Electron microscopy and atomic force microscopy studies of chromatin and metaphase chromosome structure

Daban, Joan‐Ramon

doi:10.1016/j.micron.2011.05.002

Cited by 59 publications

(62 citation statements)

References 97 publications

(176 reference statements)

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“…The present results provide evidence that relevant proportions of the DNA fibres in COPD NETs are comprised of nucleosome-bearing single double-helix molecules in ‘beads-on-a-string’ conformation (Fig. 6G,H), as known from previous experimental TEM analyses of chromosomal DNA [34], [35] and recent atomic force microscopy based models of chromatin structure [40]. ‘Beads-on-a-string’ DNA has as yet been only shown in the dilated perinuclear spaces and cytoplasmic vesicles of human neutrophils during a rapid form of NETosis in response to in vitro induction by Staphylococcus aureus [32].…”

Section: Discussionsupporting

confidence: 77%

New Aspects on the Structure of Neutrophil Extracellular Traps from Chronic Obstructive Pulmonary Disease and In Vitro Generation

et al. 2014

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Polymorphonuclear neutrophils have in recent years attracted new attention due to their ability to release neutrophil extracellular traps (NETs). These web-like extracellular structures deriving from nuclear chromatin have been depicted in ambiguous roles between antimicrobial defence and host tissue damage. NETs consist of DNA strands of varying thickness and are decorated with microbicidal and cytotoxic proteins. Their principal structure has in recent years been characterised at molecular and ultrastructural levels but many features that are of direct relevance to cytotoxicity are still incompletely understood. These include the extent of chromatin decondensation during NET formation and the relative amounts and spatial distribution of the microbicidal components within the NET. In the present work, we analyse the structure of NETs found in induced sputum of patients with acutely exacerbated chronic obstructive pulmonary disease (COPD) using confocal laser microscopy and electron microscopy. In vitro induced NETs from human neutrophils serve for purposes of comparison and extended analysis of NET structure. Results demonstrate that COPD sputa are characterised by the pronounced presence of NETs and NETotic neutrophils. We provide new evidence that chromatin decondensation during NETosis is most extensive and generates substantial amounts of double-helix DNA in ‘beads-on-a-string’ conformation. New information is also presented on the abundance and location of neutrophil elastase (NE) and citrullinated histone H3 (citH3). NE occurs in high densities in nearly all non-fibrous constituents of the NETs while citH3 is much less abundant. We conclude from the results that (i) NETosis is an integral part of COPD pathology; this is relevant to all future research on the etiology and therapy of the disease; and that (ii) release of ‘beads-on-a-string’ DNA studded with non-citrullinated histones is a common feature of in vivo NETosis; this is of relevance to both the antimicrobial and the cytotoxic effects of NETs.

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Section: Discussionsupporting

confidence: 77%

New Aspects on the Structure of Neutrophil Extracellular Traps from Chronic Obstructive Pulmonary Disease and In Vitro Generation

et al. 2014

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“…1 nN) is high enough to produce significant deformations of the chromatids. As expected for structures with components having large charges on their surfaces [74][75][76], divalent cations cause chromatin [2,22,27] and chromosome [67,73] condensation, but it has been observed [79] that crowding agents such as polyethylene glycol can also compact chromosomes even when the cation concentrations are very low. This suggests that the crowded environment of the cell can generate entropic forces [80] which may also contribute to the recovery of the initial size of deformed chromatids.…”

Section: Discussionmentioning

confidence: 88%

“…Chromosome dimensions and structural models for metaphase chromatin folding. (a) Schematic illustration of different models for chromatin structure in metaphase chromosomes: (a) chromatin fibres (grey lines) form loops that are bound to a central protein scaffold (brown) [8][9][10], (b) chromatin fibres are irregularly folded [11][12][13], and (c) chromatin has a laminar structure with many staked layers oriented perpendicular to the chromatid axis [2,[14][15][16][17][18]; parallel lines represent the side view of the stacked layers. (b) Examples showing the differences in metaphase chromosome sizes that exist among animal species: crested newt (T. cristatus) (a), human (b), and D. melanogaster (c); the three chromosome sets are represented at the same magnification; figure reproduced from [19] with permission from the Royal Society.…”

Section: Introductionmentioning

confidence: 99%

The energy components of stacked chromatin layers explain the morphology, dimensions and mechanical properties of metaphase chromosomes

Daban

2014

J. R. Soc. Interface.

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The measurement of the dimensions of metaphase chromosomes in different animal and plant karyotypes prepared in different laboratories indicates that chromatids have a great variety of sizes which are dependent on the amount of DNA that they contain. However, all chromatids are elongated cylinders that have relatively similar shape proportions (length to diameter ratio approx. 13). To explain this geometry, it is considered that chromosomes are self-organizing structures formed by stacked layers of planar chromatin and that the energy of nucleosome-nucleosome interactions between chromatin layers inside the chromatid is approximately 3.6 Â 10 220 J per nucleosome, which is the value reported by other authors for internucleosome interactions in chromatin fibres. Nucleosomes in the periphery of the chromatid are in contact with the medium; they cannot fully interact with bulk chromatin within layers and this generates a surface potential that destabilizes the structure. Chromatids are smooth cylinders because this morphology has a lower surface energy than structures having irregular surfaces. The elongated shape of chromatids can be explained if the destabilizing surface potential is higher in the telomeres (approx. 0.16 mJ m 22 ) than in the lateral surface (approx. 0.012 mJ m 22 ). The results obtained by other authors in experimental studies of chromosome mechanics have been used to test the proposed supramolecular structure. It is demonstrated quantitatively that internucleosome interactions between chromatin layers can justify the work required for elastic chromosome stretching (approx. 0.1 pJ for large chromosomes). The high amount of work (up to approx. 10 pJ) required for large chromosome extensions is probably absorbed by chromatin layers through a mechanism involving nucleosome unwrapping.

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“…The higher order structure of chromosomes has been widely studied by transmission and scanning electron microscopy, and recently by AFM (Daban, 2011;Ushiki et al, 2002;de Grooth and Putman, 1992;Summer, 1991;Earnshaw and Laemmli, 1983). The advantage of AFM is its ability to image at high resolution in a liquid environment, which is also expected by SICM.…”

Section: Chromosomesmentioning

confidence: 99%

Scanning ion conductance microscopy for imaging biological samples in liquid: A comparative study with atomic force microscopy and scanning electron microscopy

Ushiki

Nakajima

Choi³

et al. 2012

Micron

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Electron microscopy and atomic force microscopy studies of chromatin and metaphase chromosome structure

Cited by 59 publications

References 97 publications

New Aspects on the Structure of Neutrophil Extracellular Traps from Chronic Obstructive Pulmonary Disease and In Vitro Generation

New Aspects on the Structure of Neutrophil Extracellular Traps from Chronic Obstructive Pulmonary Disease and In Vitro Generation

The energy components of stacked chromatin layers explain the morphology, dimensions and mechanical properties of metaphase chromosomes

Scanning ion conductance microscopy for imaging biological samples in liquid: A comparative study with atomic force microscopy and scanning electron microscopy

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