Ontology highlight
ABSTRACT:
PROVIDER: PRJEB43689 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| ERR9884890.fastq.gz | Fastqsanger.gz | |||
| ERR9884900.fastq.gz | Fastqsanger.gz | |||
| ERR9884910.fastq.gz | Fastqsanger.gz | |||
| ERR9884920.fastq.gz | Fastqsanger.gz | |||
| ERR9884930.fastq.gz | Fastqsanger.gz |
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Project description:Non-stop mRNAs generate a ground state of mitochondrial gene expression noise
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Project description:Pervasive transcription generates mainly unstable non-coding transcripts. Although a few examples of pervasive antisense have been shown to mediate gene regulation by transcriptional interference, whether pervasive transcription has a general functional role or merely represents transcriptional noise remains to be determined. In a mutant context that revealed pervasive transcripts, we characterized more than 800 antisenses genome wide and analysed the corresponding sense mRNA behaviour. We observed that antisense non-coding transcription is commonly found associated with genes tightly repressed during the exponential phase with an opposite level of mRNA and the associated antisense. This suggested that antisense transcription might participate to a tight repression of genes during the exponential phase. We thus specifically interrupted the antisense transcription of a subset of genes, and found that it resulted in a de-repression of the corresponding mRNAs. We further validated that this repression acts in cis through a transcriptional interference mechanism, which involves several chromatin modification factors. Transcription interference is observed for the first time as a general mean of gene regulation between cellular states.
2018-07-23 | GSE101368 | GEO
Project description:Proline-rich antimicrobial peptide apidaecin (Api) inhibits bacterial protein synthesis in a distinctive way. In vitro biochemical and structural studies showed that Api binds in the nascent peptide exit tunnel of the ribosome that has completed translation of the gene and has released the newly-synthesized protein. Once in the tunnel, Api traps the release factors on the post-termination ribosome leading to cessation of translation. The mode of Api action in the bacterial cell, however, had remained unknow. By analyzing distribution of ribosomes on mRNAs in Api-treated cells using Ribo-seq approach we uncovered a range of effects that stem from the unique mechanism of Api action. Exposure of bacterial cells to Api results in arrest of translation at stop codons, likely in a post-release state, due to the immediate Api action, as well as in a pre-release state due to the depletion of available release factors. In addition, arrest of translation at the end of the open reading frame (ORF) leads to a pronounced queuing of the translating ribosomes behind the one arrested at the stop codon. One of the major consequences of the Api action is a dramatically increased stop codon bypass by the ribosomes paused in a pre-release state, leading to accumulation of proteins with C-terminal extensions, as confirmed by whole-cell proteomics analysis. Stop codon bypass, that occurs either in 0-frame, by misincorporation of a near-cognate aminoacyl-tRNA at the stop codon, or via frameshifting allows for a fraction of the translating ribosomes to reach the 3’ ends of mRNA transcripts. The pervasive stalling of pre-release ribosomes at the stop codons and at the RNA transcripts ends triggers activation of the cellular ribosome rescue systems which, likely due to Api action as well, remain ineffective. While major Api effects are directed towards the ribosome at the end of the ORFs, cells exposed to Api show a somewhat increased ribosome occupancy of the start codons. Understanding the unique mode of Api to inhibit translation termination in the cell can help envisioning the development of treatments for genetic diseases and research tools for genome exploration.
2020-10-05 | PXD019012 | Pride
Project description:Pluripotency is established in E4.5 preimplantation epiblast. Embryonic stem cells (ESCs) represent the immortalization of pluripotency, however, they only partially resemble the gene expression signature of developmental ground-state. Induced PRAMEL7 expression, a protein highly expressed in the ICM but lowly expressed in ESCs, reprograms developmentally advanced ESC+serum into ground-state pluripotency by inducing a gene expression signature close to developmental ground-state. However, how PRAMEL7 reprograms gene expression remains elusive. Here we show that PRAMEL7 associates with Cullin2 (CUL2) and this interaction is required to establish ground-state gene expression. PRAMEL7 recruits CUL2 to chromatin and targets for proteasomal degradation regulators of repressive chromatin, including NuRD complex. PRAMEL7 antagonizes NuRD-mediated repression of genes implicated in pluripotency by decreasing NuRD stability and promoter association in a CUL2-dependent manner. Our data link proteasome degradation pathways to ground-state gene expression, offering insights to generate in vitro models to reproduce the in vivo ground-state pluripotency.
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Project description:The genetic code that specifies the identity of amino acids incorporated into proteins during protein synthesis is almost universally conserved. Mitochondrial translation deviates from the standard genetic code which includes the reassignment of two arginine codons into stop codons {Jukes, 1993 #438}. Translation termination at these non-canonical stop codons requires a protein factor to release the newly synthesized polypeptide chain, however, the identity of this factor is not known currently{Nadler, 2021 #406}. Here, we used gene editing and ribo-profiling in combination with cryo-electron microscopy to establish that the unusual mitochondrial release factor 1 (mtRF1) detects the non-canonical stop codons. We show that loss of mtRF1 leads to stalling of mitochondrial ribosomes on non-canonical stop codons and consequent reduced translation of cytochrome C oxidase subunit 1 that results in decreased mitochondrial respiration. We show that binding of mtRF1 to the decoding center of the ribosome stabilizes a highly unusual distortion in the mRNA conformation and that the ribosomal RNA importantly participates in the specific recognition of the non-canonical stop codons.
2023-03-21 | GSE216979 | GEO
Project description:The genetic code that specifies the identity of amino acids incorporated into proteins during protein synthesis is almost universally conserved. Mitochondrial translation, however, exhibits deviations from the standard genetic code including reassigning of two arginine codons to stop codons. Translation termination at these non-canonical stop codons requires a protein factor to release the newly synthesized polypeptide chain. Currently it is not known how, and by which release factor, these stop codons are recognized. Here, we used biochemical experiments on knockout mutants in human cells in combination with cryo-electron microscopy to establish that the unusual mitochondrial release factor 1 (mtRF1) detects the non-canonical stop codons. We show that loss of this factor leads to stalling of mitochondrial ribosomes on non-canonical stop codons. As a result, reduced levels of cytochrome C oxidase subunit 1 of the oxidative phosphorylation complex IV are synthesized leading to a defect in mitochondrial respiration. We further show that binding of mtRF1 to the decoding center of the ribosome stabilizes a highly unusual distortion in the mRNA conformation and that the ribosomal RNA importantly participates in the specific recognition of the non-canonical stop codons.
2023-05-09 | GSE216779 | GEO
Project description:The proline-rich antimicrobial peptide apidaecin (Api) inhibits bacterial protein synthesis in a distinctive way. In vitro biochemical and structural studies showed that Api binds in the nascent peptide exit tunnel of a ribosome that has completed translation of a gene and traps the release factors on the ribosome. By analyzing the distribution of ribosomes on mRNAs in Api-treated cells using Ribo-seq, we uncovered a range of effects that stem from the unique mechanism of Api action. Exposure of bacterial cells to Api results in arrest of ribosomes at stop codons, likely in a post-release state due to direct Api action, as well as in a pre-release state due to the depletion of available release factors. In addition, Api action leads to a pronounced queuing of translating ribosomes behind those arrested at stop codons. One of the major consequences of Api action is a dramatically increased level of stop codon bypass by ribosomes paused in a pre-release state, leading to an accumulation of proteins with C-terminal extensions, as confirmed by whole-cell proteomics analysis. Stop codon bypass occurs either in 0-frame, by misincorporation of a near-cognate aminoacyl-tRNA at the stop codon, or via frameshifting, allowing for a fraction of the translating ribosomes to reach the 3’ ends of mRNA transcripts. The pervasive stalling of pre-release ribosomes at the stop codons and at the RNA transcripts ends triggers activation of the cellular ribosome rescue systems which, likely due to Api action as well, remain ineffective. Understanding the unique mode of Api action as translation termination inhibitor in the cell can advance the development of treatments for infection and genetic diseases as well as new research tools for genome exploration.
2020-10-02 | GSE150034 | GEO
Project description:Current human pluripotent stem cells lack the transcription factor circuitry that governs the ground state of mouse embryonic stem cells (ESC). Here we report that short-term expression of two components, NANOG and KLF2, is sufficient to ignite other elements of the network and reset the human pluripotent state. Inhibition of ERK and protein kinase C signalling sustains a transgene-independent rewired state. Reset cells self-renew continuously without ERK signalling, are phenotypically stable and karyotypically intact. They differentiate in vitro and form teratomas in vivo. Metabolism is reprogrammed in reset cells with activation of mitochondrial respiration as in ESC. DNA methylation is dramatically reduced and transcriptome state is globally realigned across multiple cell lines. Depletion of ground state transcription factors, TFCP2L1 or KLF4 has marginal impact on conventional human pluripotent stem cells, but collapses the reset state. These findings demonstrate feasibility of installing and propagating functional control circuitry for ground state pluripotency in human cells. DNA methylation analysis in Conventional and Reset human embryonic stem cells by whole genome bisulfite sequencing, in triplicate, using the Illumina platform
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Project description:Translational control plays a central role in regulation of gene expression and can lead to significant divergence between mRNA- and protein-abundance. The translational landscape of early mammalian development and its impact on cellular proteome, however, remains largely un-explored. Here we used genome-wide approaches combined with time-course analysis to measure the mRNA-abundance, mRNA-translation rate and protein expression during the transition of naïve into primed embryonic stem cells (ESCs). We found that the ground state ESCs cultured with GSK3- and MEK-inhibitors and LIF (2iL) display higher ribosome density on a selective set of mRNAs. These mRNAs show reduced translation during the exit from ground state pluripotency and transition to serum/LIF (SL) culture or upon commitment to primed epiblast-like stem cells (EpiLSCs). Strikingly, integrative analysis with cellular proteome indicate a strong translational buffering of this set of mRNAs in 2iL-ESCs leading to stable protein expression levels. Our data reveal that the global alteration of cellular proteome is largely accompanied by transcriptional rewiring. Furthermore, we identified a set of genes (including UHRF1 and KRAS) that undergo selective post-translational regulation during the transition of naïve into primed pluripotency and linked the observed changes to upstream GSK- and MEK/MAPK-signaling pathways using single inhibitor treated ESCs. Thus, we provide a comprehensive and detailed overview of the global changes in gene expression during the transition of naïve to primed pluripotency and dissect the relative contributions of RNA-transcription, translation and regulation of protein stability in controlling protein abundance.
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Project description:Current human pluripotent stem cells lack the transcription factor circuitry that governs the ground state of mouse embryonic stem cells (ESC). Here we report that short-term expression of two components, NANOG and KLF2, is sufficient to ignite other elements of the network and reset the human pluripotent state. Inhibition of ERK and protein kinase C signalling sustains a transgene-independent rewired state. Reset cells self-renew continuously without ERK signalling, are phenotypically stable and karyotypically intact. They differentiate in vitro and form teratomas in vivo. Metabolism is reprogrammed in reset cells with activation of mitochondrial respiration as in ESC. DNA methylation is dramatically reduced and transcriptome state is globally realigned across multiple cell lines. Depletion of ground state transcription factors, TFCP2L1 or KLF4 has marginal impact on conventional human pluripotent stem cells, but collapses the reset state. These findings demonstrate feasibility of installing and propagating functional control circuitry for ground state pluripotency in human cells.
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