Single-cell RNA-seq of mouse embryoid body cells, day 3 to day 7. Includes wild type and Tet TKO.
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA552269 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR9625960_1.fastq.gz | Fastqsanger.gz | |||
| SRR9625960_2.fastq.gz | Fastqsanger.gz | |||
| SRR9625970_1.fastq.gz | Fastqsanger.gz | |||
| SRR9625970_2.fastq.gz | Fastqsanger.gz | |||
| SRR9625980_1.fastq.gz | Fastqsanger.gz |
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Project description:Single-cell methylation and accessibility of mouse embryoid body cells, day 3 to day 7. Includes wild type and Tet TKO.
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Project description:Tet enzymes (Tet1/2/3) convert 5-methylcytosine (5mC) to 5-hydroxy-methylcytosine (5hmC) and are dynamically expressed in various embryonic and adult cell types. While loss of individual Tet enzymes or combined deficiency of Tet1/2 allows for embryogenesis, the effect of complete loss of Tet activity and 5hmC marks in development is not established. We have generated Tet1/2/3 triple knockout (TKO) mouse embryonic stem cells (ESCs) and examined their developmental potential. Combined deficiency of all three Tets depleted 5hmC and impaired ESC differentiation as seen in poorly differentiated TKO embryoid bodies (EBs) and teratomas. Consistent with impaired differentiation, TKO-ESCs contributed poorly to chimeric embryos and could not support embryonic development. Global gene expression and methylome analyses of TKO-EBs revealed promoter hypermethylation and deregulation of genes implicated in embryonic development and differentiation. These findings suggest a requirement for Tet- and 5hmC-mediated DNA demethylation in proper regulation of gene expression during differentiation of ESCs and development. To quantify global gene expression in differentiating embryoid bodies (EBs) derived from wild type (WT) and Tet triple knockout (TKO), TKO and WT mouse embryonic stem cells (ESCs) were differentiated in vitro to EBs and cultured for 10 days. RNA was extracted using Qiagen RNeasy kit and subjected to microarray analysis. Global gene expression profile of two technical replicas of WT embryoid bodies (2 samples in total) was compared to two technical replicas of two independent TKO embryoid bodies (4 TKO samples in total).
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Project description:Tet enzymes (Tet1/2/3) catalyze the conversion of 5-methylcytosine (5mC) to 5-hydroxy-methylcytosine (5hmC) and are dynamically expressed in various embryonic and adult cell types. While loss of individual Tet enzymes or combined deficiency of Tet1/2 allows for embryogenesis, the effect of complete loss of Tet activity and 5hmC marks in development has not been established. To define the role of Tet enzymes and 5hmC in development we have generated Tet1, Tet2 and Tet3 triple knockout (TKO) mouse embryonic stem cells (ESCs) and examined their developmental potential in vitro and in vivo. Combined deficiency of all three Tet enzymes led to complete depletion of 5hmC and impaired ESC differentiation as seen in poorly differentiated TKO embryoid bodies and teratomas. Consistent with impaired differentiation, TKO ES cells exhibited limited contribution to the chimeric embryos and could not support embryonic development in tetraploid complementation assays. Gene expression profiles and genome wide methylome analyses of TKO embryoid bodies revealed promoter hypermethylation and deregulation of genes implicated in embryonic development and differentiation. These findings suggest a requirement for Tet and 5hmC-mediated DNA demethylation in proper regulation of gene expression during differentiation of embryonic stem cells and development.
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