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Mus musculus

ABSTRACT: Single Cell Sequencing Identifies Key Epigenetic Regulators in Nuclear Transfer Mediated Reprogramming [RRBS]

PROVIDER: PRJNA289144 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR2089670_1.fastq.gz	Fastqsanger.gz
	SRR2089670_2.fastq.gz	Fastqsanger.gz
	SRR2089680_1.fastq.gz	Fastqsanger.gz
	SRR2089680_2.fastq.gz	Fastqsanger.gz
	SRR2089690_1.fastq.gz	Fastqsanger.gz

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Single Cell Sequencing Identifies Key Epigenetic Regulators in Nuclear Transfer Mediated Reprogramming.

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Project description:Here we performed genome-wide RNA-seq and Reduced Representation Bisulfite Sequencing (RRBS-seq) in isogenic human induced pluripotent stem cells (iPSCs) and somatic cell nuclear transfer-derived embryonic stem cells (nt-ESCs), genetically matched in vitro fertilization-derived ESCs (IVF-ESCs), and their respective differentiated cells (cardiomyocytes and endothelial cells). We generated the transcriptome and DNA methylome map in human pluripotent stem cells and their differentiated cells with single-nucleotide resolution. We compared the genetic (genetic makeup) and epigenetic (reprogramming approach) influence on the gene expression and DNA methylation profiles and found that genetic composition is the major contributor of the transcriptional and epigenetic variances observed in the undifferentiated and differentiated cells originated from different reprogramming mechanisms.

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Project description:Tet-mediated DNA oxidation is a new type of epigenetic modification in mammals and its role in the regulation of cell fate transition remains poorly understood. Here, we derive mouse embryonic fibroblasts (MEFs) deleted in all three Tet genes and examine their capability to be reprogrammed into iPS cells. We demonstrate that these Tet-deficient MEFs cannot be reprogrammed due to a blockage in the mesenchymal-to-epithelial transition (MET). Reprogramming of MEFs deficient in TDG is similarly blocked. The blockage is caused by impaired activation of crucial microRNAs, which depends on oxidative demethylation promoted by Tet and TDG. Reintroduction of either miR-200c or catalytically active Tet and TDG restores reprogramming to the respective knockout MEFs. Thus, oxidative demethylation is essential for somatic cell reprogramming. These findings provide mechanistic insights into the operation of epigenetic barriers in cell lineage conversion. Reduced Representation Bisulfite (RRBS, MspI,~75-400bp size fraction) and Tet-Assisted RRBS (TARRBS) of MEFs & reprogramming MEFs at Day 5

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