A global overview of translation efficiency and ribosome occupancy at nucleotide resolution in HEK293 cells
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ABSTRACT:
PROVIDER: PRJNA556704 | ENA |
REPOSITORIES: ENA
Similar Datasets
Project description:Looking at ribosome distribution and translational efficiency in control conditions in HEK293 cell line. Combining this information with our other datasets from HEK293 cells has allowed us to look at ribosome occupancy of specific groups of mRNAs
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Project description:Ribosome profiling in HEK293 cells with and without the depletion of CNOT1. This allows the determination of translational efficiency and nucleotide resolution of ribosome occupancy.
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Project description:We report the genomic occupancy of the BAHD1 protein in HEK293 cells over-expressing the BAHD1 gene (HEK293-HPT-BAHD1)
2015-12-11 | GSE53372 | GEO
Project description:We report the genomic occupancy of the BAHD1 protein in HEK293 cells over-expressing the BAHD1 gene (HEK293-HPT-BAHD1) We used Native ChIP-seq to identify DNA regions bound to BAHD1 (native ChIP involves use of native chromatin before precipitation of immune complexes, without formaldehyde crosslinking of proteins with nucleic acids)
2015-12-11 | E-GEOD-53372 | biostudies-arrayexpress
Project description:Technological limitations precluded transcriptome-wide analyses of translation at single cell resolution. To solve this challenge, we developed a novel microfluidic isotachophoresis approach, named RIBOsome profiling via IsoTachoPhoresis (Ribo-ITP), and characterized translation in single oocytes and embryos during early mouse development. We identified differential translation efficiency as a key regulatory mechanism of genes involved in centrosome organization and N6-methyladenosine modification of RNAs. Our high coverage measurements enabled the first analysis of allele-specific ribosome engagement in early development and led to the discovery of stage-specific differential engagement of zygotic RNAs with ribosomes and reduced translation efficiency of transcripts with allelic-biased expression. Finally, by integrating our measurements with proteomics data, we discovered that ribosome occupancy in germinal vesicle stage oocytes is the predominant determinant of protein abundance in the zygote. The novel Ribo-ITP approach will enable numerous applications by providing high coverage and high resolution ribosome occupancy measurements from ultra-low input samples including single cells.
2023-02-06 | GSE185732 | GEO
Project description:Protein-RNA interactions are fundamental to core biological processes, such as mRNA splicing, localization, degradation and translation. We have developed a photoreactive nucleotide-enhanced UV crosslinking and oligo(dT) purification approach to identify the mRNA-bound proteome using quantitative proteomics and to display the protein occupancy on mRNA transcripts by next-generation sequencing (Baltz and Munschauer et al. 2012). Our current work focuses on streamlining and extending protein occupancy profiling on poly(A)-RNA. Our objectives are to identify previously unknown protein-bound transcripts and, more importantly, to assess global and local differences in protein occupancy across different biological conditions. To this end, we have implemented poppi, the first pipeline for differential analysis of protein occupancy profiles. We have applied our analysis pipeline to pinpoint changes in occupancy profiles of MCF7 cells against already published HEK293 cells [GSE38157].
2013-12-05 | GSE49831 | GEO
Project description:The alteration of gene expression due to variations in the sequences of transcriptional regulatory elements has been a focus of substantial inquiry in humans and model organisms. However, less is known about the extent to which natural variation contributes to post-transcriptional regulation. Allelic Expression Imbalance(AEI) is a classical approach for studying the association of specific haplotypes with relative changes in transcript abundance. Here, we benchmarked a new TRAP based approach to associate genetic variation with transcript occupancy on ribosomes in specific cell types, to determine if it will allow examination of Allelic Translation Imbalance(ATI), and Allelic Translation Efficiency Imbalance, using as a test case mouse astrocytes in vivo. We show that most changes of the mRNA levels on ribosomes were reflected in transcript abundance, though ~1.5% of transcripts have variants that clearly alter loading onto ribosomes orthogonally to transcript levels. These variants were often in conserved residues and altered sequences known to regulate translation such as upstream ORFs, PolyA sites, and predicted miRNA binding sites. Such variants were also common in transcripts showing altered abundance, suggesting some genetic regulation of gene expression may function through post-transcriptional mechanisms. Overall, our work shows that naturally occurring genetic variants can impact ribosome occupancy in astrocytes in vivo and suggests that mechanisms may also play a role in genetic contributions to disease.
2023-08-17 | GSE156414 | GEO
Project description:Protein-RNA interactions are fundamental to core biological processes, such as mRNA splicing, localization, degradation and translation. We have developed a photoreactive nucleotide-enhanced UV crosslinking and oligo(dT) purification approach to identify the mRNA-bound proteome using quantitative proteomics and to display the protein occupancy on mRNA transcripts by next-generation sequencing (Baltz and Munschauer et al. 2012). Our current work focuses on streamlining and extending protein occupancy profiling on poly(A)-RNA. Our objectives are to identify previously unknown protein-bound transcripts and, more importantly, to assess global and local differences in protein occupancy across different biological conditions. To this end, we have implemented poppi, the first pipeline for differential analysis of protein occupancy profiles. We have applied our analysis pipeline to pinpoint changes in occupancy profiles of MCF7 cells against already published HEK293 cells. We generated protein occupancy cDNA libraries for two biological replicates. Briefly, we crosslinked 4SU-labeled MCF7 cells and purified protein-mRNA complexes using oligo(dT)-beads. The precipitate was treated with RNAse I to reduce the protein-crosslinked RNA fragments to a length of about 30-60 nt. To remove non-crosslinked RNA, protein-RNA complexes were precipitated with ammonium sulfate and blotted onto nitrocellulose. The RNA was recovered by Proteinase K treatment, ligated to cloning adapters, and reverse transcribed. The resulting cDNA libraries were PCR-amplified and next-generation sequenced.
2013-12-05 | E-GEOD-49831 | biostudies-arrayexpress
Project description:The stress of nucleotide pool reduction regulates transcription in neural crest and melanoma cells. To better understand the molecular response caused by nucleotide stress, we designed a chemical suppressor screen for leflunomide, an inhibitor of dihydroorate dehydrogenase. We found that alterations in the progesterone receptor (Pgr) activity suppressed the neural crest effects of leflunomide. To clarify the mechanism of action, we found that the RNA helicase protein, Ddx21, binds to Pgr, and loss of function of Ddx21 conferred resistance to nucleotide stress in zebrafish embryos. At the molecular level, nucleotide stress reduces DDX21 chromatin occupancy and thus, target gene expression. Together our results show that DDX21 is a transcriptional sensor and mediator of the nucleotide stress response.
2020-02-06 | PXD014433 | Pride
Project description:We perform polyA independent deep sequencing of chromatin associated primary transcripts across three different cell lines to obtain a global view on in vivo microRNA processing. We use these data to define a MicroProcessing Index (MPI), to quantify the cleavage efficiency of the Microprocessor complex. Hallmarks of efficient Drosha-mediated processing are confirmed by means of deep sequencing of chromatin-associated transcripts upon Drosha knockdown. Our results suggest that both sequence features and thermodynamic properties, e.g. secondary structure of the regions flanking the pre-miRNA hairpins are determinants for efficient processing. Our data furthermore enables us to observe endogenous microprocessor cleavage sites at nucleotide resolution. This analysis reveals the presence of non-canonical processing events occurring one helical turn distal of most efficiently cleaved miRNA precursors. We performed polyA independent deep sequencing of the chromatin-isolated RNA fraction for 5 samples: 2 replicates in HeLa cells, 1 Drosha knock down in HeLa cells, 1 sample for A549 cells and 1 sample for HEK293 cells. We also performed deep sequencing of the small RNA fraction in the same cell lines.
2014-04-17 | E-GEOD-56862 | biostudies-arrayexpress