Chromatin tracing and multiplexed imaging of nucleome architectures (MINA) and RNAs in single mammalian cells and tissue

Liu, Miao; Yang, Bing; Hu, Mengwei; Radda, Jonathan S. D.; Chen, Yanbo; Jin, Shengyan; Cheng, Yubao; Wang, Siyuan

doi:10.1038/s41596-021-00518-0

Cited by 18 publications

(19 citation statements)

References 52 publications

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“…This selected 840-kb region spans a TAD boundary, based on previously published IMR-90 Hi-C datasets 12,16 . We partitioned this region of interest into consecutive 30-kb segments, labeled and imaged each segment following our previously established chromatin tracing protocol 30,41 . In brief, we first labeled and imaged the entire region with a library of dye-labeled primary oligonucleotide probes.…”

Section: Resultsmentioning

confidence: 99%

TAD-like single-cell domain structures exist on both active and inactive X chromosomes and persist under epigenetic perturbations

Cheng

Liu

et al. 2021

Preprint

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Background: Topologically associating domains (TADs) are important building blocks of three-dimensional genome architectures. The formation of TADs was shown to depend on cohesin in a loop-extrusion mechanism. Recently, advances in an image-based spatial genomics technique known as chromatin tracing led to the discovery of cohesin-independent TAD-like structures, also known as single-cell domains - highly variant self-interacting chromatin domains with boundaries that occasionally overlap with TAD boundaries but tend to differ among single cells and among single chromosome copies. Several recent computational modeling studies suggest that single-cell variations of epigenetic profiles may underlie the formation of the single-cell domains. Results: Here we use chromatin tracing to visualize in female human cells the fine-scale chromatin folding of inactive and active X chromosomes, which are known to have distinct global epigenetic landscapes and distinct population-averaged TAD profiles, with inactive X chromosomes largely devoid of TADs and cohesin. We show that both inactive and active X chromosomes possess highly variant single-cell domains across the same genomic region despite the fact that only active X chromosomes show clear TAD structures at the population level. These X chromosome single-cell domains exist in distinct cell lines. Perturbations of major epigenetic components did not significantly affect the frequency or strength of the single-cell domains. Increased chromatin compaction of inactive X chromosomes occurs at a length scale above that of the single-cell domains. Conclusions: In sum, this study suggests that single-cell domains are genome architecture building blocks independent of variations in major epigenetic landscapes.

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Section: Resultsmentioning

confidence: 99%

TAD-like single-cell domain structures exist on both active and inactive X chromosomes and persist under epigenetic perturbations

Cheng

Liu

et al. 2021

Preprint

Self Cite

View full text Add to dashboard Cite

show abstract

“…This selected 840-kb region spans a TAD boundary, based on previously published IMR-90 Hi-C datasets [ 12 , 16 ]. We partitioned this region of interest into consecutive 30-kb segments, labeled each segment with 400 primary oligonucleotide probes, and imaged each segment following our previously established chromatin tracing protocol [ 30 , 41 ]. In brief, we first labeled and imaged the entire region with a library of dye-labeled primary oligonucleotide probes.…”

Section: Resultsmentioning

confidence: 99%

“…For epifluorescence illumination, we used a 750-nm laser (2RU-VFL-P-500-750-B1R, MPB Communications) to excite and image Alexa Fluor 750, a 647-nm laser (2RU-VFL-P-1000-647-B1R, MPB Communications) to excite and image Cy5 and Alexa Fluor 647 fluorophores, a 560-nm laser (2RU-VFL-P-1000-560-B1R, MPB Communications) to excite and image ATTO 565 and Alexa Fluor 568 fluorophores, and a 488-nm laser (2RU-VFL-P-500-488-B1R, MPB Communications) to excite and image the yellow-green fiducial beads for drift correction. The microscope body hosts a penta-band dichroic mirror (ZT405/488/561/647/752rpc-UF2, Chroma), a penta-band emission filter (Chroma, ZET405/488/561/647-656/752 m), a motorized x–y sample stage (SCAN IM 112×74, Marzhauser), a piezo z positioner (Mad City Labs, Nano-F100S), and a home-built focus-lock system as described previously [ 41 ]. For datasets collected with “Combined chromatin tracing primary hybridization with mH2A1 immunofluorescence staining”, we installed a Duel-View setup on the emission path to prevent the fluorescent bleed-through between the mH2A1 and chromatin imaging channels.…”

Section: Methodsmentioning

confidence: 99%

TAD-like single-cell domain structures exist on both active and inactive X chromosomes and persist under epigenetic perturbations

Cheng

Liu

et al. 2021

Genome Biol

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Background Topologically associating domains (TADs) are important building blocks of three-dimensional genome architectures. The formation of TADs has been shown to depend on cohesin in a loop-extrusion mechanism. Recently, advances in an image-based spatial genomics technique known as chromatin tracing lead to the discovery of cohesin-independent TAD-like structures, also known as single-cell domains, which are highly variant self-interacting chromatin domains with boundaries that occasionally overlap with TAD boundaries but tend to differ among single cells and among single chromosome copies. Recent computational modeling studies suggest that epigenetic interactions may underlie the formation of the single-cell domains. Results Here we use chromatin tracing to visualize in female human cells the fine-scale chromatin folding of inactive and active X chromosomes, which are known to have distinct global epigenetic landscapes and distinct population-averaged TAD profiles, with inactive X chromosomes largely devoid of TADs and cohesin. We show that both inactive and active X chromosomes possess highly variant single-cell domains across the same genomic region despite the fact that only active X chromosomes show clear TAD structures at the population level. These X chromosome single-cell domains exist in distinct cell lines. Perturbations of major epigenetic components and transcription mostly do not affect the frequency or strength of the single-cell domains. Increased chromatin compaction of inactive X chromosomes occurs at a length scale above that of the single-cell domains. Conclusions In sum, this study suggests that single-cell domains are genome architecture building blocks independent of the tested major epigenetic components.

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“…Together, these methods paved the way for a diversity of multimodal imaging setups in which highly multiplexed DNA FISH is combined with both RNA FISH and protein visualization using immunofluorescence (IF) to identify nuclear landmarks and histone modifications (Figure 2A). The first method to do this was the recent multiplexed imaging of nucleome architectures (MINA) [74,75], which combined multiscale chromatin tracing (50 DNA loci at 1 Mb and 19 DNA loci at 5 kb resolution) along chr 19 with RNA FISH against 137 targets (applying the same barcoding approach used in the MERFISH [58] method described in the following text) and IF for fibrillarin to detect associations with nucleoli in a single experiment. Moreover, MINA represents the first instance of chromatin tracing in a mammalian tissue, and its application to mouse fetal liver sections not only allowed the visualization of dozens of TADs and regulatory regions but also uncovered cell type-specific as well as cell type-independent principles of chromatin organization [74,75].…”

Section: Trends In Geneticsmentioning

confidence: 99%

The era of 3D and spatial genomics

2022

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Chromatin tracing and multiplexed imaging of nucleome architectures (MINA) and RNAs in single mammalian cells and tissue

Cited by 18 publications

References 52 publications

TAD-like single-cell domain structures exist on both active and inactive X chromosomes and persist under epigenetic perturbations

TAD-like single-cell domain structures exist on both active and inactive X chromosomes and persist under epigenetic perturbations

TAD-like single-cell domain structures exist on both active and inactive X chromosomes and persist under epigenetic perturbations

The era of 3D and spatial genomics

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