Visualizing the genome in high resolution challenges our textbook understanding

Lakadamyali, Melike; Cosma, María Pía

doi:10.1038/s41592-020-0758-3

Cited by 77 publications

(72 citation statements)

References 89 publications

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“…While three-dimensional (3D) organization of the genome has been directly linked to gene regulation, information regarding nuclear-scale 3D chromatin organization under native physiological conditions is limited. Recent advances in imaging, sequencing and modeling approaches have greatly enhanced our understanding of 3D genome organization, bridging the gap between single-cell spatial information and genome-wide linear sequence interactions 1 . However, the absence of experiments that visualize the global 3D chromatin organization within the native tissue environment limits the interpretation of current data.…”

Section: Introductionmentioning

confidence: 99%

“…Nucleosomes are organized into 10-nm "beads on a string" fiber that together with additional proteins define the chromatin. While the textbook view suggests hierarchical higher-order folding of chromatin fibers (10 nm, 30 nm, 100 nm, and higher), recent advanced imaging under more physiological conditions challenged this view, describing a more complex and heterogeneous chromatin organization into clutches and domains of nucleosomes of varying sizes and densities [1][2][3][4] . On the nuclear scale, a range of imaging approaches, as well as Hi-C analysis, suggest that chromatin of an interphase nucleus is partitioned into distinct chromosomal territories several µm long, as was documented in various mammalian and Drosophila cells [5][6][7][8] .…”

Section: Introductionmentioning

confidence: 99%

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Live imaging of chromatin distribution in muscle nuclei reveals novel principles of nuclear architecture and chromatin compartmentalization

Pavlov¹,

Lorber²,

Bajpai³

et al. 2020

Preprint

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Packaging of the chromatin within the nucleus serves as an important factor in the regulation of transcriptional output. However, information on chromatin architecture on nuclear scale in fully differentiated cells, under physiological conditions and in live organisms, is largely unavailable. Here, we imaged nuclei and chromatin in muscle fibers of live, intact Drosophila larvae. In contrast to the common view that chromatin is distributed throughout the nuclear volume, we show that the entire chromatin, including active and repressed regions, forms a peripheral layer underneath the nuclear lamina, leaving a chromatin-devoid compartment at the nucleus center. Importantly, visualization of nuclear compartmentalization required imaging of un-fixed nuclei embedded within their intrinsic tissue environment, with preserved nuclear volume. Upon fixation of similar muscle nuclei, we observed an average of three-fold reduction in nuclear volume caused by dehydration and evidenced by nuclear flattening. In these conditions, the peripheral chromatin layer was not detected anymore, demonstrating the importance of preserving native biophysical tissue environment. We further show that nuclear compartmentalization is sensitive to the levels of lamin C, since over-expression of lamin C-GFP in muscle nuclei resulted in detachment of the peripheral chromatin layer from the lamina and its collapse into the nuclear center. Computer simulations of chromatin distribution recapitulated the peripheral chromatin organization observed experimentally, when binding of lamina associated domains (LADs) was incorporated with chromatin selfattractive interactions. Reducing the number of LADs led to collapse of the chromatin, similarly to our observations following lamin C over-expression. Taken together, our findings reveal a novel mode of mesoscale organization of chromatin within the nucleus in a live organism, in which the chromatin forms a peripheral layer separated from the nuclear interior.This architecture may be essential for robust transcriptional regulation in fully differentiated cells.

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Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Live imaging of chromatin distribution in muscle nuclei reveals novel principles of nuclear architecture and chromatin compartmentalization

Pavlov¹,

Lorber²,

Bajpai³

et al. 2020

Preprint

View full text Add to dashboard Cite

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“…Super-resolution techniques and tools SRM comprises an array of techniques such as photoactivated localization microscopy (PALM), stochastic optical reconstruction microscopy (STORM) and stimulated emission depletion (STED) microscopy that overcome the diffraction barrier and can achieve a resolution <10 nm and thereby generate novel biological insights (64)(65)(66)(67).…”

Section: Section Ii: Imaging Methods For Visualizing Mtdna In Cellsmentioning

confidence: 99%

Visualizing, quantifying, and manipulating mitochondrial DNA in vivo

Prole

Chinnery

Jones

2020

Journal of Biological Chemistry

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Mitochondrial DNA (mtDNA) encodes proteins and RNAs that support the functions of mitochondria and thereby numerous physiological processes. Mutations of mtDNA can cause mitochondrial diseases and are implicated in ageing. The mtDNA within cells is organized into nucleoids within the mitochondrial matrix, but how mtDNA nucleoids are formed and regulated within cells remains incompletely resolved. Visualization of mtDNA within cells is a powerful means by which mechanistic insight can be gained. Manipulation of the amount, and sequence of, mtDNA within cells is important experimentally and for developing therapeutic interventions to treat mitochondrial disease. This review details recent developments and opportunities for improvements in the experimental tools and techniques that can be used to visualize, quantify and manipulate the properties of mtDNA within cells.

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“…FISH methods (Giepmans et al, 2006). A variety of methods exist to recruit multiple fluorophores to targets to improve detection ( Figure 3B) (Lakadamyali and Cosma, 2020). In tiled dCas9-, TetR-TetO, and TALE-based systems (see Glossary), multiple proteins are recruited to adjacent locations on the genome (Chen et al, 2013;Miyanari et al, 2013;Straight et al, 1996;Tasan et al, 2018;Zhu and Cheng, 2020).…”

Section: Ll Open Accessmentioning

confidence: 99%

Understanding and Engineering Chromatin as a Dynamical System across Length and Timescales

Johnstone¹,

Wang²,

Sevier

et al. 2020

Cell Systems

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Connecting the molecular structure and function of chromatin across length and timescales remains a grand challenge to understanding and engineering cellular behaviors. Across five orders of magnitude, dynamic processes constantly reshape chromatin structures, driving spaciotemporal patterns of gene expression and cell fate. Through the interplay of structure and function, the genome operates as a highly dynamic feedback control system. Recent experimental techniques have provided increasingly detailed data that revise and augment the relatively static, hierarchical view of genomic architecture with an understanding of how dynamic processes drive organization. Here, we review how novel technologies from sequencing, imaging, and synthetic biology refine our understanding of chromatin structure and function and enable chromatin engineering. Finally, we discuss opportunities to use these tools to enhance understanding of the dynamic interrelationship of chromatin structure and function. INTRODUCTION TO CHROMATIN AND DYNAMIC MODES OF GENE REGULATION Chromatin can facilitate pattern formation and information transfer across multiple time and length scales (Figure 1A). Nanometer-scale interactions such as transcription factor binding occur on the order of milliseconds (Fierz and Poirier, 2019), while chromosome territories, the largest subnuclear structures, persist over many cell divisions (Dekker and Mirny, 2016). Because of the short-ranged stochastic nature of DNA-protein ll

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Visualizing the genome in high resolution challenges our textbook understanding

Cited by 77 publications

References 89 publications

Live imaging of chromatin distribution in muscle nuclei reveals novel principles of nuclear architecture and chromatin compartmentalization

Live imaging of chromatin distribution in muscle nuclei reveals novel principles of nuclear architecture and chromatin compartmentalization

Visualizing, quantifying, and manipulating mitochondrial DNA in vivo

Understanding and Engineering Chromatin as a Dynamical System across Length and Timescales

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