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Abstract Chromatin organization over multiple length scales plays a critical role in the regulation of transcription. Deciphering the interplay of these processes requires high-resolution, three-dimensional, quantitative imaging of chromatin structure in vitro . Herein, we introduce ChromSTEM, a method that utilizes high-angle annular dark-field imaging and tomography in scanning transmission electron microscopy combined with DNA-specific staining for electron microscopy. We utilized ChromSTEM for an in-depth quantification of 3D chromatin conformation with high spatial resolution and contrast, allowing for characterization of higher-order chromatin structure almost down to the level of the DNA base pair. Employing mass scaling analysis on ChromSTEM mass density tomograms, we observed that chromatin forms spatially well-defined higher-order domains, around 80 nm in radius. Within domains, chromatin exhibits a polymeric fractal-like behavior and a radially decreasing mass-density from the center to the periphery. Unlike other nanoimaging and analysis techniques, we demonstrate that our unique combination of this high-resolution imaging technique with polymer physics-based analysis enables us to (i) investigate the chromatin conformation within packing domains and (ii) quantify statistical descriptors of chromatin structure that are relevant to transcription. We observe that packing domains have heterogeneous morphological properties even within the same cell line, underlying the potential role of statistical chromatin packing in regulating gene expression within eukaryotic nuclei.
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Physical and data structure of 3D genome
With the textbook view of chromatin folding based on the 30-nm fiber being challenged, it has been proposed that interphase DNA has an irregular 10-nm nucleosome polymer structure whose folding philosophy is unknown. Nevertheless, experimental advances suggest that this irregular packing is associated with many nontrivial physical properties that are puzzling from a polymer physics point of view. Here, we show that the reconciliation of these exotic properties necessitates modularizing three-dimensional genome into tree data structures on top of, and in striking contrast to, the linear topology of DNA double helix. These functional modules need to be connected and isolated by an open backbone that results in porous and heterogeneous packing in a quasi–self-similar manner, as revealed by our electron and optical imaging. Our multiscale theoretical and experimental results suggest the existence of higher-order universal folding principles for a disordered chromatin fiber to avoid entanglement and fulfill its biological functions.
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- PAR ID:
- 10161019
- Date Published:
- Journal Name:
- Science Advances
- Volume:
- 6
- Issue:
- 2
- ISSN:
- 2375-2548
- Page Range / eLocation ID:
- eaay4055
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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- Free Publicly Accessible Full Text
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Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.1126/sciadv.aay4055
