A guide to visualizing the spatial epigenome with super‐resolution microscopy

Xu, Jianquan; Liu, Yang

doi:10.1111/febs.14938

Cited by 21 publications

(15 citation statements)

References 140 publications

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“…Simply put, Hi-C does not measure absolute contact frequency, and 3D reconstructions of higher order chromatin organization from Hi-C data have sometimes been used to argue for relatively open and accessible lariat-like chromatin loop structures [69, 73–76]. In contrast, recent studies using advanced microscopic strategies have suggested nucleosome clusters (NCs) as basic entities of chromatin organization beyond the nucleosome level [77–79]. Chromatin loops built from NCs imply more compacted and less accessible CDs with profound consequences for their accessibility for macromolecules (see below).…”

Section: Discussionmentioning

confidence: 99%

“…In order to test this hypothesis, it is necessary to explore the space-time compaction and accessibility of CDs. Super-resolved fluorescence microscopy, including single molecule localization microscopy (SMLM) and stochastic optical reconstruction microscopy (STORM), may become the methods of choice to measure absolute differences of DNA/chromatin compaction with spatial resolution at the nanometer scale [79, 84, 85], whereas chromatin accessibility can be probed indirectly with methods that allow to measure molecular diffusion rates [86–89].…”

Section: Discussionmentioning

confidence: 99%

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Cohesin depleted cells rebuild functional nuclear compartments after endomitosis

Cremer

Brandstetter

Maiser

et al. 2019

Preprint

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43The human genome forms thousands of "contact domains", which are intervals of enhanced 44 contact frequency. Some, called "loop domains" are thought to form by cohesin-mediated loop 45 extrusion. Others, called "compartmental domains", form due to the segregation of active and 46 inactive chromatin into A and B compartments. Recently, Hi-C studies revealed that the 47 depletion of cohesin leads to the disappearance of all loop domains within a few hours, but 48 strengthens compartment structure. Here, we combine live cell microscopy, super-resolution 49 microscopy, Hi-C, and studies of replication timing to examine the longer-term consequences 50 of cohesin degradation in HCT-116 human colorectal carcinoma cells, tracking cells for up to 51 30 hours. Surprisingly, cohesin depleted cells proceed through an aberrant mitosis, yielding a 52 single postmitotic cell with a multilobulated nucleus. Hi-C reveals the continued disappearance 53 of loop domains, whereas A and B compartments are maintained. In line with Hi-C, microscopic 54 observations demonstrate the reconstitution of chromosome territories and chromatin 55 domains. An interchromatin channel system (IC) expands between chromatin domain clusters 56 and carries splicing speckles. The IC is lined by active chromatin enriched for RNA Pol II and 57 depleted in H3K27me3. Moreover, the cells exhibit typical early-, mid-, and late-DNA 58 replication timing patterns. Our observations indicate that the functional nuclear 59 compartmentalization can be maintained in cohesin depleted pre-and postmitotic cells.60 However, we find that replication foci -sites of active DNA synthesis -become physically 61 larger consistent with a model where cohesin dependent loop extrusion tends to compact 62 intervals of replicating chromatin, whereas their genomic boundaries are associated with 63 compartmentalization, and do not change.64 65 3 Abbreviations 66 3D FISH = 3D fluorescence in situ hybridization 67 3D SIM = 3D structured illumination microscopy 68 AID = auxin inducible degron 69 ANC / INC = active / inactive nuclear compartment 70 CT = chromosome territory 71 CD(C) = chromatin domain (cluster) 72 CTCF = CCCTC binding factor 73 DAPI = 4',6-diamidino-2-phenylindole 74 EdU = 5-Ethynyl-2'-deoxyuridine 75 Hi-C = chromosome conformation capturing combined with deep sequencing 76 IC = interchromatin compartment 77 MLN = multilobulated nucleus 78 NC = nucleosome cluster 79 PBS = phosphate buffered saline 80 PBST = phosphate buffered saline with 0.02% Tween 81 PR = perichromatin region 82 RD = replication domain 83 RL = replication labeling 84 TAD = topologically associating domain 85 86 109 strengthened, leading to the presence of compartment domains and even compartment loops110 (but no loop domains) in the treated cells [18]. Other studies, using different cell types and 111 approaches for cohesin elimination yielded similar results [19-21], (reviewed in [22]). 112Here, we study the longer-term consequences of cohesin depletion and its effects on 113 the higher order nuclear a...

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Cohesin depleted cells rebuild functional nuclear compartments after endomitosis

Cremer

Brandstetter

Maiser

et al. 2019

Preprint

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“…Integrative biophysical and biochemical research strategies are imperative for a better understanding of the dynamic nuclear architecture and the molecular mechanisms involved in the interplay between nuclear architecture and function, for review, see ref. []. Such strategies will be discussed in a subsequent review with a focus on high‐resolution studies of chromatin compaction amd accessibility (Cremer et al., in preparation).…”

Section: Concluding Remarks and Future Perspectivesmentioning

confidence: 99%

The Interchromatin Compartment Participates in the Structural and Functional Organization of the Cell Nucleus

et al. 2020

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This article focuses on the role of the interchromatin compartment (IC) in shaping nuclear landscapes. The IC is connected with nuclear pore complexes (NPCs) and harbors splicing speckles and nuclear bodies. It is postulated that the IC provides routes for imported transcription factors to target sites, for export routes of mRNA as ribonucleoproteins toward NPCs, as well as for the intranuclear passage of regulatory RNAs from sites of transcription to remote functional sites (IC hypothesis). IC channels are lined by less-compacted euchromatin, called the perichromatin region (PR). The PR and IC together form the active nuclear compartment (ANC). The ANC is co-aligned with the inactive nuclear compartment (INC), comprising more compacted heterochromatin. It is postulated that the INC is accessible for individual transcription factors, but inaccessible for larger macromolecular aggregates (limited accessibility hypothesis). This functional nuclear organization depends on still unexplored movements of genes and regulatory sequences between the two compartments.

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“…The past decade not only witnessed the emerging application of super-resolution microscopy in centrosome research but also in the study of many other subcellular organelles and structures, including chromatin, which is mostly resolved by SMLM (Bintu et al 2018;Birk 2019;Sieben et al 2018b;Wang et al 2016;Xu and Liu 2019), the centromere and kinetochore (Joglekar et al 2009;Wan et al 2009;Wynne and Funabiki 2016), the contractile ring (Laplante et al 2016;McDonald et al 2017), the nuclear pore complex (Hurt and Beck 2015;Loschberger et al 2012;Szymborska et al 2013), and the mitochondria (Brown et al 2010;Jakobs and Wurm 2014;Jans et al 2013).…”

Section: Applications Beyond Centrosomementioning

confidence: 99%

Super-resolution microscopy: successful applications in centrosome study and beyond

Zhang

2019

Biophys Rep

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Centrosome is the main microtubule-organizing center in most animal cells. Its core structure, centriole, also assembles cilia and flagella that have important sensing and motility functions. Centrosome has long been recognized as a highly conserved organelle in eukaryotic species. Through electron microscopy, its ultrastructure was revealed to contain a beautiful nine-symmetrical core 60 years ago, yet its molecular basis has only been unraveled in the past two decades. The emergence of super-resolution microscopy allows us to explore the insides of a centrosome, which is smaller than the diffraction limit of light. Super-resolution microscopy also enables the compartmentation of centrosome proteins into different zones and the identification of their molecular interactions and functions. This paper compiles the centrosome architecture knowledge that has been revealed in recent years and highlights the power of several super-resolution techniques.

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A guide to visualizing the spatial epigenome with super‐resolution microscopy

Cited by 21 publications

References 140 publications

Cohesin depleted cells rebuild functional nuclear compartments after endomitosis

Cohesin depleted cells rebuild functional nuclear compartments after endomitosis

The Interchromatin Compartment Participates in the Structural and Functional Organization of the Cell Nucleus

Super-resolution microscopy: successful applications in centrosome study and beyond

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