Splicing towards noncoding isoforms in colorectal carcinoma is associated with tumor hypoxia and the DNA damage response
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE81513 | GEO | 2016/07/28
SECONDARY ACCESSION(S): PRJNA321835
REPOSITORIES: GEO
Similar Datasets
Project description:Tumor hypoxia is associated with poor patient outcome and resistance to therapy. It is associated with a rapid decline in protein production mediated through mTOR signalling. Here we show that it also leads to widespread changes in splicing and a global shift towards the expression of noncoding isoforms, thus providing a novel and orthogonal mechanism by which cells can modulate protein expression. Examination of mRNA levels in HCT116 cells after 0 hr, 1 hr, 2 hr and 24 hr in hypoxia. Three biological replicates each.
2016-07-28 | E-GEOD-81513 | biostudies-arrayexpress
Project description:Tumour hypoxia is associated with poor patient outcome and resistance to therapy. It is accompanied by widespread changes in gene expression mediated largely through the transcription factors HIF1/2/3α. Hypoxia impacts on multiple pathways throughout the cell and has widespread effects on phenotype. Here we use sample-specific annotation approaches to determine the changes in transcript architecture that arise as result of alternative splicing in hypoxic cells. Using in vivo data generated from a time course in reduced oxygenation we identified genome-wide switching between coding and noncoding isoforms, including a significant number of components of the DNA damage response pathway. Notably, HDAC6, a master regulator of the cytotoxic response, and TP53BP1, which sits at the nexus of the double-strand break repair pathway, both underwent a marked transition towards an intron-retention pattern with a concomitant decline in protein levels. These transitions from coding to noncoding isoforms were recapitulated in a large and independent cohort of 499 colorectal samples taken from The Cancer Genome Atlas (TCGA). The set of altered genes was enriched for multiple components of the Fanconi Anaemia, nucleotide excision and double-strand break repair pathways, and together correlating with tumour status at last contact. Altogether, these data demonstrate a new role for hypoxia-driven alternative splicing in regulating DNA damage response, and highlight the importance of considering alternative splicing as a critical factor in our understanding of human disease.
| S-EPMC5417364 | biostudies-literature
Project description:The Ser/Thr protein kinase mTOR controls metabolic pathways, including the catabolic process of autophagy. Autophagy plays additional, catabolism-independent roles in homeostasis of cytoplasmic endomembranes and whole organelles. How signals from endomembrane damage are transmitted to mTOR to orchestrate autophagic responses is not known. Here we show that mTOR is inhibited by lysosomal damage. Lysosomal damage, recognized by galectins, leads to association of Gal8 with mTOR apparatus on the lysosome. Gal8 inhibits mTOR activity through its Ragulator-Rag signaling machinery. Thus, a novel galectin-based signal-transduction apparatus, termed here GALTOR, controls mTOR in response to lysosomal damage.
2017-12-07 | MSV000081788 | MassIVE
Project description:Recent data have linked hypoxia, a classic feature of the tumor microenvironment, to the function of specific microRNAs (miRNAs); however, whether hypoxia affects other types of noncoding transcripts is currently unknown. Starting from a genome-wide expression profiling, we demonstrate for the first time a functional link between oxygen deprivation and the modulation of long noncoding transcripts from ultraconserved regions, termed transcribed-ultraconserved regions (T-UCRs). Interestingly, several hypoxia-upregulated T-UCRs, henceforth named 'hypoxia-induced noncoding ultraconserved transcripts' (HINCUTs), are also overexpressed in clinical samples from colon cancer patients. We show that these T-UCRs are predominantly nuclear and that the hypoxia-inducible factor (HIF) is at least partly responsible for the induction of several members of this group. One specific HINCUT, uc.475 (or HINCUT-1) is part of a retained intron of the host protein-coding gene, O-linked N-acetylglucosamine transferase, which is overexpressed in epithelial cancer types. Consistent with the hypothesis that T-UCRs have important function in tumor formation, HINCUT-1 supports cell proliferation specifically under hypoxic conditions and may be critical for optimal O-GlcNAcylation of proteins when oxygen tension is limiting. Our data gives a first glimpse of a novel functional hypoxic network comprising protein-coding transcripts and noncoding RNAs (ncRNAs) from the T-UCRs category.
| S-EPMC3824588 | biostudies-literature
Project description:Type-I interferon (IFN)-induced activation of the mammalian target of rapamycin (mTOR) signaling pathway has been implicated in translational control of mRNAs encoding interferon-stimulated genes (ISGs). However, mTOR-sensitive translatomes commonly include mRNAs with a 5’ terminal oligopyrimidine tract (TOP), such as those encoding ribosomal proteins, but not ISGs. Because these translatomes were obtained under conditions when ISG expression is not induced, we examined the mTOR-sensitive translatome in human WISH cells stimulated with IFNβ. The mTOR inhibitor Torin1 resulted in a repression of global protein synthesis, including that of ISG products, and translation of all but 3 ISG mRNAs (TLR3, NT5C3A, and RNF19B) was not selectively more sensitive to mTOR inhibition. Detailed studies of NT5C3A revealed an IFN-induced change in transcription start site resulting in a switch from a non-TOP to a TOP-like transcript variant and mTOR sensitive translation. Thus, we show that, in the cell model used, translation of the vast majority of ISG mRNAs is not selectively sensitive to mTOR activity and describe an uncharacterized mechanism wherein the 5’-UTR of an mRNA is altered in response to a cytokine, resulting in a shift from mTOR-insensitive to mTOR-sensitive translation.
2015-07-07 | GSE70270 | GEO
Project description:Little is known about the role of mTOR signaling in plasma cell differentiation and function. Furthermore, for reasons not understood, mTOR inhibition reverses antibody-associated disease in a murine model of systemic lupus erythematosus. Here, we have demonstrated that induced B-lineage specific deletion of the gene encoding RAPTOR, an essential signaling adaptor for the rapamycin-sensitive mTOR complex 1 (mTORC1) abrogated the generation of antibody-secreting plasma cells in mice. Acute treatment with rapamycin recapitulated the effects of RAPTOR deficiency, while both strategies led to the ablation of newly formed plasma cells in the spleen and bone marrow while also obliterating pre-existing germinal centers. Surprisingly, although perturbing mTOR activity caused a profound decline in serum antibodies that were specific for exogenous antigen or DNA, frequencies of long-lived bone marrow plasma cells were unaffected. Instead, mTORC1 inhibition led to decreased expression of immunoglobulin binding protein (BiP) and other factors needed for robust protein synthesis. Consequently, blockade of antibody synthesis was rapidly reversed after termination of rapamycin treatment. We conclude that mTOR signaling plays critical but diverse roles in early and late phases of antibody responses and plasma cell differentiation.
2016-09-20 | GSE87035 | GEO
Project description:Long-read nanopore sequencing is capable of continuously reading thousands of base pairs. Here, we report the application of nanopore sequencing technology for analysing hypoxia-specific transcriptomes in normal liver and liver cancer cell lines, which help us to better understand the mechanism of cell survival and metastasis under hypoxia. We conducted transcriptomic sequencing of normal liver and liver cancer cell lines under normoxia or hypoxia, founding that long-read sequencing could detect thousands of hypoxia-specific isoforms and unannotated novel genes. At the cellular level, we have verified that the novel gene 644 is most up-regulated novel gene under hypoxic conditions. We observed that the novel gene 644 possesses classical mRNA characteristics, with its CDS region encoding 91 amino acids. Through protein structure predictions, we discovered that the novel gene 644 contains a transmembrane domain. Experimental validation confirmed that the novel gene 644 is a membrane protein, and knockdown of the novel gene 644 significantly suppresses the proliferative capacity of liver cancer cells under hypoxic conditions. Furthermore, we revealed the protein structure alteration of hypoxia specific isoforms with molecular dynamics simulation. The results indicate that our hypoxic dataset can serve as a foundation for the structural simulation of hypoxic-specific proteins. In summary, we have presented a hypoxic-specific third-generation sequencing dataset for further analysis, which provides sufficient details on cellular transcriptomes and is particularly valuable for understanding the survival and metastasis of tumor under hypoxic conditions. From this dataset, we have validated the importance of novel gene 644 for tumor survival under hypoxia and identified alterations in hypoxic-specific protein structures. These discoveries can bring us new knowledge about hypoxia, leading to the identification of potential tumor treatment targets and biomarkers.
2024-05-28 | GSE263126 | GEO
Project description:Purpose: mRNA translation into protein is highly regulated, but the role of mRNA isoforms, noncoding RNAs (ncRNAs), and genetic variants has yet to be systematically studied. Using high-throughput sequencing (RNA-seq), we have measured cellular levels of mRNAs and ncRNAs, and their isoforms, in lymphoblast cell lines (LCL) and in polysomal fractions, the latter shown to yield strong correlations of mRNAs with expressed protein levels. Analysis of allelic RNA ratios at heterozygous SNPs served to reveal genetic factors in ribosomal loading. Methods: RNA-seq was performed on cytosolic extracts and polysomal fractions (3 ribosomes or more) from three lymphoblastoid cell lines. As each RNA fraction was amplified (NuGen kit), and relative contributions from various RNA classes differed between cytosol and polysomes, the fraction of any given RNA species loaded onto polysomes was difficult to quantitate. Therefore, we focused on relative recovery of the various RNA classes and rank order of single RNAs compared to total RNA. Results: RNA-seq of coding and non-coding RNAs (including microRNAs) in three LCLs revealed significant differences in polysomal loading of individual RNAs and isoforms, and between RNA classes. Moreover, correlated distribution between protein-coding and non-coding RNAs suggests possible interactions between them. Allele-selective RNA recruitment revealed strong genetic influence on polysomal loading for multiple RNAs. Allelic effects can be attributed to generation of different RNA isoforms before polysomal loading or to differential loading onto polysomes, the latter defining a direct genetic effect on translation. Several variants and genes identified by this approach are also associated with RNA expression and clinical phenotypes in various databases. Conclusions: These results provide a novel approach using complete transcriptome RNA-seq to study polysomal RNA recruitment and regulatory variants affecting protein translation. cells from 3 samples were grown to 5x105 cells/mL density in T75 tissue culture flask and harvested, total RNA and polysome bound RNA was sequenced by Ion Proton
2015-06-30 | E-GEOD-66135 | biostudies-arrayexpress
Project description:Skeletal muscle weakness has been associated with different pathological conditions, including sarcopenia and muscular dystrophy, and is accompanied by altered mTOR signaling. Here we wanted to better elucidate the functional role of mTOR on muscle contractility. Most loss of function studies for mTOR signaling have used the drug rapamycin to inhibit some of the signaling downstream of mTOR. However, as rapamycin does not completely inhibit all mTOR signaling, we generated a double k.o. for mTOR and for the scaffold protein of mTORC1, Raptor, in skeletal muscle. We found that dk.o. mice results in a more severe phenotype compared to Raptor or mTOR deletion alone. Indeed, they display muscle weakness, increased fiber denervation, and a slower muscle relaxation following tetanic stimulation. This is accompanied by a shift towards slow-twitch fibers and changes in the expression levels of calcium-related genes, like Serca1 and Casq1. Indeed, dk.o. mice show a decrease in calcium decay kinetics after tetanus in vivo, suggestive of a reduced calcium reuptake. In addition, RNA sequencing analysis revealed that many downregulated genes are linked to sarcomere organization, like Tcap and Fhod3. These results suggest a key role for mTOR signaling in maintaining a proper fiber relaxation in skeletal muscle.
2022-10-19 | GSE215314 | GEO
Project description:The liver is the central hub for processing and maintaining homeostatic levels of dietary nutrients especially essential amino acids such as tryptophan (Trp). Trp is required not only to sustain protein synthesis but also as a precursor for the production of NAD, neurotransmitters and immunosuppressive metabolites. In light of these roles of Trp and its metabolic products, maintaining homeostatic levels of Trp is essential for health and well-being. The liver regulates global Trp supply by the immunosuppressive enzyme tryptophan-2,3-dioxygenase (TDO2), which degrades Trp down the kynurenine pathway (KP). In the current study, we show that isolated primary hepatocytes when exposed to hypoxic environments, extensively rewire their Trp metabolism by reducing constitutive Tdo2 expression and differentially regulating other Trp pathway enzymes and transporters. Mathematical modelling of Trp metabolism in liver cells under hypoxia predicted decreased flux through the KP while metabolic flux through the tryptamine branch significantly increased. In line, the model also revealed an increased accumulation of tryptamines under hypoxia, at the expense of kynurenines. Metabolic measurements in hypoxic hepatocytes confirmed the predicted reduction in KP metabolites as well as accumulation of tryptamine. Tdo2 expression in cultured primary hepatocytes was reduced upon hypoxia inducible factor (HIF) stabilisation by dimethyloxalylglycine (DMOG), demonstrating that HIFs are involved in the hypoxic downregulation of hepatic Tdo2. DMOG abrogated hepatic luciferase signals in Tdo2 reporter mice, indicating that HIF stability also recapitulates hypoxic rewiring of Trp metabolism in vivo. Also in WT mice HIF stabilization drove homeostatic Trp metabolism away from the KP towards enhanced tryptamine production, leading to enhanced levels of tryptamine in liver, serum and brain. As tryptamines are the most potent hallucinogens known, the observed upregulation of tryptamine in response to hypoxic exposure of hepatocytes may be involved in the generation of hallucinations occurring at high altitude. KP metabolites are known to activate the aryl hydrocarbon receptor (AHR). The AHR-activating properties of tryptamines may explain why immunosuppressive AHR activity is maintained under hypoxia despite downregulation of the KP. In summary our results identify hypoxia as an important factor controlling Trp metabolism in the liver with possible implications for immunosuppressive AHR activation and mental disturbances.
| S-EPMC7933006 | biostudies-literature