De novo DNA methylation at imprinted loci during reprogramming into naïve and primed pluripotency
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA436116 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR6786630.fastq.gz | Fastqsanger.gz | |||
| SRR6786621_1.fastq.gz | Fastqsanger.gz | |||
| SRR6786621_2.fastq.gz | Fastqsanger.gz | |||
| SRR6786631.fastq.gz | Fastqsanger.gz | |||
| SRR6786622_1.fastq.gz | Fastqsanger.gz |
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Project description:CpG islands (CGIs) including those at imprinting control regions (ICRs) are protected from de novo methylation in somatic cells. However, many cancers often exhibit CGI hypermethylation, implying that the machinery is impaired in cancer cells. Here, we conducted a comprehensive analysis of CGI methylation during somatic cell reprogramming. Although most CGIs remain hypomethylated, a small subset of CGIs, particularly at several ICRs, was often de novo methylated in reprogrammed pluripotent stem cells (PSCs). Such de novo ICR methylation was linked with the silencing of reprogramming factors, which occurs at a late stage of reprogramming. The ICR-preferred CGI hypermethylation was similarly observed in human PSCs. Mechanistically, ablation of Dnmt3a prevented PSCs from de novo ICR methylation. Notably, the ICR-preferred CGI hypermethylation was observed in pediatric cancers, while adult cancers exhibit genome-wide CGI hypermethylation. These results may have important implications in the pathogenesis of pediatric cancers and the application of PSCs.
| S-EPMC6524733 | biostudies-literature
Project description:CpG islands (CGIs) including those at imprinting control regions (ICRs) are protected from de novo methylation in somatic cells. However, many cancers often exhibit CGI hypermethylation, implying that the machinery is impaired in cancer cells. Here, we conducted a comprehensive analysis of CGI methylation during the somatic cell reprogramming. Although most CGIs remain hypomethylated, a small subset of CGIs, particularly at several ICRs, were often de novo methylated in reprogrammed pluripotent stem cells (PSCs). Such de novo ICR methylation was linked with the silencing of reprogramming factors, which occurs at a late stage of reprogramming. The ICR-preferred CGI hypermethylation was similarly observed in human PSCs. Mechanistically, ablation of Dnmt3a prevented PSCs from de novo ICR methylation. Notably, the ICR-preferred CGI hypermethylation was observed in pediatric cancers, while adult cancers exhibit genome-wide CGI hypermethylation. These results may have important implications in the pathogenesis of pediatric cancers and the application of PSCs.
2019-06-18 | GSE111173 | GEO
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:Naïve and primed pluripotency is characterized by distinct signaling requirements, transcriptomes and developmental properties, but both cellular states share key transcriptional regulators, Oct4, Sox2 and Nanog. Here we demonstrate that transition between these two pluripotent states is associated with widespread Oct4 relocalization, mirrored by global rearrangement of enhancer chromatin landscapes. Our genomic and biochemical analyses identified candidate mediators of primed state-specific Oct4 binding, including Otx2 and Zic2/3. Even in the absence of other differentiation cues, premature Otx2 overexpression is sufficient to exit the naïve state, induce transcription of a large subset of primed pluripotency-associated genes and redirect Oct4 to thousands of previously inaccessible sites. However, ability of Otx2 to engage new enhancer regions is determined by its levels, cis-encoded properties of the sites and signaling environment. Our results illuminate regulatory mechanisms underlying pluripotency and suggest that capacity of transcription factors such as Otx2 and Oct4 to function as pioneers is highly context-dependent transcription profile of ESCs and EpiLCs to analzye changes during differentiation and the effect of Otx2 loss and overexpression on the differentiation properties
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Project description:Deciphering the regulatory network for human naïve and primed pluripotency is of fundamental theoretical and applicable significance. Here, by combining quantitative proteomics, phosphoproteomics and acetylproteomics analyses, we revealed RNA processing and translation as the most differentially-regulated processes between naïve and primed human embryonic stem cells (hESCs). While glycolytic primed hESCs rely predominantly on eIF4E-mediated cap-dependent pathway for protein translation, naïve hESCs with reduced mTORC1 activity are more tolerant to blockage of eIF4E-dependent translation, and their bivalent metabolism allows for translating selective mRNAs via both eIF4E-dependent and eIF4E-independent/eIF4A2-dependent pathways to form a more compact naïve proteome. Globally up-regulated proteostasis system and down-regulated post-translational modification system help to further refine and maintain the naïve proteome that is compatible with the more rapid cycling of naïve hESCs, where CDK1 plays an indispensable coordinative role.
2021-12-10 | PXD030230 | Pride
Project description:Naïve and primed pluripotency is characterized by distinct signaling requirements, transcriptomes and developmental properties, but both cellular states share key transcriptional regulators, Oct4, Sox2 and Nanog. Here we demonstrate that transition between these two pluripotent states is associated with widespread Oct4 relocalization, mirrored by global rearrangement of enhancer chromatin landscapes. Our genomic and biochemical analyses identified candidate mediators of primed state-specific Oct4 binding, including Otx2 and Zic2/3. Even in the absence of other differentiation cues, premature Otx2 overexpression is sufficient to exit the naïve state, induce transcription of a large subset of primed pluripotency-associated genes and redirect Oct4 to thousands of previously inaccessible sites. However, ability of Otx2 to engage new enhancer regions is determined by its levels, cis-encoded properties of the sites and signaling environment. Our results illuminate regulatory mechanisms underlying pluripotency and suggest that capacity of transcription factors such as Otx2 and Oct4 to function as pioneers is highly context-dependent ChIP-seq analysis was performed to map enhancers and associated transcription factors. We used H3K27ac, H3K4me1 and p300 to call enhancers from 2 different pluripotent cell states: ESC and EpiLC. In addition we performed ChIP-seq for Oct4 and Otx2 from these cell states. All these experiments were carried out in replicates, for the EpiLC state the replicates were performed with and without ActivinA. Additionally we carried out ChIPseq for Otx2 and Oct4 in Otx2ko cell lines in which we integrated an inducible Otx2 gene before and after induction with doxycycline.
2014-06-05 | E-GEOD-56098 | biostudies-arrayexpress
Project description:Naïve and primed pluripotent stem cells (PSCs) and germ cells express the Pou5f1 (Oct4) gene. The Oct4 gene contains two cis-regulatory elements, the distal (DE) and proximal enhancer (PE), which differentially control Oct4 expression in a cell-type- and stage-specific manner. Here, we generated double transgenic mice carrying both Oct4-ΔPE-GFP and Oct4-ΔDE-tdTomato (RFP), enabling us to simultaneously monitor the activity of DE and PE. Oct4 expression is stage-specifically regulated by DE and PE during embryonic and germ cell development. Using this dual reporter system, we successfully cultured pure populations of naïve (GFP+RFP-) and primed (GFP-RFP+) PSCs. We found that GFP+RFP- cells were metastable in serum-containing medium; stable naïve pluripotent cells were observed in medium containing two inhibitors (Meki and GSKi) but lacking serum. Finally, we suggest that the activity of Oct4 DE and PE is regulated by the repressive histone marks H3K9me3 and DNA methylation.
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Project description:Naïve and Primed MSC derived exosomes
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Project description:Although cell therapies require large numbers of quality-controlled hPSCs, existing technologies are limited in their ability to efficiently grow and scale stem cells. We report here that cell-state conversion from primed-to-naïve pluripotency enhances the biomanufacturing of hPSCs. Naïve hPSCs exhibit superior growth kinetics and aggregate formation characteristics in stirred suspension bioreactors compared to their primed counterparts. Moreover, we demonstrate the role of the bioreactor mechanical environment in the maintenance of naïve pluripotency, through transcriptomic enrichment of mechano-sensing signaling for cells in the bioreactor along with a decrease in expression of lineage-specific and primed pluripotency hallmarks. Bioreactor-cultured, naïve hPSCs express epigenetic regulatory transcripts associated with naïve pluripotency, and display hallmarks of X-chromosome reactivation. They exhibit robust production of naïve pluripotency metabolites and display reduced expression of primed pluripotency cell surface markers. We also show that these cells retain the ability to undergo targeted differentiation into beating cardiomyocytes, hepatocytes, and neural rosettes. They additionally display faster kinetics of teratoma formation compared to their primed counterparts. Naïve bioreactor hPSCs also retain structurally stable chromosomes. Our research corroborates that converting hPSCs to the naïve state enhances hPSC manufacturing and indicates a potentially important role for the bioreactor’s mechanical environment in maintaining naïve pluripotency.
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