all four kinases act on one main substrate, eIF2, published literature demonstrates both common and unique effects by each kinase in response to its specific activating stress. This suggests that the activated eIF2 kinases regulate the translation of both a global and a specific set of mRNAs. Up to now, few studies have attempted to test such a hypothesis; none has been done in mammals. We use array analysis to determine the global mRNA shift into polysomes following a stress response, and to compare the translational response following activation of GCN2 versus PERK, two of the four eIF2alpha kinases. Experiment Overall Design: Gcn2 wild-type or knockout mouse liver were perfused with complete amino acids media or media lacking methionine for RNA extraction and hybridization of Affymetrix microarrays. RNA was extracted from unfractionated liver samples and polysome fraction of samples separated on sucrose density gradient. To minimize biological variations, we pooled RNA from two perfused liver samples to use in each array analysis. The conditions were total and polysome fraction of Gcn2+/+, +Met or -Met; total and polysome fraction of Gcn2-/-, +Met or -Met. Each array analysis was done in duplicate.
2009-11-23 | E-GEOD-11496 | biostudies-arrayexpress
Project description:General control nonderepressible 2 (GCN2) phosphorylates eIF2α, regulating translation in response to nutritional stress. Here, we show that mammalian ribosomes are potent GCN2 activators. Hydrogen/deuterium exchange–mass spectrometry (HDX-MS) showed GCN2 interacting with domain II of the uL10 P-stalk protein. The P-stalk is a uL10/P12/P22 pentameric complex that is part of the ribosomal GTPase-associated center. Recombinant human P-stalk greatly stimulates GCN2. Both domain II of uL10 and the C-terminal tails of P1 and P2 are necessary for maximal GCN2 activation. On actively translating ribosomes, the C-terminal tails of P1 and P2 are sequestered by elongation factors, suggesting P-stalk availability could link translational stress to GCN2 activation.
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