Single-cell CUT&amp;Tag analysis of chromatin modifications in differentiation and tumor progression

Wu, Steven J.; Furlan, Scott N.; Mihalas, Anca B.; Kaya-Okur, Hatice S; Feroze, Abdullah H.; Emerson, Samuel; Zheng, Yi; Carson, K. Jane; Cimino, Patrick J.; Keene, C. Dirk; Sarthy, Jay F.; Gottardo, Raphaël; Ahmad, Kami; Henikoff, Steven; Patel, Anoop P.

doi:10.1038/s41587-021-00865-z

Cited by 149 publications

(117 citation statements)

References 29 publications

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“…Using 10X Genomics Cell Ranger software, we obtained G4 profiles of an average of 593 MCF7 and 2,071 U2OS cells with a median of 739 and 939 unique fragments per cell, respectively (Supplementary Table S2 ). Compared to other studies, albeit in different cell types, we observed a greater number of median unique fragments per cell (~ 650–1200) compared to scCUT&Tag for transcription factors (< 300) 7 but similar to that of H3K27me3 (~ 400–6000) 8 . The tracks of normalised read coverage from aggregated single cells correlate well (r s > 0.8, Supplementary Fig.…”

Section: Resultscontrasting

confidence: 77%

“…Previously, imaging 12 – 14 and cytometry 15 studies of single cells showed that G4 formation is dynamic and cell-cycle dependent but precise genomic location and sequence context of G4s in individual cells were elusive. Here, we have adapted Cleavage Under Targets and Tagmentation (CUT&Tag) 6 – 8 for G4s and applied it to map G4 locations at single-cell level, for the first time.…”

Section: Introductionmentioning

confidence: 99%

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Single-cell mapping of DNA G-quadruplex structures in human cancer cells

Hui

Simeone

Zyner

et al. 2021

Sci Rep

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G-quadruplexes (G4s) are four-stranded DNA secondary structures that form in guanine-rich regions of the genome. G4s have important roles in transcription and replication and have been implicated in genome instability and cancer. Thus far most work has profiled the G4 landscape in an ensemble of cell populations, therefore it is critical to explore the structure–function relationship of G4s in individual cells to enable detailed mechanistic insights into G4 function. With standard ChIP-seq methods it has not been possible to determine if G4 formation at a given genomic locus is variable between individual cells across a population. For the first time, we demonstrate the mapping of a DNA secondary structure at single-cell resolution. We have adapted single-nuclei (sn) CUT&Tag to allow the detection of G4s in single cells of human cancer cell lines. With snG4-CUT&Tag, we can distinguish cellular identity from a mixed cell-type population solely based on G4 features within individual cells. Our methodology now enables genomic investigations on cell-to-cell variation of a DNA secondary structure that were previously not possible.

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Section: Resultscontrasting

confidence: 77%

Section: Introductionmentioning

confidence: 99%

Single-cell mapping of DNA G-quadruplex structures in human cancer cells

Hui

Simeone

Zyner

et al. 2021

Sci Rep

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“…The past several years have seen a rapid proliferation of techniques for measuring transcript levels ( Ramsköld et al, 2012 ; Tang et al, 2009 ), DNA methylation ( Guo et al, 2013 ), chromosome conformation ( Nagano et al, 2013 ), chromatin accessibility ( Buenrostro et al, 2015 ; Cusanovich et al, 2015 ), and protein-DNA interactions ( Bartosovic et al, 2021 ; Grosselin et al, 2019 ; Rotem et al, 2015 ; Wu et al, 2021 ) in single cells. However, little work has been done on cell-to-cell variation in TSS usage: to our knowledge, only two such studies have been performed.…”

Section: Future Prospectsmentioning

confidence: 99%

Global approaches for profiling transcription initiation

Policastro

Zentner

2021

Cell Reports Methods

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SUMMARY Transcription start site (TSS) selection influences transcript stability and translation as well as protein sequence. Alternative TSS usage is pervasive in organismal development, is a major contributor to transcript isoform diversity in humans, and is frequently observed in human diseases including cancer. In this review, we discuss the breadth of techniques that have been used to globally profile TSSs and the resulting insights into gene regulation, as well as future prospects in this area of inquiry.

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“…Single cell chromatin integration labeling followed by sequencing (scChIL-seq) ( Harada et al, 2019 ) and the single cell cleavage under targets and tagmentation (scCUT&Tag) ( Kaya-Okur et al, 2019 ) methods use specific antibodies against histone modifications and integrate a sequencing tag into double stranded DNA via transposase. This method was recently adapted to scalable nanowell and droplet-based single-cell platforms to profile polycomb group (PcG) silenced regions marked by H3K27me3 in tissue culture as well as patient samples ( Wu et al, 2021 ). Similarly, combinatorial barcoding and targeted chromatin release (COBATCH) uses the enrichment of a Protein A-Tn5 transposase fusion protein to genomic regions by specific antibodies and produced for H3K27 acetylation marks ∼12,000 unique non-duplicated reads per cell ( Wang Q. et al, 2019 ).…”

Section: Single-molecule Approaches To Monitor Histone Ptms and Effector Proteinsmentioning

confidence: 99%

Single-Molecule Techniques to Study Chromatin

Chanou

Hamperl

2021

Front. Cell Dev. Biol.

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Besides the basic organization in nucleosome core particles (NCPs), eukaryotic chromatin is further packed through interactions with numerous protein complexes including transcription factors, chromatin remodeling and modifying enzymes. This nucleoprotein complex provides the template for many important biological processes, such as DNA replication, transcription, and DNA repair. Thus, to understand the molecular basis of these DNA transactions, it is critical to define individual changes of the chromatin structure at precise genomic regions where these machineries assemble and drive biological reactions. Single-molecule approaches provide the only possible solution to overcome the heterogenous nature of chromatin and monitor the behavior of individual chromatin transactions in real-time. In this review, we will give an overview of currently available single-molecule methods to obtain mechanistic insights into nucleosome positioning, histone modifications and DNA replication and transcription analysis—previously unattainable with population-based assays.

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Single-cell CUT&Tag analysis of chromatin modifications in differentiation and tumor progression

Cited by 149 publications

References 29 publications

Single-cell mapping of DNA G-quadruplex structures in human cancer cells

Single-cell mapping of DNA G-quadruplex structures in human cancer cells

Global approaches for profiling transcription initiation

Single-Molecule Techniques to Study Chromatin

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