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Mus musculus

ABSTRACT: Small RNA profiling of WT E14, Ago2_KO and Ago1_KO mouse embryonic stem cells before and after XEN conversion and small RNA profiling after Immunoprecipitation of AGO1&2 proteins in WT E14, Ago1_KO and Ago2_KO mESCs.

PROVIDER: PRJNA318281 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR3370950.fastq.gz	Fastqsanger.gz
	SRR3370951.fastq.gz	Fastqsanger.gz
	SRR3370942.fastq.gz	Fastqsanger.gz
	SRR3370952.fastq.gz	Fastqsanger.gz
	SRR3370943.fastq.gz	Fastqsanger.gz

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Small RNA profiling of WT E14, Ago2_KO and Ago2_KO-AGO1 mouse embryonic stem cells.

Project description:Purpose: The goals of this study was to obtain the profile total miRNAs in WT E14, Ago2_KO mESCs to compare with Ago2_KO-AGO1 overexpressing AGO1.

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Small RNA profiling of WT E14, Ago2_KO and Ago1_KO mouse embryonic stem cells before and after XEN conversion and small RNA profiling after Immunoprecipitation of AGO1&2 proteins in WT E14, Ago1_KO and Ago2_KO mESCs.

Project description:Purpose: The goals of this study was to obtain the profile total miRNAs in WT E14, Ago2_KO and Ago1_KO mESCs to compare it with Immunoprecipitated microRNAs in AGO1&2 proteins.

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Project description:Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study was to obtain the trasncriptome of Ago2_KO mESCs to compare it with the transcriptome of WT mESCs before and after XEN conversion (deposit separately).

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Project description:Small RNA profiling of WT E14, Ago2_KO and Ago2_KO-AGO1 mouse embryonic stem cells.

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Convergence of distinct RNA-silencing pathways on GW182/Tnrc6

Project description:The RNA-binding protein Trim71/Lin41 is a phylogenetically conserved developmental regulator that functions in mammalian stem cell reprogramming, brain development and cancer. Trim71 recognizes target mRNAs through hairpin motifs and silences them through molecular mechanisms that await identification. Deposited data are mass spectrometry data from immunoprecipitation experiments with endogenously tagged Trim71, Ago2, and Tnrc6a. Experiments are carried out in mouse embryonic stem cells (mESCs) and uncover reciprocal interactions of Trim71, Ago2, and Tnrc6a. Trim71 protein interactions are largely independent of Ago2 levels, but strongly depend on the presence of RNA. A second set of experiments is mass spectrometry data from immunoprecipitation experiments in mESCs overexpressing a tagged peptide derived from Homo sapiens TNRC6B (‘FLAG-HA::T6BWT::Cherry’), that is known to block the Tnrc6-Ago2 interaction. We show that a wild-type version strongly binds Ago1 and Ago2, but not Trim71, while a mutant version binds neither Ago1, 2, nor Trim71.

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Project description:The transcription factor Sox17 is expressed in early primitive endoderm-fated cells of the mouse embryo and in embryo-derived extraembryonic endoderm (ExEn) stem (XEN) cells. We have shown that overexpression of Sox17 in mouse embryonic stem cells (ESCs) drives cell fate to a committed XEN-like cell state (Sox17-XEN cells). When placed back into the embryo, Sox17-XEN cells contribute exclusively to the ExEn. Transient Sox17 expression is sufficient to drive this fate change during which time cells transit through distinct intermediate states prior to the generation of functional XEN-like cells. We identified dynamic regulatory networks driving Sox17-mediated XEN conversion by analyzing a dynamic regulatory map of gene expression bifurcation points throughout conversion, created using RNA-seq time series data. We found that Sox17 orchestrates this conversion process by acting in autoregulatory and feed-forward network motifs, regulating dynamic gene regulatory networks (GRNs) directing cell fate. We have shown that Sox17-mediated XEN conversion provides a powerful tool for understanding the regulation of cell fate changes and for the elucidation of GRNs regulating lineage decisions in the mouse embryo. Total RNA was extracted during a time course of Sox17 overexpression in mouse ESCs at 7 time points as well as from wild-type ESCs and wild-type XEN cells.

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Project description:The inner cell mass of the mouse pre-implantation blastocyst is comprised of epiblast progenitor and primitive endoderm cells of which cognate embryonic (mESCs) or extra-embryonic (XEN) stem cell lines can be derived. Importantly, each stem cell type retains the defining properties and lineage restriction of their in vivo tissue of origin. Recently, we demonstrated that XEN-like cells arise within mESC cultures. This raises the possibility that mESCs can generate self-renewing XEN cells without the requirement for gene manipulation. We have developed a novel approach to convert mESCs to XEN cells (cXEN) using growth factors. We confirm that the down-regulation of the pluripotency transcription factor Nanog and the expression of primitive endoderm-associated genes Gata6, Gata4, Sox17 and Pdgfra are necessary for cXEN cell derivation. This approach highlights an important function for Fgf4 in cXEN cell derivation. Paracrine FGF-signalling compensates for the loss of endogenous Fgf4, which is necessary to exit mESC self-renewal, but not for XEN cell maintenance. Our cXEN protocol also reveals that distinct pluripotent stem cells respond uniquely to differentiation promoting signals. cXEN cells can be derived from mESCs cultured with Erk- and Gsk3-inhibitors (2i) and LIF, similarly to conventional mESCs. However, we find that epiblast stem cells (EpiSCs) derived from the post-implantation embryo are refractory to cXEN cell establishment, consistent with the hypothesis that EpiSCs represent a pluripotent state distinct from mESCs. In all, these findings suggest that the potential of mESCs includes the capacity to give rise to both extra-embryonic and embryonic lineages. A total of eight samples were analyzed. Three mouse embryonic stem cell (mESC) lines were used as a negative control, 2 embryo-derived extraembryonic endoderm (XEN) cell lines were used as a positive control, 3 converted XEN (cXEN) cells were used as the experiemental cell lines.

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