Project description:Many transcriptional and epigenetic networks must be integrated to maintain self-renewal and pluripotency in embryonic stem cells (ESCs) and to enable induced pluripotent stem cell (iPSC) reprogramming. Here, we explore the role of Zfp217and Mettl3 as RNA-binding proteins. Identification and characterization of RNAs associated with Zfp217 and Mettl3 in mouse embryoinc stem cells
Project description:Many transcriptional and epigenetic networks must be integrated to maintain self-renewal and pluripotency in embryonic stem cells (ESCs) and to enable induced pluripotent stem cell (iPSC) reprogramming. Here, we explore the role of Zfp217 as a key transcriptional factor in maintaining ES cell self-renewal by permorming genome-wide ChIP-Seq analyses. Examination of Zfp217 binding profiling by high throughput sequencing in mouse stem cells
Project description:Many transcriptional and epigenetic networks must be integrated to maintain self-renewal and pluripotency in embryonic stem cells (ESCs) and to enable induced pluripotent stem cell (iPSC) reprogramming. Here, we explore the role of Zfp217 as a key transcriptional factor in maintaining ES cell self-renewal by performing meRIP analysis in control and Zfp217-depleted mouse stem cells. Examination of m6A levels from total RNA in control and Zfp217 shRNA infected mouse stem cells
Project description:Comprehensive RNA-seq experiments in control and zfp217 shRNA infected cells delineate a Zfp217-driven network required for ES cell self-renewal. RNA-seq was used to measure gene expression levels in luciferase control and Zfp217 short hairpin RNA (shRNA) infected mouse stem cells
Project description:Bmi1 is a component of the Polycomb-repressive complexes (PRC) and essential for maintaining the pool of adult stem cells. PRC are key regulators for embryonic development by modifying chromatin architecture and maintaining gene repression. To assess the role of Bmi1 in pluripotent stem cells and upon exit from pluripotency during differentiation, we studied forced Bmi1 expression in mouse embryonic stem cells (ESC). We found that ESC do not express detectable levels of Bmi1 RNA and protein and that forced Bmi1 expression had no obvious influence on ESC self-renewal. However, upon ESC differentiation Bmi1 effectively enhanced development of hematopoietic cells. Global transcriptional profiling identified a large array of genes that were differentially regulated during ESC differentiation by Bmi1. Importantly, we found that Bmi1 induced a prominent up-regulation of Gata2, a zinc finger transcription factor, which is essential for primitive hematopoietic cell generation from mesoderm. In addition, Bmi1 caused sustained growth and a more than 100-fold expansion of ESC-derived hematopoietic stem/progenitor cells within 2-3 weeks of culture. The enhanced proliferative capacity was associated with reduced Ink4a/Arf expression in Bmi1-transduced cells. Taken together, our experiments demonstrate distinct activities of Bmi1 in ESC and ESC-derived hematopoietic progenitor cells. In addition, Bmi1 enhances the propensity of ESC in differentiating towards the hematopoietic lineage. Thus, Bmi1 could be a candidate gene for engineered adult stem cell derivation from ESC. 8 samples in total. Bmi1 embryonic stem cells sample_1 (Bmi1_ESC_1) Bmi1 embryonic stem cells sample_2 (Bmi1_ESC_2) Untreated CCE embryonic stem cells (CCE_ESC_Control) Empty vector CCE embryonic stem cells (CCE_ESC_Vector) Bmi1 embryoid body sample_1 (Bmi1_EB_1) Bmi1 embryoid body sample_2 (Bmi1_EB_2) Empty vector control embryoid body sample_1 (Vector_EB_1) Empty vector control embryoid body sample_2 (Vector_EB_2)
Project description:The embryonic stem (ES) cell transcriptional and epigenetic networks are critical for the maintenance of ES cell self-renewal. However, it remains unclear whether components of these networks functionally interact and if so, what factors mediate such interactions. Here we show that WD-repeat protein-5 (Wdr5), a core member of the mammalian Trithorax (trxG) complex, positively correlates with the undifferentiated state and is a novel regulator of ES cell self-renewal. We demonstrate that Wdr5, an ‘effector’ of H3K4 methylation, interacts with the pluripotency transcription factor Oct4. In concordance, genome-wide ChIP-Seq and transcriptome analyses demonstrate overlapping gene regulatory functions between Oct4 and Wdr5. We show that the Oct4-Sox2-Nanog circuitry cooperates with trxG for transcriptional activation of key self-renewal regulators. Furthermore, Wdr5 expression is required for the efficient formation of induced pluripotent stem (iPS) cells. Collectively, these findings implicate an integrated model of transcriptional and epigenetic control, mediated by select trxG members, for the maintenance of ES cell self-renewal and somatic cell reprogramming. 7 Samples
Project description:Mouse stem cells were differenciated with retinoic acid and cells were harvested at the indicated time points. RNA-seq was used to measure gene expression levels in retinoic acid-differentiated mouse stem cells
Project description:Whole-genome single-base resolution methylcytosine map reveals profound changes that occur after Lsh deletion during embryonic development in primary WT and Lsh-/- MEFs. Lsh deletion leads to widespread decreases of CG methylation level at uniquely mapped genomic regions compared to wild type, including TSSs at protein-coding genes, and non-coding RNA genes. MethylC-Seq from Mus musculus primary MEFs.
Project description:Purpose: The aim of this study was to profile the transcriptome of differentiating embryonic stem cells (ES) with nonsilencing shRNA mediated knockdown and Setdb1 geneTrap heterozygous cells with Setdb1 shRNA mediated knockdown. Methods: Female Setdb1+/+ XistâA/+ ES cells were induced to differentiate and RNA sampled on day 0, day 3 and day 5 of differentiation. RNAseq libraries were prepared with TruSeq RNA sample preparation v2 kit. Libraries were pooled and sequenced on the Illumina HiSeq 2000 platform for 100 bp single-end reads. Image analysis was performed in real time by the HiSeq Control Software (HCS) v1.4.8 and Real Time Analysis (RTA) v1.12.4.2, running on the instrument computer. Real-time base calling on the HiSeq instrument computer was performed with the RTA software. Illumina CASAVA1.8 pipeline was used to generate the sequence data. Total RNA was extracted and purified from each cell type and their transcriptomes analysed by RNA-Seq.