Chromatin In Situ Proximity (ChrISP): Single-cell analysis of chromatin proximities at a high resolution
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE42009 | GEO | 2014/02/20
SECONDARY ACCESSION(S): PRJNA178758
REPOSITORIES: GEO
Similar Datasets
Project description:Mammals contain over 200 different cell types, yet nearly all have the same genomic DNA sequence. It is a key question in biology how the genetic instructions in DNA are selectively interpreted by cells to specify various transcriptional programs and therefore cellular identity. The structural and functional organization of chromatin governs the transcriptional state of individual genes. To understand how genomic loci adopt different levels of gene expression, it is critical to characterize all local chromatin factors as well as long-range interactions in the 3D nuclear compartment. Much of our current knowledge regarding protein interactions in a chromatin context is based on affinity purification of chromatin components coupled to mass spectrometry (AP-MS). AP-MS has been invaluable to map strong protein-protein interactions in the nucleus. However, the interaction is detected after cell lysis and biochemical enrichment, allowing for loss or gain of false positive or negative interaction partners. Recently, proximity-dependent labeling methods have emerged as powerful tools for studying chromatin in its native context. These methods take advantage of engineered enzymes that are fused to a chromatin factor of interest and can directly label all factors in proximity. Subsequent pull-down assays followed by mass spectrometry or sequencing approaches provide a comprehensive snapshot of the proximal chromatin interactome. By combining this method with dCas9, this approach can also be extended to study chromatin at specific genomic loci. Here, we review and compare current proximity-labeling approaches available for studying chromatin, with a particular focus on new emerging technologies that can provide important insights into the transcriptional and chromatin interaction networks essential for cellular identity.
| S-EPMC7235407 | biostudies-literature
Project description:3D structure of chromatin is thought to be critical for the regulation of gene expression during development. Here we employ the Micro-C assay to achieve 100 bp resolution for the genome organization of Drosophila melanogaster throughout the first half of embryogenesis. The resulting contact maps enable the identification of fine-scaled structures such as individual loops and boundaries delineating TADs. We observe that 3D structures form prior to zygotic genome activation in precellular embryos, and many of these organizational features persist during successive mitotic cycles. 3D structures are classified through the use of 149 ChIP-seq datasets. We present evidence for specialized elements associated with housekeeping genes that are enriched for BEAF-32 but not CTCF. We also characterize a distinct class of elements associated with developmental gene regulation that are enriched with GAF and Zld binding. This binding is maintained during mitotic cycles and exhibits evolutionary conservation of both sequence and 3D structures. In sum, this work provides a comprehensive genome-wide characterization at unprecedented resolution of the role of 3D chromatin organization in gene regulation during development. We propose that the 3D organization of the pre-cellular embryo facilitates deployment of the developmental control genes defining the Drosophila fate map.
2024-12-06 | GSE265818 | GEO
Project description:We assessed whether changes in DNA methylation were associated with the regulation of ISC signature genes. Methylated regions of DNA from whole genome were isolated using Methyl Binding Domain pull-down followed by sequencing (MBD-seq). The size of DNA fragments was between 120 to 170 bp, which provides a resolution at nucleosome level.
2017-03-29 | GSE89682 | GEO
Project description:Rapid advances in high-throughput DNA sequencing technologies are accelerating the pace of research into personalized medicine. While methods for variant discovery and genotyping from whole genome sequencing (WGS) datasets have been well established, linking variants together into a single haplotype remains a challenge. An understanding of complete haplotypes of an individual will help clarify the consequences of inheriting multiple alleles in combination, identify novel disease associations, and augment studies of gene regulation. Although numerous methods have been developed to reconstruct haplotypes from WGS data, chromosome-span haplotypes at high resolution have been difficult to obtain. Here we present a novel method to accurately reconstruct chromosome-span haplotypes from proximity-ligation and DNA shotgun sequencing. We demonstrate the utility of this approach in producing high-resolution chromosome-span haplotype phasing in mouse and human. While proximity-ligation based methods were originally designed to investigate spatial organization of the genome, our results lend support for their use as a general tool for haplotyping in the future. Hi-C experiments in two replicates of Human GM12878 Lymphoblastoid cells and two replicates of F123 mouse ES cells (4 total samples)
2013-11-03 | E-GEOD-48592 | biostudies-arrayexpress
Project description:Rapid advances in high-throughput DNA sequencing technologies are accelerating the pace of research into personalized medicine. While methods for variant discovery and genotyping from whole genome sequencing (WGS) datasets have been well established, linking variants together into a single haplotype remains a challenge. An understanding of complete haplotypes of an individual will help clarify the consequences of inheriting multiple alleles in combination, identify novel disease associations, and augment studies of gene regulation. Although numerous methods have been developed to reconstruct haplotypes from WGS data, chromosome-span haplotypes at high resolution have been difficult to obtain. Here we present a novel method to accurately reconstruct chromosome-span haplotypes from proximity-ligation and DNA shotgun sequencing. We demonstrate the utility of this approach in producing high-resolution chromosome-span haplotype phasing in mouse and human. While proximity-ligation based methods were originally designed to investigate spatial organization of the genome, our results lend support for their use as a general tool for haplotyping in the future.
2013-11-03 | GSE48592 | GEO
Project description:Proof-of-concept of a new method involving the limited digestion and subsequent ligation of intramolecular RNA structures in situ followed by deep sequencing Proof-of-concept of RPL in S. cerevisiae and H. sapiens tissue culture
2015-08-01 | E-GEOD-69472 | biostudies-arrayexpress
Project description:Advanced stem cell-based in vitro models of embryogenesis offer great promise to decipher developmental programs with high-throughput and resolution. However, the rapid changes in morphology and cell state that underly these models mask multiple higher order parameters, including the regulative balance between differentiation and self-renewal that govern transient progenitor cell populations. Here, we apply a continuous single-cell Cas9-based molecular recorder to mouse embryonic stem cells, which enables the recovery of can reproducibly recovere and high-content information regarding the historical relationship between individual cells. We apply our system to Trunk-Like Structures (TLS), a complex in vitro model of embryogenesis that captures differentiation and morphological dynamics from pluripotency exit to the specification of the somitic and neural lineages. By recording cell lineages alongside the transcriptional identities of extant cells, we are able to build high-resolution phylogenetic trees, identify clonal subpopulations, and reveal recover the step-wisestepwise series of events that govern germ layer specification. Moreover, we examine are able to functionally characterize the specification and maintenance of multipotent neuro-mesodermal progenitor (NMP) populations as they coordinate self-renewal and commitment into more specialized cell types, including how thehow highly disparateplastic individual nature of individual branches trajectories ultimately aggregateultimately aggregate to ensure to generate the generation of highly consistent embryonic structures. Finally, we demonstrate the potential of this technology for high-throughput perturbation screening by combining small molecule treatment with individual structure labeling, allowing us to we demonstrate that these parameters can be independently tuned to modulate TLS morphology, including in reversible ways that extend the duration through which NMPs can contribute to both lineages of the embryonic trunkconnect perturbation of early differentiation pathways to progenitor cell behavior and structure morphology. Our work represents the first continuous molecular recording approach applied to dynamic in vitro models, a toolkit that can be combined with inhibitor libraries or other screening modalities to uncover the genetic and epigenetic mechanisms that enable highly orchestrated and robust changes in cell state composition during critical windows of morphogenesis.
2024-03-05 | GSE220949 | GEO
Project description:We report Proximity Ligation Assisted ChIP-sequencing (PLAC-seq), a method for comprehensive detection of long-range interactions associated with proteins of interest. PLAC-seq requires up to 500-fold less starting material compared to ChIA-PET and using experimentally determined input as control precisely reveals protein associated interaction upto single-element resolution. Application of PLAC-seq to mouse embryonic stem cells revealed a comprehensive map of regulatory interactions.
2016-11-15 | GSE86150 | GEO
Project description:Mycobacterium polyketide synthase 13 (Pks13) is a protein complex comprised of a closely symmetric parallel dimer of chains, each encoding several enzymatic and transport functions. This module carries out the condensation of two different very long chain fatty acids to produce mycolic acids that are essential components of the mycobacterial cell wall. Here, we use cryo-EM, XL-MS, and integrative modeling to determine the structure and domain trajectories of the dimeric multi-enzyme Pks13 purified endogenously from mycobacteria under normal growth conditions. Structures of the multi-domain assembly revealed by cryogenic electron microscopy (cryoEM) define the ketosynthase (KS), linker, and acyltransferase (AT) domains, each at atomic resolution (1.8Å), with bound substrates defined at 2.4Å and 2.9Å resolution. Image classification reveals two distinct structures with alternate locations of the N-terminal acyl carrier protein (termed ACP1a, ACP1b) seen at 3.6Å and 4.6Å resolution respectively. These two structures suggest plausible intermediate states, related by a ~60Å movement of ACP1, on the pathway for substrate delivery from the fatty acyl-ACP ligase (FadD32) to the ketosynthase domain. The linking sequence between ACP1 and the KS includes an 11 amino acid sequence with 6 negatively charged side chains that lies in different positively charged grooves on the KS in ACP1a versus ACP1b structures. This charge complementarity between the extended chain and the grooves suggests some stabilization of these two distinct orientations. Other domains are visible at lower resolution and indicate flexibility relative to the KS-AT core. The chemical structures of three bound endogenous long chain fatty acid substrates with their proximal regions defined in the structures were determined by electrospray ionization mass spectrometry. The domain proximities were probed by chemical cross-linking and identified by mass spectrometry (XL-MS). These were incorporated into integrative structure modeling to define multiple domain configurations that transport the very long fatty acid chains throughout the multistep Pks13 mediated synthetic pathway.
2023-06-13 | PXD033471 | Pride
Project description:Anticancer drug therapy generally elicits the drug tolerance after long-term treatment. Recent studies suggested that 3D chromatin structures of cancer cells were intimately linked to drug resistance. However, 3D chromatin structures in drug-tolerant cancer cells at single-cell resolution haven’t been elucidated. Here we performed single-cell Hi-C (scHi-C) analysis to examine the 3D chromatin structures in three stages of breast cancer cells.
2024-08-14 | GSE194308 | GEO