DSP-MNase-DNA-seq
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA914212 | ENA |
REPOSITORIES: ENA
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Project description:Mechanotransduction leads to a variety of biological responses including changes in cell shape, migration, tissue development, immune responses, and gene expression. Dysregulation of mechanotransduction is implicated in the progression of various diseases such as cancer and cardiovascular diseases. The actin cytoskeleton plays a crucial role in transmitting mechanical stimuli. Actin filaments, essential for cell motility and shape changes, respond to mechanical cues by remodeling, influencing gene expression via the LINC complex and mechanosensitive transcription factors. This study employs the DSP-MNase DNA-seq method to explore the relationship between cellular mechanosensing and chromatin architecture.
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Project description:Characterization of open chromatin sensitive to actin polymerization using DSP-MNase-DNA-seq
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Project description:Chromatin is the highly complex structure consisting of DNA, histone, and trans-acting factors (TAFs). Binding of TAF proteins to specific chromosomal loci regulates gene expression. Therefore, profiling chromatin is essential to understand gene expression mechanism. Although various chromatin profiling techniques using genome sequencing are available, high-throughput proteomics of TAFs on the chromatin is still challenging and their relationship with DNA sequences needs deeper understanding. Here, we develop a new method to extract the open-chromatin region and elucidate the DNA which is mainly regulated by TAFs in low and high density cell by DNA-seq.
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Project description:We developed a new method to extract open-chromatins that contain trans-acting factors (TAFs)-DNA complex crosslinked with dithiobis(succinimidyl propionate) (DSP). Using this method we identified CBFB as a cell density-sensitive TAF, which translocates between the cytoplasm and the nucleus in cell density-dependent manner. Here, we demonstrate that the extracted TAF/DNA complex is also amenable to ChIP-seq analysis.
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Project description:MNase-seq Experiments from Calorie Restricted and Non-Restricted Yeast from WT, ISW2DEL and ISW2K215R strains We used MNase-seq to study genome-wide nucleosome positions under Calorie Restricted and Non-restricted Saccharomyces cerevisiae
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Project description:CBFB-DSP-ChIPseq and YAP-DSP-ChIPseq
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Project description:Chromatin mapping using micrococcal nuclease (MNase) has been the standard tool for mapping nucleosomes for >40 years. When coupled with DNA sequencing, MNase-seq can provide base-pair-resolution nucleosome maps. However, determining nucleosome occupancy using MNase-seq has been hampered by its aggressive endo-/exo-nuclease activities, whereby cleavages within linker regions produce oligo- and mono-nucleosomes whereas cleavages within nucleosomes destroy them. Here we introduce a theoretical framework for predicting nucleosome occupancies and an experimental protocol with appropriate spike-in normalization that confirms our theory and provides accurate occupancy levels over an MNase digestion time-course. As expected, DNaseI hypersensitive sites and transcription units are digested by MNase at elevated rates, and the apparent deficiency of nucleosomes at 3’ ends of Drosophila genes is an artifact of MNase preference for AT-rich DNA. Surprisingly, we observed no overall differences between Drosophila euchromatin and heterochromatin, which implies that heterochromatin compaction does not render nucleosomal DNA less accessible than euchromatin.
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Project description:We performed ChIP-Seq for hallmark TFs (Ets1, Runx1), histone modification marks (H3K4me1, H3K4me2, H3K4me3, H3K27me3, H3K36me3), total RNA Pol II, short RNA-Seq as well as nucleosome mapping mainly in murine Rag2 -/- thymocytes. We also performed ChIP-Seq for E47 as well as nucleosome mapping, gene expression microarray analysis in CD4+ CD8+ DP thymocytes. Overall, we find a key role for the transcription factor Ets1, contributing towards alpha beta T cell lineage commitment via differential transactivation of stage-specific genes orchestrated by dynamic, co-association -mediated chromatin remodeling, as well as transcription dependent generation of a specialized chromatin structure at the TCR beta locus. Genome-wide analysis via ChIP-Seq for Ets1, Runx1, total RNA Pol II binding, H3K4me1, H3K4me2, H3K4me3, H3K27me3, H3K36me3, short RNA-Seq, Mnase-Seq in murine Rag2 -/- thymocytes, ChIP-Seq for E47, Mnase-Seq and gene expression microarray analysis in DP thymocytes This Series represents Mnase-Seq data.
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Project description:MNase-seq was performed in order to analyze changes in nucleosomal occupancy after depletion of CTCF/P190 and ISWI from Drosophila S2 cells MNase-seq from Drosophila S2 nuclei after CTCF/CP190 or ISWI-specific RNAi treatment
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