Effect of emtine treatment on RNA expression and stablization of RNA otherwise degraded by NMD pathway in head and neck cell lines
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
SUBMITTER: Sanjai Sharma
PROVIDER: E-GEOD-29788 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
Similar Datasets
Project description:The data shows the effect of NMD inhbition on cell lines and the change in RNA transcripts. The data also shows comparison of non-transformed cells (tert kert) to a Head and Neck tumorigenic cell line SCC12.
2011-06-08 | GSE29788 | GEO
Project description:In this study, nonsense-mediated mRNA decay (NMD) inhibition was used for the discovery of truncating mutations. Six prostate cancer (PRCA) patients and their healthy brothers were selected from a group of HPCX1-linked families. mRNA was isolated from their lymphoblastic cells after pharmacological treatment. Expression analyses were done using Agilent 44K oligoarrays, and selected genes were screened for mutations by sequencing. In order to identify genes containing inactivating mutations in the Xq27-q28 region, an NMD microarray analysis with Agilent 44K Whole Human Genome oligonucleotide microarrays was performed in the families showing the strongest linkage to HPCX1. Five families were chosen with 6 affected and 6 healthy controls. Lymphoblastoid cell lines were derived from whole blood and treated with emetine to block NMD pathway. Total RNA was extracted from harvested cells (treated and untreated) and labeled with Cy5 and Cy3. Labeled samples were hybridized to the Agilent 44K Whole Human Genome Oligonucleotide Microarrays. After hybridization microarrays were scanned and data was extracted using Feature Extraction software. The candidate genes (n=17) for subsequent sequence analysis were selected according to the microarray analysis.
2011-12-31 | E-GEOD-24204 | biostudies-arrayexpress
Project description:Translation of messenger RNAs (mRNAs) with premature translation termination codons produces truncated proteins with potentially deleterious effects. This is prevented by nonsense-mediated mRNA decay (NMD) of these mRNAs. NMD is triggered by ribosomes terminating upstream of a splice site marked by an exon-junction complex (EJC), but also acts on many mRNAs lacking a splice junction after their termination codon. We developed a genome-wide CRISPR flow cytometry screen to identify regulators of mRNAs with premature termination codons in K562 cells. This screen recovered essentially all core NMD factors and suggested a role for EJC factors in degradation of PTCs without downstream splicing. Among the strongest hits were the translational repressors GIGYF2 and EIF4E2. GIGYF2 and EIF4E2 mediate translational repression but not mRNA decay of a subset of NMD targets and interact with NMD factors genetically and physically. Our results suggest a model wherein recognition of a stop codon as premature can lead to its translational repression through GIGYF2 and EIF4E2.
2021-10-12 | PXD027479 | Pride
Project description:Inhibition of the nonsense mediated decay (NMD) mechanism in cells results in stabilization of transcripts carrying premature translation termination codons. A strategy referred to as gene indentification by NMD inhibition (GINI) has been proposed to identify genes carrying nonsense mutations (Noensie & Dietz, 2001). Genes containing frameshift mutations in colon cancer cell line have been identifying mutatnt genes using GINI, we have now further improved the strategy. In this approach, inhibition of NMD with emetine is complemented with inhibiting NMD by blocking the phosphorylation of the hUpf1 protein with caffeine. In addition, to enhance the GINI strategy, comparing mRNA level alterations produced by inhibiting transcription alone or inhbiiting transcription together with NMD following caffeine pretreatment were used for the efficient identification of false positives produced as a result of stress response to NMD inhibition. To demonstrate the improved efficiency of this approach, we analyzed colon cancer cell lines showing microstellite instability. Bi-allelic inactivating mutations were found in the FXR1, SEC1L1, NCOR1, BAT3, PHD14, ZNF294, C190ORF5 genes as well as genes coding for proteins with yet unknown functions. Experiment Overall Design: Using GINI2 as a method of identifying novel biallelic truncating mutations in MSI+ colon cancer cells, various treatment conditions were applied to cultured colon cancer cell lines LS180 (MSI+) and SW480 (MSI-) and gene expression profiling was measured using Affymetrix GeneChip U133 Plus 2 arrays.
2008-06-13 | E-GEOD-5486 | biostudies-arrayexpress
Project description:Identification of potential tumor suppressor genes using the GINI strategy in Mantle Cell Lymphoma cell lines Experiment Overall Design: Inhibition of the NMD pathway was performed in Mantle Cell Lymphoma cell lines according to a modified protocol previously described (Noensie EN et al Nat Biotechnol 2001;19:434-439; Huusko P et al, Nat Gentet 2004; 36:979-983). The cells were treated with emetine or with emetine followed by incubation with actinomycin D during a time course. Mock treated cells were used as reference.
2008-06-15 | E-GEOD-6728 | biostudies-arrayexpress
Project description:Vitamin D is the strongest known natural anti-proliferative. A large number of studies in a wide spectrum of cancers, including epidemiological, in vitro and animal models, demonstrate that the active form of Vitamin D has anti-cancer benefits, affecting both progression and metastasis. Alike the role in calcium Vitamin D regulation, its anti-proliferative effect is thought to function through the Vitamin D receptor (VDR), although convincing evidence is lacking. Notwithstanding, separation of the calcemic and the anti-proliferative activity of Vitamin D analogues has been a major obstacle in developing new drugs for the treatment of cancer. The work presented attempts to unveil the molecular mechanism behind the anti-proliferative action of Vitamin D using genomic tools. For that purpose four independently developed Vitamin D sensitive/resistant MCF7 cell line pairs were collected. These unique biological replicates enabled us, both to increase the power of our study and to omit the use of Vitamin D. We deem this omission crucial since in the presence of Vitamin D only downstream genes involved in proliferation and cell cycle would be identified rather than causal resistance genes. The use of a variety of genomic techniques including expression, NMD and oligo CGH arrays reveal in the resistant cell lines the 11q13-14 as a region of DNA copy number loss and an altered expression of EGFR signaling pathway genes. Surprisingly, no genes known from calcium Vitamin D regulation were identified, nor did the VDR silencing by RNAi induce resistance to the sensitive cell lines. Keywords: Expression microarray, NMD array, cell type comparision Several MCF7 breast tumor cell lines independently derived from the human breast cancer cell line, both resistant and sensitive to Vitamin D, were used. The pairs of cell lines (parental-resistant) were developed in different laboratories and using different methodologies; while some cell lines acquired resistance by long exposure to the physiological concentration of Vitamin D (100nM), others were induced to resistant by being exposed to increasing amounts of Vitamin D. A total of 4 microarray expression experiments were carried out. In all microarray experiments RNA from the Vitamin D resistant cell line treated with Emetine was hybrizided agains RNA from the respsective sensitive/parental cell line treated with emetine.
2008-06-01 | E-GEOD-9959 | biostudies-arrayexpress
Project description:Nonsense mediated mRNA decay is a translation-dependent surveillance pathway that detects and eliminates RNA harboring premature termination codon. Our NMD complexes analysis led to the description of strong in vivo interaction between Nmd4, Ebs1 and Upf1, the major factor of NMD. We wondered if these proteins affected the degradation of RNA sensitive to NMD and if they could act on a specific class of targets. We performed RNAseq experiments in strains deleted for NMD4, EBS1 or both in comparison with wild type and upf1∆ strains. Our analysis confirmed that Nmd4 and Ebs1 have a role in the degradation of NMD substrates. The effect of Nmd4 or Ebs1 deletion was weak compared to upf1∆, fold change compared with WT was between -1 and 1 in log2. The double mutant had a global stronger effect. We have not detected any target specificity for Nmd4 or Ebs1, their profiles were very well correlated. We have also compared our data with upf1∆ RNAseq results. Among the more stabilized RNA in ebs1 and nmd4∆ strains, the majority were also stabilized in upf1∆. We observed a good correlation between upf1∆ and ebs1∆ profiles. For nmd4∆ the correlation was weaker, probably due to the low effect of the deletion, on NMD substrates stabilisation. We conclude that Nmd4 and Ebs1 play a specific role in the stabilization of all NMD substrates.
2018-02-19 | GSE102099 | GEO
Project description:Nonsense mediated decay (NMD) is a translation termination coupled quality control system. NMD identifies and degrades aberrant mRNAs containing premature termination codons, thereby preventing the accumulation of detrimental truncated proteins. NMD also controls the expression of several normal mRNAs and non-coding transcripts. RNA-seq assays were conducted to identify the NMD regulated genes in Arabidopsis. However, NMD targets have not been studied in other plants. To identify the conserved NMD targets, we wanted to identify the NMD regulated genes in Nicotiana benthamiana model species. Virus induced gene silencing was used to inactivate UPF1 NMD key factor in N. benthamiana, and then comparative RNA-seq experiment was carried out using UPF1-silenced and control–silenced plants. We found that in N. benthamiana significantly more genes are controlled by NMD than in Arabidopsis and that, surprisingly few conserved NMD targets can be recognized. These results suggest that while the mechanism of NMD is highly conserved, the set of NMD controlled genes can be quickly changed in plants.
2019-07-01 | GSE109130 | GEO
Project description:The RNA helicase UPF1 interacts with mRNAs, mRNA decay machinery, and the terminating ribosome to promote nonsense-mediated mRNA decay (NMD). Structural and biochemical data have revealed that UPF1 exists in an enzymatically autoinhibited “closed” state. Upon binding the NMD protein UPF2, UPF1 undergoes an extensive conformational change into a more enzymatically active “open” state, which exhibits enhanced ATPase and helicase activity. However, mechanically deficient UPF1 mutants can support efficient NMD, bringing into question the roles of UPF1 enzymatic autoinhibition and activation in NMD. Here, we identify two additional important features of the activated open state: slower nucleic acid binding kinetics and enhanced ATP-stimulated nucleic acid dissociation kinetics. Computational modeling based on empirical measurements of UPF1, UPF2, and RNA interaction kinetics predicts that the majority of UPF1-RNA binding and dissociation events in cells occur independently of UPF2 binding. We find that UPF1 mutants with either reduced or accelerated dissociation from RNA have NMD defects, whereas UPF1 mutants that are more dependent on UPF2 for catalytic activity remain active on well-established NMD targets. These findings support a model in which the kinetics of UPF1-mRNA interactions are important determinants of cellular NMD efficiency.
2024-05-29 | GSE246483 | GEO
Project description:The RNA helicase UPF1 interacts with mRNAs, mRNA decay machinery, and the terminating ribosome to promote nonsense-mediated mRNA decay (NMD). Structural and biochemical data have revealed that UPF1 exists in an enzymatically autoinhibited “closed” state. Upon binding the NMD protein UPF2, UPF1 undergoes an extensive conformational change into a more enzymatically active “open” state, which exhibits enhanced ATPase and helicase activity. However, mechanically deficient UPF1 mutants can support efficient NMD, bringing into question the roles of UPF1 enzymatic autoinhibition and activation in NMD. Here, we identify two additional important features of the activated open state: slower nucleic acid binding kinetics and enhanced ATP-stimulated nucleic acid dissociation kinetics. Computational modeling based on empirical measurements of UPF1, UPF2, and RNA interaction kinetics predicts that the majority of UPF1-RNA binding and dissociation events in cells occur independently of UPF2 binding. We find that UPF1 mutants with either reduced or accelerated dissociation from RNA have NMD defects, whereas UPF1 mutants that are more dependent on UPF2 for catalytic activity remain active on well-established NMD targets. These findings support a model in which the kinetics of UPF1-mRNA interactions are important determinants of cellular NMD efficiency.
2024-05-29 | GSE246484 | GEO