Similar Datasets
Project description:Transcriptional bursting is the stochastic activation and inactivation of promoters, contributing to cell-to-cell heterogeneity in gene expression. However, the mechanism underlying the regulation of transcriptional bursting kinetics (burst size and frequency) in mammalian cells remains elusive. In this study, we performed single-cell RNA sequencing to analyze the intrinsic noise and mRNA levels for elucidating the transcriptional bursting kinetics in mouse embryonic stem cells. Informatics analyses and functional assays revealed that transcriptional bursting kinetics was regulated by a combination of promoter- and gene body-binding proteins, including the polycomb repressive complex 2 and transcription elongation factors. Furthermore, large-scale CRISPR-Cas9-based screening identified that the Akt/MAPK signaling pathway regulated bursting kinetics by modulating transcription elongation efficiency. These results uncovered the key molecular mechanisms underlying transcriptional bursting and cell-to-cell gene expression noise in mammalian cells.
| S-EPMC7299619 | biostudies-literature
Project description:Genome-wide kinetic properties of transcriptional bursting in mouse embryonic stem cells
| PRJNA548515 | ENA
Project description:Cell-to-cell heterogeneity in gene expression can even be observed in the same type of cells, present in a similar environment. Transcriptional bursting is thought to be one of contributing factors to the heterogeneity, but it remains elusive how the kinetic properties of transcriptional bursting (e.g. burst size, burst frequency, and noise induced by transcriptional bursting) are regulated in mammalian cells. To unbiasedly identify genes regulating the kinetic properties of transcriptional bursting, we performed large-scale CRISPR/-Cas9 based screening and functional analysis, and found that Akt/MAPK signaling pathways are involved in the regulation of the kinetic properties of transcriptional bursting.
2020-06-04 | GSE132590 | GEO
Project description:Recent studies have demonstrated that the Ten-eleven translocation (Tet) family proteins can enzymatically convert 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC). While 5mC has been studied extensively, little is known about the distribution and function of 5hmC. Here we present a genome-wide profile of 5hmC in mouse embryonic stem (ES) cells. A combined analysis of global 5hmC distribution and gene expression profile in wild-type and Tet1-depleted ES cells suggests that 5hmC is enriched at both gene bodies of actively transcribed genes and extended promoter regions of Polycomb-repressed developmental regulators. Thus, our study reveals the first genome-wide 5hmC distribution in pluripotent stem cells, and supports its dual function in regulating gene expression.
| S-EPMC3070931 | biostudies-literature
Project description:Smad7 has been identified as a negative regulator of the transforming growth factor TGF-β pathway by direct interaction with the TGF-β type I receptor (TβR-I). Although Smad7 has also been shown to play TGF-β unrelated functions in the cytoplasm and in the nucleus, a comprehensive analysis of its nuclear function has not yet been performed. Here, we show that in ESCs Smad7 is mainly nuclear and acts as a general transcription factor regulating several genes unrelated to the TGF-β pathway. Loss of Smad7 results in the downregulation of several key stemness master regulators, including Pou5f1 and Zfp42, and in the upregulation of developmental genes, with consequent loss of the stem phenotype. Integrative analysis of genome-wide mapping data for Smad7 and ESC self-renewal and pluripotency transcriptional regulators revealed that Smad7 co-occupies promoters of highly expressed key stemness regulators genes, by binding to a specific consensus response element NCGGAAMM. Altogether, our data establishes Smad7 as a new, integral component of the regulatory circuitry that controls ESC identity.
| S-EPMC8708723 | biostudies-literature
Project description:Inner cell mass (ICM) cells of a blastocyst, the source of embryonic stem (ES) cells, are characterized by their unique ability to give rise to all cell types in adult organisms. The epigenomes of germ cells and developing zygotes undergo extensive reprogramming to acquire such a pluripotent state. A major reprogramming event during early embryonic development is the erasure and subsequent re-establishment of patterns of methylation at the 5-position of cytosine (5mC). The recent demonstration that Ten-eleven translocation family proteins, Tet1-3 have the capacity to convert 5mC to 5-hydroxymethylcytosine (5hmC) raises the possibility that 5hmC may act as an distinct epigenetic state contributing to dynamic changes in DNA methylation and transcriptional regulation during embryonic development. In ES cells, Tet1 is highly expressed and 5hmC is present at relatively high levels compared to most differentiated cells, but the functional significance of Tet1 and 5hmC in these pluripotent cells are not clear. Recently, a flurry of papers that profile the distribution of Tet1 and/or 5hmC across the genome of mouse ES cells provide new insights into the role of Tet proteins and 5hmC in regulating expression of genes related to pluripotency and cellular differentiation. Through integrative analyses of datasets from different groups, we reveal the common Tet1 and 5hmC targets in undifferentiated mouse ES cells, which suggest that Tet1 may play a key role in orchestrating the balance between pluripotent and lineage committed states.
| S-EPMC3180185 | biostudies-literature
Project description:The nucleosome remodeling and deacetylase (NuRD) complex plays an important role in gene expression regulation, stem cell self-renewal, and lineage commitment. However, little is known about the dynamics of NuRD during cellular differentiation. Here, we study these dynamics using genome-wide profiling and quantitative interaction proteomics in mouse embryonic stem cells (ESCs) and neural progenitor cells (NPCs). We find that the genomic targets of NuRD are highly dynamic during differentiation, with most binding occurring at cell-type specific promoters and enhancers. We identify ZFP296 as an ESC-specific NuRD interactor that also interacts with the SIN3A complex. ChIP-sequencing in Zfp296 knockout (KO) ESCs reveals decreased NuRD binding both genome-wide and at ZFP296 binding sites, although this has little effect on the transcriptome. Nevertheless, Zfp296 KO ESCs exhibit delayed induction of lineage-specific markers upon differentiation to embryoid bodies. In summary, we identify an ESC-specific NuRD-interacting protein which regulates genome-wide NuRD binding and cellular differentiation.
| S-EPMC6214896 | biostudies-other
Project description:Epigenetic modification as an intrinsic fine-tune program cooperates with key transcription factors to regulate the cell fate determination. The histone acetylation participating in neural differentiation of pluripotent stem cells is expected but not well studied. Here, using acetylated histone H3 ChIP-sequencing (ChIP-seq), we demonstrate that the histone H3 acetylation level is gradually increased on the neural gene loci while decreased on the neural-inhibitory gene loci during mouse embryonic stem cell (mESC) neural differentiation. We further show that histone deacetylase 1 (HDAC1) is essential for neural commitment by targeting Nodal signaling. Thus, our study reveals a mechanism by which the epigenetic modification of histone acetylation/deacetylation interacts with extracellular signaling in mESC neural fate determination. Data were deposited in Gene Expression Omnibus (GEO) datasets under reference number GSE66025.
| S-EPMC4583617 | biostudies-literature
Project description:Mouse Embryonic Stem (ES) cells express a unique set of microRNAs (miRNAs), the miR-290-295 cluster. To elucidate the role of these miRNAs and how they integrate into the ES cell regulatory network requires identification of their direct regulatory targets. The difficulty, however, arises from the limited complementarity of metazoan miRNAs to their targets, with the interaction requiring as few as six nucleotides of the miRNA seed sequence. To identify miR-294 targets, we used Dicer1-null ES cells, which lack all endogenous mature miRNAs, and introduced just miR-294 into these ES cells. We then employed two approaches to discover miR-294 targets in mouse ES cells: transcriptome profiling using microarrays and a biochemical approach to isolate mRNA targets associated with the Argonaute2 (Ago2) protein of the RISC (RNA Induced Silencing Complex) effector, followed by RNA-sequencing. In the absence of Dicer1, the RISC complexes are largely devoid of mature miRNAs and should therefore contain only transfected miR-294 and its base-paired targets. Our data suggest that miR-294 may promote pluripotency by regulating a subset of c-Myc target genes and upregulating pluripotency-associated genes such as Lin28.
| S-EPMC2958809 | biostudies-literature
Project description:We describe a screen for cellular response to drugs that makes use of haploid embryonic stem cells. We generated ten libraries of mutants with piggyBac gene trap transposon integrations, totalling approximately 100,000 mutant clones. Random barcode sequences were inserted into the transposon vector to allow the number of cells bearing each insertion to be measured by amplifying and sequencing the barcodes. These barcodes were associated with their integration sites by inverse PCR. We exposed these libraries to commonly used cancer drugs and profiled changes in barcode abundance by Ion Torrent sequencing in order to identify mutations that conferred sensitivity. Drugs tested included conventional chemotherapeutics as well as targeted inhibitors of topoisomerases, poly(ADP-ribose) polymerase (PARP), Hsp90 and WEE1.
| S-EPMC5332012 | biostudies-literature