Project description:We determined genome-wide nucleosome occupancy in mouse embryonic stem cells and their neural progenitor and embryonic fibroblast counterparts to assess features associated with nucleosome positioning during lineage commitment. Cell type and protein specific binding preferences of transcription factors to sites with either low (e.g. Myc, Klf4, Zfx) or high (e.g. Nanog, Oct4 and Sox2) nucleosome occupancy as well as complex patterns for CTCF were identified. Nucleosome depleted regions around transcription start and termination sites were broad and more pronounced for active genes, with distinct patterns for promoters classified according to their CpG-content or histone methylation marks. Throughout the genome nucleosome occupancy was dependent on the presence of certain histone methylation or acetylation modifications. In addition, the average nucleosome-repeat length increased during differentiation by 5-7 base pairs, with local variations for specific genomic regions. Our results reveal regulatory mechanisms of cell differentiation acting through nucleosome repositioning. We have determined genome-wide nucleosome position maps in mouse embryonic stem cells (ESCs), neural progenitor cells (NPCs) derived from these ESCs by retinoic acid induced differentiation as well as mouse embryonic fibroblasts (MEFs) from the corresponding mouse strain
Project description:The nucleosome is a fundamental unit of chromatin architecture, but it has been under-evaluated during the analysis of histone modification using ChIP-Seq. Thus, we developed a novel approach for defining histone modification at single nucleosome resolution by synergistic analyses of histone modification data from ChIP-Seq and high-resolution nucleosome mapping data from native MNase-Seq using a PCR-free strategy. With this approach, we have generated histone modification data at the single nucleosome resolution in human embryonic stem cells, normal hepatocytes, and liver cancer cells of both genders, and performed quantitative analysis of epigenetic regulation in stem cell differentiation into hepatocytes and neoplastic transformation of liver cancer.
Project description:We determined genome-wide nucleosome occupancy in mouse embryonic stem cells and their neural progenitor and embryonic fibroblast counterparts to assess features associated with nucleosome positioning during lineage commitment. Cell type and protein specific binding preferences of transcription factors to sites with either low (e.g. Myc, Klf4, Zfx) or high (e.g. Nanog, Oct4 and Sox2) nucleosome occupancy as well as complex patterns for CTCF were identified. Nucleosome depleted regions around transcription start and termination sites were broad and more pronounced for active genes, with distinct patterns for promoters classified according to their CpG-content or histone methylation marks. Throughout the genome nucleosome occupancy was dependent on the presence of certain histone methylation or acetylation modifications. In addition, the average nucleosome-repeat length increased during differentiation by 5-7 base pairs, with local variations for specific genomic regions. Our results reveal regulatory mechanisms of cell differentiation acting through nucleosome repositioning.
Project description:We determined genome-wide nucleosome occupancy in mouse embryonic stem cells and their neural progenitor and embryonic fibroblast counterparts to assess features associated with nucleosome positioning during lineage commitment. Cell type and protein specific binding preferences of transcription factors to sites with either low (e.g. Myc, Klf4, Zfx) or high (e.g. Nanog, Oct4 and Sox2) nucleosome occupancy as well as complex patterns for CTCF were identified. Nucleosome depleted regions around transcription start and termination sites were broad and more pronounced for active genes, with distinct patterns for promoters classified according to their CpG-content or histone methylation marks. Throughout the genome nucleosome occupancy was dependent on the presence of certain histone methylation or acetylation modifications. In addition, the average nucleosome-repeat length increased during differentiation by 5-7 base pairs, with local variations for specific genomic regions. Our results reveal regulatory mechanisms of cell differentiation acting through nucleosome repositioning.
Project description:We determined genome-wide nucleosome occupancy in mouse embryonic stem cells and their neural progenitor and embryonic fibroblast counterparts to assess features associated with nucleosome positioning during lineage commitment. Cell type and protein specific binding preferences of transcription factors to sites with either low (e.g. Myc, Klf4, Zfx) or high (e.g. Nanog, Oct4 and Sox2) nucleosome occupancy as well as complex patterns for CTCF were identified. Nucleosome depleted regions around transcription start and termination sites were broad and more pronounced for active genes, with distinct patterns for promoters classified according to their CpG-content or histone methylation marks. Throughout the genome nucleosome occupancy was dependent on the presence of certain histone methylation or acetylation modifications. In addition, the average nucleosome-repeat length increased during differentiation by 5-7 base pairs, with local variations for specific genomic regions. Our results reveal regulatory mechanisms of cell differentiation acting through nucleosome repositioning.
Project description:Embryonic stem cells (ESCs) and induced-pluripotent stem cells (iPSCs) self-renew and differentiate into an array of cell types in vitro and in vivo. A complex network of genetic and epigenetic pathways regulates the self-renewal and differentiation of these pluripotent cells, and the structure and covalent modifications of chromatin play a prominent role in this process. We examine nucleosome occupancy in mouse and human embryonic stem cells (ESCs), induced-pluripotent stem cells (iPSCs), and differentiated cell types using MNase-seq. To address variability inherent in this technique, we developed a bioinformatic approach that enabled the identification of regions of difference (RoD) in nucleosome occupancy between pluripotent and somatic cells. The majority of changes in nucleosomal signatures that occur in differentiation are reset during reprogramming. We conclude that changes in nucleosome occupancy are a hallmark of pluripotency and likely identify key regulatory regions that play a role in determining cell identity. A six chip study using total RNA recovered from three cell types with 2 replicates each
Project description:The SWI/SNF ATP-dependent chromatin remodeler is a master regulator of the epigenome; controlling pluripotency, cell fate determination and differentiation. There is a sparsity of information on the autoregulation of SWI/SNF, the domains involved and their mode of action. We find a DNA or RNA binding module conserved from yeast to humans located in the C-terminus of the catalytic subunit of SWI/SNF called the AT-hook that positively regulates the chromatin remodeling activity of yeast and mouse SWI/SNF. The AT-hook in yeast SWI/SNF interacts with the SnAC and ATPase domains, which after binding to nucleosome switches to contacting the N-terminus of histone H3. Deletion of the AT-hooks in yeast SWI/SNF and mouse esBAF complexes reduces the remodeling activity of SWI/SNF without affecting complex integrity or its recruitment to nucleosomes. In addition, deletion of the AT-hook impairs the ATPase and nucleosome mobilizing activities of yeast SWI/SNF without disrupting the interactions of the ATPase domain with nucleosomal DNA. The AT-hook is also important in vivo for SWI/SNF-dependent response to amino acid starvation in yeast and for cell lineage priming in mouse embryonic stem cells. In summary, the AT-hook is shown to be an evolutionarily conserved autoregulatory domain of SWI/SNF that positively regulates SWI/SNF both in vitro and in vivo.
Project description:Embryonic stem cells (ESCs) and induced-pluripotent stem cells (iPSCs) self-renew and differentiate into an array of cell types in vitro and in vivo. A complex network of genetic and epigenetic pathways regulates the self-renewal and differentiation of these pluripotent cells, and the structure and covalent modifications of chromatin play a prominent role in this process. We examine nucleosome occupancy in mouse and human embryonic stem cells (ESCs), induced-pluripotent stem cells (iPSCs), and differentiated cell types using MNase-seq. To address variability inherent in this technique, we developed a bioinformatic approach that enabled the identification of regions of difference (RoD) in nucleosome occupancy between pluripotent and somatic cells. The majority of changes in nucleosomal signatures that occur in differentiation are reset during reprogramming. We conclude that changes in nucleosome occupancy are a hallmark of pluripotency and likely identify key regulatory regions that play a role in determining cell identity. Micrococcal nuclease digestion of chromatin in crosslinked cells was followed by high throughput sequencing. These experiments were carried out in four mouse cell types: embryonic stem cells, induced pluripotent stem cells, somatic tail-tip fibroblasts and liver, and three human cell types: H1-OGN embryonic stem cells, H1-OGN induced pluripotent stem cells, and fibroblasts differentiated from H1-OGN ESCs. At least two replicates performed with each cell type were sequenced.
Project description:Embryonic stem cells (ESCs) and induced-pluripotent stem cells (iPSCs) self-renew and differentiate into an array of cell types in vitro and in vivo. A complex network of genetic and epigenetic pathways regulates the self-renewal and differentiation of these pluripotent cells, and the structure and covalent modifications of chromatin play a prominent role in this process. We examine nucleosome occupancy in mouse and human embryonic stem cells (ESCs), induced-pluripotent stem cells (iPSCs), and differentiated cell types using MNase-seq. To address variability inherent in this technique, we developed a bioinformatic approach that enabled the identification of regions of difference (RoD) in nucleosome occupancy between pluripotent and somatic cells. The majority of changes in nucleosomal signatures that occur in differentiation are reset during reprogramming. We conclude that changes in nucleosome occupancy are a hallmark of pluripotency and likely identify key regulatory regions that play a role in determining cell identity.
Project description:Embryonic stem cells (ESCs) and induced-pluripotent stem cells (iPSCs) self-renew and differentiate into an array of cell types in vitro and in vivo. A complex network of genetic and epigenetic pathways regulates the self-renewal and differentiation of these pluripotent cells, and the structure and covalent modifications of chromatin play a prominent role in this process. We examine nucleosome occupancy in mouse and human embryonic stem cells (ESCs), induced-pluripotent stem cells (iPSCs), and differentiated cell types using MNase-seq. To address variability inherent in this technique, we developed a bioinformatic approach that enabled the identification of regions of difference (RoD) in nucleosome occupancy between pluripotent and somatic cells. The majority of changes in nucleosomal signatures that occur in differentiation are reset during reprogramming. We conclude that changes in nucleosome occupancy are a hallmark of pluripotency and likely identify key regulatory regions that play a role in determining cell identity.