Reduced MEK inhibition preserves genomic stability in naïve human ES cells
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA435590 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR6748810_1.fastq.gz | Fastqsanger.gz | |||
| SRR6748810_2.fastq.gz | Fastqsanger.gz | |||
| SRR6748820_1.fastq.gz | Fastqsanger.gz | |||
| SRR6748820_2.fastq.gz | Fastqsanger.gz | |||
| SRR6748811_1.fastq.gz | Fastqsanger.gz |
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Project description:This SuperSeries is composed of the SubSeries listed below.
2018-07-10 | GSE111020 | GEO
Project description:Human embryonic stem cells (hESCs) can be captured in either a primed state resembling the post-implantation epiblast or in a naïve state resembling the pre-implantation epiblast. Naïve culture conditions allow study of pre-implantation development ex vivo but reportedly lead to chromosomal abnormalities, compromising their utility in research and potential therapeutic applications. Although MEK inhibition is essential for the naïve state, here we show that reduced MEK inhibition facilitates the establishment and maintenance of naïve hESCs that retain naïve-specific transcriptional and DNA methylation patterns yet accrue fewer chromosomal abnormalities. We further show that hESCs cultured under these modified conditions are less prone to apoptosis and proliferate more rapidly. Our results point to the importance of residual MAPK signaling and provide a simple modification to current protocols to enable robust growth and genomic stability in naïve hESCs.
2018-07-10 | GSE111018 | GEO
Project description:Human embryonic stem cells (hESCs) can be captured in either a primed state resembling the post-implantation epiblast or in a naïve state resembling the pre-implantation epiblast. Naïve culture conditions allow study of pre-implantation development ex vivo but reportedly lead to chromosomal abnormalities, compromising their utility in research and potential therapeutic applications. Although MEK inhibition is essential for the naïve state, here we show that reduced MEK inhibition facilitates the establishment and maintenance of naïve hESCs that retain naïve-specific transcriptional and DNA methylation patterns yet accrue fewer chromosomal abnormalities. We further show that hESCs cultured under these modified conditions are less prone to apoptosis and proliferate more rapidly. Our results point to the importance of residual MAPK signaling and provide a simple modification to current protocols to enable robust growth and genomic stability in naïve hESCs.
2018-07-10 | GSE110805 | GEO
Project description:Reduced MEK inhibition confers a growth advantage and reduces genomic instability in naïve human ES cells (RNA-seq data set)
| PRJNA434588 | ENA
Project description:Reduced MEK inhibition confers a growth advantage and reduces genomic instability in naïve human ES cells (RRBS-seq data set)
| PRJNA435603 | ENA
Project description:Human embryonic stem cells (hESCs) can be captured in a primed state in which they resemble the postimplantation epiblast, or in a naive state where they resemble the preimplantation epiblast. Naive-cell-specific culture conditions allow the study of preimplantation development ex vivo but reportedly lead to chromosomal abnormalities, which compromises their utility in research and potential therapeutic applications. Although MEK inhibition is essential for the naive state, here we show that reduced MEK inhibition facilitated the establishment and maintenance of naive hESCs that retained naive-cell-specific features, including global DNA hypomethylation, HERVK expression, and two active X chromosomes. We further show that hESCs cultured under these modified conditions proliferated more rapidly; accrued fewer chromosomal abnormalities; and displayed changes in the phosphorylation levels of MAPK components, regulators of DNA damage/repair, and cell cycle. We thus provide a simple modification to current methods that can enable robust growth and reduced genomic instability in naive hESCs.
| S-EPMC6127858 | biostudies-literature
Project description:Embryonic stem cell (ESC) cultures display a heterogeneous gene expression profile, ranging from a pristine naïve pluripotent state to a primed epiblast state. While it is known that the addition of inhibitors of GSK3? and MEK (so-called 2i conditions) push ESC cultures towards a more homogeneous naïve pluripotent state, the molecular underpinnings of this naïve transition are not completely understood. Here we demonstrate that Dazl, a RNA-binding protein previously thought to be expressed specifically in developing primordial germ cells (PGCs), marks a subpopulation of ESCs in vitro that is actively transitioning toward naïve pluripotency. In the absence of Dazl expression, ESCs fail to induce proper expression of Tet enzymes required for 5-hydroxymethylation in 2i-culture conditions. As a result, 5-hydroxymethylation of methylated cystosine residues is impaired. Indeed, we demonstrate that Tet1 and Tet2 are mRNA targets of Dazl, indicating that Dazl might play a role in protection or stabilizing these mRNA molecules. Our results provide insight in the regulation of the acquisition of naïve pluripotency and demonstrate that Dazl is required for TET-mediated cytosine hydroxymethylation in cells that are actively reprogramming to a pluripotent ground state. Two independent mouse ES cell lines, Dazl-GFP and Stella-GFP, were cultured on ?-irradiated feeder MEFs in DMEM containing 15% FBS or serum-free B27N2 medium both supplemented with leukemia inhibitory factor (LIF) (Ying 2008). For the 2i experiments, 1µM MEK inhibitor PD0325901 (Axon Medchem), and 5 µM GSK3? inhibitor Kenpaullone (Tocris), were used. Cells were harvest at 0, 3 and 10 days in each condition for expression microarray analysis.
2015-07-01 | E-GEOD-69055 | biostudies-arrayexpress
Project description:Since gastric cancer (GC) is characterized by amplifications of receptor tyrosine kinases (RTK) and KRAS, targeting the RTK/KRAS downstream pathways could contribute to broader applicability of molecular targeted therapy in GC. Here, we assembled a panel of 49 GC cell lines and screened predictors for responsiveness to RAF/MEK/ERK pathway inhibition by comparing their sensitivity to MEK inhibition with alteration status of RTK/KRAS and phosphorylation status of RTK/KRAS downstream molecules. We found that GC cells with MET amplification or KRAS mutation tend to be sensitive to MEK inhibition. Furthermore, lower phosphorylation levels of mTORC1 targets, p70S6K and 4EBP1, was also significantly associated with sensitivity to MEK inhibition. Interestingly, the sensitive cells showed reduced mTORC1 activity accompanied by decreased rate of protein synthesis after MEK inhibition, suggesting that antiproliferative effect of MEK inhibition may be exerted through inhibition of protein synthesis, which was caused by reduced mTORC1 activity. Mechanistically, expression of some mTORC1 negative regulators, such as DDIT4, FOXO3, SESN1 and TSC1, were induced by MEK inhibition in highly-sensitive cell lines and knockdown of these negative regulators partially alleviated the suppression of mTORC1 activity and antiproliferative effect of MEK inhibition. Our data suggest that a subset of GCs, which would be distinguishable according to the predictive markers we identified, may be sensitive to MEK inhibition and have implications for the strategy in clinical trial of MEK inhibitors for GC.
2016-12-31 | GSE74842 | GEO
Project description:The transition of pluripotent stem cells (PSCs) from primed to naïve states constitutes a prototypical example of cellular plasticity. The naïve state can be stabilized by defined chemical cocktails that block extracellular signals, notably including the MEK pathway. However, little is known regarding the underlying transcriptional mechanisms. Here, we report that the transcriptional landscape of the naïve state can be mimicked in mouse and human PSCs by stimulating transcriptional enhancers. This is attained by inhibiting the CDK8 and CDK19 kinases, which are negative regulators of Mediator, a critical component of enhancer function. Mechanistically, CDK8/19i triggers a global increase in the recruitment of RNA Pol II at promoters and enhancers, hyperactivating enhancers and their target genes. Lastly, the emergence of naïve pluripotency in the pre-implantation epiblast coincides with a marked reduction in CDK8/19 activity, and CDK8/19i blocks its subsequent developmental progression. These findings suggest that naïve pluripotency during development includes hyperactivation of enhancers and can be captured in vitro, either by blunting extracellular signaling, or by stimulating enhancer-driven transcription. These principles may apply to other cellular transitions.
2020-09-29 | PXD009200 | Pride
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.
2020-04-03 | PXD014528 | Pride