Single-cell RNA-seq of naive and primed human embryonic stem cells
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PROVIDER: PRJEB26646 | ENA |
REPOSITORIES: ENA
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Project description:This study aims to profile the transcriptomes of single naive and primed human embryonic stem cells. Cells from the H9 line were cultured to select for naive or primed phenotypes, and a sequencing library was generated from each single cell using the Smart-seq2 method. This was repeated for multiple experimental batches, i.e., independent cultures. Batch 1 consists of sequencing runs 2383 and 2384; batch 2 consists of runs 2678, 2679, 2739 and 2740; and batch 3 consists of runs 2780 and 2781. Transcriptional profiles for all cells were obtained from the sequencing data and used to explore substructure and heterogeneity in the population for each phenotype.
2018-07-26 | E-MTAB-6819 | biostudies-arrayexpress
Project description:Pluripotent stem cells can give rise to the three embryonic germ layers and the characterization of their properties is crucial to exploit their therapeutic potential. Mouse embryonic stem cells (mESCs) are isolated and usually maintained in vitro in a primed state that resembles the post-implantation epiblast features. Furthermore, primed mESCs can be de-differentiated to a naive state that resembles the pre-implantation inner cell mass (ICM). Cell differentiation or genotoxic stress, among others, can alter DNA replication, which is a flexible process able to adapt to different cellular contexts. Here, we demonstrate that primed-to-naive mESC reprogramming triggers replication fork slowdown, increased fork asymmetry and a compensatory activation of dormant origins. Using iPOND (“isolation of proteins on nascent DNA”) coupled to mass spectrometry we have characterized the changes in replisome composition between naive and primed mESCs. Several DNA repair factors, including MRE11 nuclease, are enriched in naive mESCs forks, while factors involved in ubiquitin-dependent protein metabolism are enriched in primed mESC forks. We report that primed-to-naive mESC de-differentiation promotes recruitment of MRE11 to the forks in response to transcription-replication conflicts, underlying the DNA replication rewiring required for efficient mESC reprogramming.
2024-04-03 | PXD041946 | Pride
Project description:Naive and primed human pluripotent stem cells (hPSC) provide valuable models to study cellular and molecular developmental processes. The lack of detailed information about cell-surface protein expression in these two pluripotent cell types prevents an understanding of how the cells communicate and interact with their microenvironments. Here, we used plasma membrane profiling to directly measure cell-surface protein expression in naive and primed hPSC. This unbiased approach quantified over 1700 plasma membrane proteins including those involved in cell adhesion, signalling and cell interactions. Notably, multiple cytokine receptors upstream of JAK-STAT signalling were more abundant in naive hPSC. In addition, functional experiments showed that FOLR1 and SUSD2 proteins are highly expressed at the cell surface in naive hPSC but are not required to establish human naive pluripotency. This study provides a comprehensive stem cell proteomic resource that uncovers differences in signalling pathway activity and has identified new markers to define human pluripotent states.
2020-04-17 | PXD015359 | Pride
Project description:Human pluripotent cell lines were derived from blastocyst-stage embryos and propagated in self-renewal conditions that maintain features of naive pluripotency characteristic of mouse embryonic stem cells. Genome-wide DNA methylation status of HNES1 and HNES3 naive and primed cells was assessed with post-bisulfite adapter tagging (PBAT).
2016-03-01 | E-MTAB-4462 | biostudies-arrayexpress
Project description:Upon implantation, the naive pluripotent epiblast of the mouse blastocyst generates a rosette, undergoes lumenogenesis and forms the primed pluripotent egg cylinder, able to generate the embryonic tissues. How pluripotency progression and morphogenesis are linked, and whether intermediate pluripotent states exist remain controversial. We identify here a rosette pluripotent state, defined by co-expression of naive factors with transcription factor OTX2. Downregulation of blastocyst WNT signals drives transition into rosette pluripotency by inducing OTX2. The rosette then activates MEK signals that induce lumenogenesis and drive progression to primed pluripotency. Consequently, combined WNT and MEK inhibition supports rosette-like stem cells (RSCs), a self-renewing naive-primed intermediate. RSCs erase constitutive heterochromatin marks and display a primed chromatin landscape, with bivalently marked primed pluripotency genes. Nonetheless, WNT induces reversion to naive pluripotency. The rosette is therefore a reversible pluripotent intermediate where control over both pluripotency progression and morphogenesis pivots from WNT to MEK signals.
2020-04-28 | GSE112234 | GEO
Project description:Autophagy is a conserved cellular mechanism to degrade unwanted cytoplasmic proteins and organelles to recycle their components, and it is proved to be critical for embryonic stem cell (ESC) self-renewal and somatic cell reprogramming. However, the role of autophagy in embryonic development remains elusive, and no information exists regarding its functions during the transition from naive to primed pluripotency. Here by using an in vitro transition model of ESCs to epiblast-like cells (EpiLCs), we describe that the dynamic changes in Atg7-dependent autophagy is required for the naive to primed transition, and it is also necessary for germline specification. RNA-seq and ATAC-seq profiling reveal that Nanog acts as a barrier to prevent pluripotency transition, and autophagy-dependent Nanog degradation is important for dismantling the naive pluripotency expression program through decommissioning of naive-associated active enhancers. Mechanistically, we found that autophagy adaptor protein Sqstm1 (p62) is nucleus located during the pluripotency transition period and it is preferentially associated with ubiquitinated Nanog for selective protein degradation. In vivo, loss of autophagy by Atg7 depletion disrupts peri-implantation development and we observed increased chromatin association of Nanog, which affects neuronal differentiation through activation of a subset of neuroectodermal development-associated enhancers. Taken together, our findings illuminate regulatory mechanisms underlying the naive to primed transition and reveal that autophagy-dependent regulation of Nanog is essential for exit from the naive state and marks distinct cell fate allocation during lineage specification.
2020-11-07 | GSE160976 | GEO
Project description:Autophagy is a conserved cellular mechanism to degrade unwanted cytoplasmic proteins and organelles to recycle their components, and it is proved to be critical for embryonic stem cell (ESC) self-renewal and somatic cell reprogramming. However, the role of autophagy in embryonic development remains elusive, and no information exists regarding its functions during the transition from naive to primed pluripotency. Here by using an in vitro transition model of ESCs to epiblast-like cells (EpiLCs), we describe that the dynamic changes in Atg7-dependent autophagy is required for the naive to primed transition, and it is also necessary for germline specification. RNA-seq and ATAC-seq profiling reveal that Nanog acts as a barrier to prevent pluripotency transition, and autophagy-dependent Nanog degradation is important for dismantling the naive pluripotency expression program through decommissioning of naive-associated active enhancers. Mechanistically, we found that autophagy adaptor protein Sqstm1 (p62) is nucleus located during the pluripotency transition period and it is preferentially associated with ubiquitinated Nanog for selective protein degradation. In vivo, loss of autophagy by Atg7 depletion disrupts peri-implantation development and we observed increased chromatin association of Nanog, which affects neuronal differentiation through activation of a subset of neuroectodermal development-associated enhancers. Taken together, our findings illuminate regulatory mechanisms underlying the naive to primed transition and reveal that autophagy-dependent regulation of Nanog is essential for exit from the naive state and marks distinct cell fate allocation during lineage specification.
2020-07-22 | GSE154860 | GEO
Project description:Derivation of naive state of mouse embryonic stem cells (mESCs) in LIF+serum (LS) culture condition is strain dependent, whereas derivation of ground state mESCs is readily possible from all strains tested so far in “2i” culture condition. ESCs can be derived from the post-implantation stage mouse embryos (EpiSCs), showing primed characteristics. In the present study, we characterized and compared the transcriptional profile of naïve, primed and ground state mESCs. Considering the importance of genetic background of mouse model for ESCs derivation in conventional culture conditions, all ESCs lines used in the study were derived from the same strain of mice. We found distinct transcriptional profiles between naive, primed and ground state mESCs. Primed state mESCs exhibit lower expression of pluripotency markers along with higher expression of lineage specific markers compared to naive and ground state mESCs. We also demonstrate that the differentiation propensity of ESCs to specific germ layer varies depending on the pluripotency state of ESCs.
2018-04-12 | GSE78144 | GEO
Project description:Human naive pluripotent stem cells have unrestricted lineage potential. Underpinning this property, naive cells are thought to lack chromatin-based lineage barriers. However, this assumption has not been tested. Here, we apply multi-omics to comprehensively define the chromatin-associated proteome, histone post-translational modifications and transcriptome of human naive and primed pluripotent stem cells. Integrating the chromatin-bound proteome and histone modification data sets reveals differences in the relative abundance and activities of distinct chromatin modules, identifying a strong enrichment of Polycomb Repressive Complex 2 (PRC2)-associated H3K27me3 in naive pluripotent stem cell chromatin. Single-cell approaches and human blastoid models reveal that PRC2 activity acts as a chromatin barrier restricting the differentiation of naive cells towards the trophoblast lineage, and inhibiting PRC2 promotes trophoblast fate induction and cavity formation. Our results establish that human naive pluripotent stem cells are not epigenetically unrestricted, but instead possess chromatin mechanisms that oppose the induction of alternative cell fates.
2022-05-06 | PXD028111 | Pride
Project description:Autophagy is a conserved cellular mechanism to degrade unwanted cytoplasmic proteins and organelles to recycle their components, and it is proved to be critical for embryonic stem cell (ESC) self-renewal and somatic cell reprogramming. However, the role of autophagy in embryonic development remains elusive, and no information exists regarding its functions during the transition from naive to primed pluripotency. Here by using an in vitro transition model of ESCs to epiblast-like cells (EpiLCs), we describe that the dynamic changes in Atg7-dependent autophagy is required for the naive to primed transition, and it is also necessary for germline specification. RNA-seq and ATAC-seq profiling reveal that Nanog acts as a barrier to prevent pluripotency transition, and autophagy-dependent Nanog degradation is important for dismantling the naive pluripotency expression program through decommissioning of naive-associated active enhancers. Mechanistically, we found that autophagy adaptor protein Sqstm1 (p62) is nucleus located during the pluripotency transition period and it is preferentially associated with ubiquitinated Nanog for selective protein degradation. In vivo, loss of autophagy by Atg7 depletion disrupts peri-implantation development and we observed increased chromatin association of Nanog, which affects neuronal differentiation through activation of a subset of neuroectodermal development-associated enhancers. Taken together, our findings illuminate regulatory mechanisms underlying the naive to primed transition and reveal that autophagy-dependent regulation of Nanog is essential for exit from the naive state and marks distinct cell fate allocation during lineage specification.
2020-05-20 | GSE150865 | GEO