Defining Essential Regulators of Naïve Human Pluripotent Cell Reprogramming
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA691952 | ENA |
REPOSITORIES: ENA
Similar Datasets
Project description:Defining how human pluripotent cell identity is controlled and in particular how naïve pluripotency is acquired during cell reprogramming is crucial for the future applications of pluripotent stem cells. However, the regulatory pathways of naïve cell reprogramming remain incompletely understood. Here, we used genome-wide CRISPR-Cas9 screening to identify novel regulators of primed to naïve pluripotent stem cell reprogramming, including genes that are essential for reprogramming and genes that normally impede reprogramming and whose targeted deletion led to enhanced reprogramming. Integrated analysis defined specific chromatin complexes and signalling pathways as critical regulators of naïve reprogramming, and were largely distinct from regulators of somatic cell reprogramming. Mechanistically, PRC1.3 and PRDM14 are jointly required to transcriptionally repress developmental and gene regulatory factors to ensure naïve cell reprogramming. Additionally, small molecule inhibitors of reprogramming impediments increased the efficiency of naïve cell reprogramming, and are of practical benefit that can improve on current reprogramming methods. Taken together, we have identified novel regulators controlling the establishment of naïve pluripotency in human cells, which will open up new ways to exploit the full potential of pluripotent stem cells. These results also provide new insights into mechanisms that destabilise and reconfigure cell identity during cell state transitions.
2022-04-06 | PXD023547 | Pride
Project description:Defining how human pluripotent cell identity is controlled and in particular how naïve pluripotency is acquired during cell reprogramming is crucial for the future applications of pluripotent stem cells. However, the regulatory pathways of naïve cell reprogramming remain incompletely understood. Here, we used genome-wide CRISPR-Cas9 screening to identify novel regulators of primed to naïve pluripotent stem cell reprogramming, including genes that are essential for reprogramming and genes that impede reprogramming and whose targeted deletion led to enhanced reprogramming. Integrated analysis defined specific chromatin complexes and signalling pathways as critical regulators of naïve reprogramming, and were largely distinct from regulators of somatic cell reprogramming. Mechanistically, PRC1.3 and PRDM14 are jointly required to transcriptionally repress developmental and gene regulatory factors to ensure naïve cell reprogramming. Additionally, small molecule inhibitors of reprogramming impediments increased the efficiency of naïve cell reprogramming, and are of practical benefit that can improve on current reprogramming methods. Taken together, we have identified novel regulators controlling the establishment of naïve pluripotency in human cells, which will open up new ways to exploit the full potential of pluripotent stem cells. These results also provide new insights into mechanisms that destabilise and reconfigure cell identity during cell state transitions.
2022-03-25 | GSE164786 | GEO
Project description:Defining the regulons of Pseudomonas aeruginosa transcriptional regulators
| PRJNA685406 | ENA
Project description:Cnot8 Eliminates Naïve Regulation Networks and Is Essential for Naïve-to-Formative Pluripotent Transition
| PRJNA729787 | ENA
Project description:To identify regulators of exit from the naïve pluripotent state we performed an enhancer trap screen under sensitized conditions. Briefly, haploid mouse ESCs with a Puromycin resistance gene under the control of Oct4 regulatory elements were mutagenized using viruses harboring enhancer trap constructs and subjected to differentiation in the presence of 3uM CHIR with concomitant selection for Oct4 expression. Selected cells were harvested after 16, 20, and 23 days and the extracted genomic DNA, together with genomic DNA of cells taken before selection, subjected to inverse PCR and sequencing. This allowed for mapping of enhancer trap integration sites and identifying genes whose targeting is enriched during selection.
2021-07-07 | E-MTAB-9796 | biostudies-arrayexpress
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 GSK3β 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. RNA-IP experiments were used to identify the RNA species bound to DAZL.
2015-07-01 | E-GEOD-69356 | biostudies-arrayexpress
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:Defining Essential Enhancer for Pluripotent stem cells using Features Oriented CRISPR-Cas9 Screen
| PRJNA490759 | ENA
Project description:Defining Essential Enhancer for Pluripotent stem cells using Features Oriented CRISPR-Cas9 Screen (4C-Seq)
| PRJNA490760 | ENA
Project description:Defining Essential Enhancer for Pluripotent stem cells using Features Oriented CRISPR-Cas9 Screen (ChIP-Seq)
| PRJNA544263 | ENA