Browse
Submit Data
Databases
API
Help

Dataset Information

43 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Chromatin and transcription factor profiling in rare stem cell populations using CUT&Tag.

ABSTRACT: Muscle stem cells (MuSCs) are a rare stem cell population that provides myofibers with a remarkable capacity to regenerate after tissue injury. Here, we have adapted the Cleavage Under Target and Tagmentation technology to the mapping of the chromatin landscape and transcription factor binding in 50,000 activated MuSCs isolated from injured mouse hindlimb muscles. We have applied this same approach to human CD34⁺ hematopoietic stem and progenitor cells. This protocol could be adapted to any rare stem cell population. For complete details on the use and execution of this protocol, please refer to Robinson et al. (2021).

SUBMITTER: Li Y

PROVIDER: S-EPMC8384913 | biostudies-literature | 2021 Sep

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

Chromatin and transcription factor profiling in rare stem cell populations using CUT&Tag.

Li Yuefeng Y Nakka Kiran K Olender Thomas T Gingras-Gelinas Philippe P Wong Matthew Man-Kin MM Robinson Daniel C L DCL Bandukwala Hina H Palii Carmen G CG Neyret Odile O Brand Marjorie M Blais Alexandre A Dilworth F Jeffrey FJ

STAR protocols 20210819 3

Muscle stem cells (MuSCs) are a rare stem cell population that provides myofibers with a remarkable capacity to regenerate after tissue injury. Here, we have adapted the Cleavage Under Target and Tagmentation technology to the mapping of the chromatin landscape and transcription factor binding in 50,000 activated MuSCs isolated from injured mouse hindlimb muscles. We have applied this same approach to human CD34<sup>+</sup> hematopoietic stem and progenitor cells. This protocol could be adapted ...[more]

PMID: 34467227

Similar Datasets

Multimodal chromatin profiling using nanobody-based single-cell CUT&Tag.

Project description:Probing histone modifications at a single-cell level in thousands of cells has been enabled by technologies such as single-cell CUT&Tag. Here we describe nano-CUT&Tag (nano-CT), which allows simultaneous mapping of up to three epigenomic modalities at single-cell resolution using nanobody-Tn5 fusion proteins. Multimodal nano-CT is compatible with starting materials as low as 25,000-200,000 cells and has significantly higher sensitivity and number of fragments per cell than single-cell CUT&Tag. We use nano-CT to simultaneously profile chromatin accessibility, H3K27ac, and H3K27me3 in juvenile mouse brain, allowing for discrimination of more cell types and states than unimodal single-cell CUT&Tag. We also infer chromatin velocity between assay for transposase-accessible chromatin (ATAC) and H3K27ac in the oligodendrocyte lineage and deconvolute H3K27me3 repressive states, finding two sequential waves of H3K27me3 repression at distinct gene modules during oligodendrocyte lineage progression. Given its high resolution, versatility, and multimodal features, nano-CT allows unique insights in epigenetic landscapes in complex biological systems at the single-cell level.

| S-EPMC10264246 | biostudies-literature

Efficient chromatin profiling of H3K4me3 modification in cotton using CUT&Tag.

Project description:BackgroundIn 2019, Kaya-Okur et al. reported on the cleavage under targets and tagmentation (CUT&Tag) technology for efficient profiling of epigenetically modified DNA fragments. It was used mainly for cultured cell lines and was especially effective for small samples and single cells. This strategy generated high-resolution and low-background-noise chromatin profiling data for epigenomic analysis. CUT&Tag is well suited to be used in plant cells, especially in tissues from which small samples are taken, such as ovules, anthers, and fibers.ResultsHere, we present a CUT&Tag protocol step by step using plant nuclei. In this protocol, we quantified the nuclei that can be used in each CUT&Tag reaction, and compared the efficiency of CUT&Tag with chromatin immunoprecipitation with sequencing (ChIP-seq) in the leaves of cotton. A general workflow for the bioinformatic analysis of CUT&Tag is also provided. Results indicated that, compared with ChIP-seq, the CUT&Tag procedure was faster and showed a higher-resolution, lower-background signal than did ChIP.ConclusionA CUT&Tag protocol has been refined for plant cells using intact nuclei that have been isolated.

| S-EPMC7460760 | biostudies-literature

Efficient low-cost chromatin profiling with CUT&Tag.

Project description:We recently introduced Cleavage Under Targets & Tagmentation (CUT&Tag), an epigenomic profiling strategy in which antibodies are bound to chromatin proteins in situ in permeabilized nuclei. These antibodies are then used to tether the cut-and-paste transposase Tn5. Activation of the transposase simultaneously cleaves DNA and adds adapters ('tagmentation') for paired-end DNA sequencing. Here, we introduce a streamlined CUT&Tag protocol that suppresses DNA accessibility artefacts to ensure high-fidelity mapping of the antibody-targeted protein and improves the signal-to-noise ratio over current chromatin profiling methods. Streamlined CUT&Tag can be performed in a single PCR tube, from cells to amplified libraries, providing low-cost genome-wide chromatin maps. By simplifying library preparation CUT&Tag requires less than a day at the bench, from live cells to sequencing-ready barcoded libraries. As a result of low background levels, barcoded and pooled CUT&Tag libraries can be sequenced for as little as $25 per sample. This enables routine genome-wide profiling of chromatin proteins and modifications and requires no special skills or equipment.

| S-EPMC8318778 | biostudies-literature

Automated CUT&Tag profiling of chromatin heterogeneity in mixed-lineage leukemia.

Project description:Acute myeloid and lymphoid leukemias often harbor chromosomal translocations involving the KMT2A gene, encoding the KMT2A lysine methyltransferase (also known as mixed-lineage leukemia-1), and produce in-frame fusions of KMT2A to other chromatin-regulatory proteins. Here we map fusion-specific targets across the genome for diverse KMT2A oncofusion proteins in cell lines and patient samples. By modifying CUT&Tag chromatin profiling for full automation, we identify common and tumor-subtype-specific sites of aberrant chromatin regulation induced by KMT2A oncofusion proteins. A subset of KMT2A oncofusion-binding sites are marked by bivalent (H3K4me3 and H3K27me3) chromatin signatures, and single-cell CUT&Tag profiling reveals that these sites display cell-to-cell heterogeneity suggestive of lineage plasticity. In addition, we find that aberrant enrichment of H3K4me3 in gene bodies is sensitive to Menin inhibitors, demonstrating the utility of automated chromatin profiling for identifying therapeutic vulnerabilities. Thus, integration of automated and single-cell CUT&Tag can uncover epigenomic heterogeneity within patient samples and predict sensitivity to therapeutic agents.

| S-EPMC8571097 | biostudies-literature

Identification of chromatin states during zebrafish gastrulation using CUT&RUN and CUT&Tag.

Project description:BackgroundCell fate decisions are governed by interactions between sequence-specific transcription factors and a dynamic chromatin landscape. Zebrafish offer a powerful system for probing the mechanisms that drive these cell fate choices, especially in the context of early embryogenesis. However, technical challenges associated with conventional methods for chromatin profiling have slowed progress toward understanding the exact relationships between chromatin changes, transcription factor binding, and cellular differentiation during zebrafish embryogenesis.ResultsTo overcome these challenges, we adapted the chromatin profiling methods Cleavage Under Targets and Release Using Nuclease (CUT&RUN) and CUT&Tag for use in zebrafish and applied these methods to generate high-resolution enrichment maps for H3K4me3, H3K27me3, H3K9me3, RNA polymerase II, and the histone variant H2A.Z using tissue isolated from whole, mid-gastrula stage embryos. Using this data, we identify a subset of genes that may be bivalently regulated during both zebrafish and mouse gastrulation, provide evidence for an evolving H2A.Z landscape during embryo development, and demonstrate the effectiveness of CUT&RUN for detecting H3K9me3 enrichment at repetitive sequences.ConclusionsOur results demonstrate the power of combining CUT&RUN and CUT&Tag methods with the strengths of the zebrafish system to define emerging chromatin landscapes in the context of vertebrate embryogenesis.

| S-EPMC8976701 | biostudies-literature

Spatial-CUT&Tag: Spatially resolved chromatin modification profiling at the cellular level.

Project description:Spatial omics emerged as a new frontier of biological and biomedical research. Here, we present spatial-CUT&Tag for spatially resolved genome-wide profiling of histone modifications by combining in situ CUT&Tag chemistry, microfluidic deterministic barcoding, and next-generation sequencing. Spatially resolved chromatin states in mouse embryos revealed tissue-type-specific epigenetic regulations in concordance with ENCODE references and provide spatial information at tissue scale. Spatial-CUT&Tag revealed epigenetic control of the cortical layer development and spatial patterning of cell types determined by histone modification in mouse brain. Single-cell epigenomes can be derived in situ by identifying 20-micrometer pixels containing only one nucleus using immunofluorescence imaging. Spatial chromatin modification profiling in tissue may offer new opportunities to study epigenetic regulation, cell function, and fate decision in normal physiology and pathogenesis.

| S-EPMC7612972 | biostudies-literature

High-Resolution Chromatin Profiling Using CUT&RUN.

Project description:Determining the genomic location of DNA-binding proteins is essential to understanding their function. Cleavage Under Targets and Release Using Nuclease (CUT&RUN) is a powerful method for mapping protein-DNA interactions at high resolution. In CUT&RUN, a recombinant protein A-microccocal nuclease (pA-MN) fusion is recruited by an antibody targeting the chromatin protein of interest; this can be done with either uncrosslinked or formaldehyde-crosslinked cells. DNA fragments near sites of antibody binding are released from the insoluble bulk chromatin through endonucleolytic cleavage and used to build barcoded DNA-sequencing libraries that can be sequenced in pools of at least 30. Therefore, CUT&RUN provides an alternative to ChIP-seq approaches for mapping chromatin proteins, which typically have relatively high signal-to-noise ratios, while using fewer cells and at a lower cost. Here, we describe the methods for performing CUT&RUN, generating DNA-sequencing libraries, and analyzing the resulting datasets. © 2019 by John Wiley & Sons, Inc.

| S-EPMC6422702 | biostudies-literature

Protocol to capture transcription factor-mediated 3D chromatin interactions using affinity tag-based BL-HiChIP.

Project description:Pioneer transcription factors (TFs) can directly establish higher-order chromatin interactions to instruct gene transcription. Here, we present a protocol for capturing TF-mediated 3D chromatin interactions using affinity tag-based bridge linker (BL)-Hi-chromatin immunoprecipitation (HiChIP). We describe steps for constructing FLAG-tagged TF, performing BL-HiChIP, and preparing the library. We then detail procedures for sequencing, data analysis, and quality control. This protocol has potential applications in 3D chromatin analysis centered on any specific TF in any type of cells without the need of optimal antibodies. For complete details on the use and execution of this protocol, please refer to Ren et al. (2022).1.

| S-EPMC10519843 | biostudies-literature

Untargeted CUT&Tag and BG4 CUT&Tag are both enriched at G-quadruplexes and accessible chromatin.

Project description:G-quadruplex DNA structures (G4s) form within single-stranded DNA in nucleosome-free chromatin. As G4s modulate gene expression and genomic stability, genome-wide mapping of G4s has generated strong research interest. Recently, the Cleavage Under Targets and Tagmentation (CUT&Tag) method was performed with the G4-specific BG4 antibody to target Tn5 transposase to G4s. While this method generated a novel high-resolution map of G4s, we unexpectedly observed a strong correlation between the genome-wide signal distribution of BG4 CUT&Tag and accessible chromatin. To examine whether untargeted Tn5 cutting at accessible chromatin contributes to BG4 CUT&Tag signal, we examined the genome-wide distribution of signal from untargeted (i.e. negative control) CUT&Tag datasets. We observed that untargeted CUT&Tag signal distribution was highly similar to both that of accessible chromatin and of BG4 CUT&Tag. We also observed that BG4 CUT&Tag signal increased at mapped G4s, but this increase was accompanied by a concomitant increase in untargeted CUT&Tag at the same loci. Consequently, enrichment of BG4 CUT&Tag over untargeted CUT&Tag was not increased at mapped G4s. These results imply that either the vast majority of accessible chromatin regions contain mappable G4s or that the presence of G4s within accessible chromatin cannot reliably be determined using BG4 CUT&Tag alone.

| S-EPMC11463444 | biostudies-literature

Multi-CUT&Tag to simultaneously profile multiple chromatin factors.

Project description:Genome-wide chromatin mapping approaches typically focus on one protein at a time. We recently developed multi-CUT&Tag, which enables simultaneous mapping of multiple chromatin proteins in the same single cells or pools of cells. Using barcoded adapters loaded onto antibody-protein A-Tn5 transposase complexes, multi-CUT&Tag marks the locations of each chromatin protein and directly detects colocalization of different proteins in the same cell(s). Although slightly more laborious than CUT&Tag, multi-CUT&Tag provides a powerful option for generating multi-factor maps for epigenomic profiling. For complete details on the use and execution of this protocol, please refer to Gopalan et al. (2021).

| S-EPMC8783141 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data