Local changes of higher-order chromatin structure during DSB-repair

Falk, Martin; Lukášová, Emı́lie; Gabrielová, Barbora; Ondřej, Vladan; Kozubek, Stanislav

doi:10.1088/1742-6596/101/1/012018

Cited by 14 publications

(9 citation statements)

References 41 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Microscopic analyses of γH2AX nuclear foci/clusters formed at DSB sites with the participation of a number of other repair proteins, as, for instance 53BP1, MRE11, pATM, Rad51, BRCA1, etc. (e.g, [ 53 , 104 , 105 ]), have contributed significantly to the current understanding of radiation-induced DNA damage, repair and misrepair processes (see, e.g., our earlier works [ 11 , 21 , 52 , 106 , 107 , 108 , 109 ]). In addition to the biochemical processes taking place at DSB sites, microscopy discovered important relationships between the damage-surrounding chromatin architecture, DSB generation and repair mechanisms, and mechanisms of chromosomal aberration formation following the exposure of cells to various types of ionizing radiation [ 110 ].…”

Section: Discussionmentioning

confidence: 99%

“…After irradiation, aliquots of the irradiated cell cultures were fixed at the given time points It is well-known that the architecture of chromatin is not random [83][84][85][86]. Importantly, structurally distinct chromatin domains with specific functions show different sensitivity to radiation damage [11,21,52,53,87,88]. Creating additional level of complexity with the potential influence on DSB damage and repair, specific chromatin domains such as euchromatin and heterochromatin are non-randomly distributed in the cell nucleus [84,89,90].…”

Section: Clustering and Persistent Homologies Of γH2ax Foci And Heterochromatinmentioning

confidence: 99%

“…In order to investigate the hypothesized mechanisms and to apply the basic findings to cancer research and diagnosis, a comprehensive toolbox is necessary to researchers that will allow to routinely provide an insight into spatial chromatin organization and determine cell type-specific or individual differences and potential treatment outcomes. Light and confocal microscopy—as unique methods in radiobiological research—have brought extensive knowledge of the organization of repair processes in space and time [ 11 , 51 , 52 , 53 ]. However, to obtain mechanistic insights into the true nature of spatial organization behind DNA damaging, repair, and misrepair, we must delve deeper into the level of interactions between individual molecules [ 54 , 55 , 56 , 57 , 58 , 59 , 60 , 61 ].…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Elucidation of the Clustered Nano-Architecture of Radiation-Induced DNA Damage Sites and Surrounding Chromatin in Cancer Cells: A Single Molecule Localization Microscopy Approach

Hausmann

Falk

Neitzel

et al. 2021

IJMS

View full text Add to dashboard Cite

In cancer therapy, the application of (fractionated) harsh radiation treatment is state of the art for many types of tumors. However, ionizing radiation is a “double-edged sword”—it can kill the tumor but can also promote the selection of radioresistant tumor cell clones or even initiate carcinogenesis in the normal irradiated tissue. Individualized radiotherapy would reduce these risks and boost the treatment, but its development requires a deep understanding of DNA damage and repair processes and the corresponding control mechanisms. DNA double strand breaks (DSBs) and their repair play a critical role in the cellular response to radiation. In previous years, it has become apparent that, beyond genetic and epigenetic determinants, the structural aspects of damaged chromatin (i.e., not only of DSBs themselves but also of the whole damage-surrounding chromatin domains) form another layer of complex DSB regulation. In the present article, we summarize the application of super-resolution single molecule localization microscopy (SMLM) for investigations of these structural aspects with emphasis on the relationship between the nano-architecture of radiation-induced repair foci (IRIFs), represented here by γH2AX foci, and their chromatin environment. Using irradiated HeLa cell cultures as an example, we show repair-dependent rearrangements of damaged chromatin and analyze the architecture of γH2AX repair clusters according to topological similarities. Although HeLa cells are known to have highly aberrant genomes, the topological similarity of γH2AX was high, indicating a functional, presumptively genome type-independent relevance of structural aspects in DSB repair. Remarkably, nano-scaled chromatin rearrangements during repair depended both on the chromatin domain type and the treatment. Based on these results, we demonstrate how the nano-architecture and topology of IRIFs and chromatin can be determined, point to the methodological relevance of SMLM, and discuss the consequences of the observed phenomena for the DSB repair network regulation or, for instance, radiation treatment outcomes.

show abstract

Section: Discussionmentioning

confidence: 99%

Section: Clustering and Persistent Homologies Of γH2ax Foci And Heterochromatinmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Elucidation of the Clustered Nano-Architecture of Radiation-Induced DNA Damage Sites and Surrounding Chromatin in Cancer Cells: A Single Molecule Localization Microscopy Approach

Hausmann

Falk

Neitzel

et al. 2021

IJMS

View full text Add to dashboard Cite

show abstract

“…Such studies may be relevant to the repair of DNA in genomic chromatin in view of the topological similarity of the minichromosome to chromatin loops and its position in regions of lower chromatin density within the nucleus [15], [17] where double strand breaks in genomic DNA and sites of their repair are predominantly localised [100], [101].…”

Section: Discussionmentioning

confidence: 99%

Repair of DNA Strand Breaks in a Minichromosome In Vivo: Kinetics, Modeling, and Effects of Inhibitors

et al. 2013

View full text Add to dashboard Cite

To obtain an overall picture of the repair of DNA single and double strand breaks in a defined region of chromatin in vivo, we studied their repair in a ∼170 kb circular minichromosome whose length and topology are analogous to those of the closed loops in genomic chromatin. The rate of repair of single strand breaks in cells irradiated with γ photons was quantitated by determining the sensitivity of the minichromosome DNA to nuclease S1, and that of double strand breaks by assaying the reformation of supercoiled DNA using pulsed field electrophoresis. Reformation of supercoiled DNA, which requires that all single strand breaks have been repaired, was not slowed detectably by the inhibitors of poly(ADP-ribose) polymerase-1 NU1025 or 1,5-IQD. Repair of double strand breaks was slowed by 20–30% when homologous recombination was supressed by KU55933, caffeine, or siRNA-mediated depletion of Rad51 but was completely arrested by the inhibitors of nonhomologous end-joining wortmannin or NU7441, responses interpreted as reflecting competition between these repair pathways similar to that seen in genomic DNA. The reformation of supercoiled DNA was unaffected when topoisomerases I or II, whose participation in repair of strand breaks has been controversial, were inhibited by the catalytic inhibitors ICRF-193 or F11782. Modeling of the kinetics of repair provided rate constants and showed that repair of single strand breaks in minichromosome DNA proceeded independently of repair of double strand breaks. The simplicity of quantitating strand breaks in this minichromosome provides a usefull system for testing the efficiency of new inhibitors of their repair, and since the sequence and structural features of its DNA and its transcription pattern have been studied extensively it offers a good model for examining other aspects of DNA breakage and repair.

show abstract

“…high-LET and low-LET) might differently interact with structurally and functionally distinct higher order chromatin domains (discussed in [ 1 ] and citations therein); this might be reflected by DNA double strand break (DSB) repair efficiency and the mechanism of how cancerogenous chromosomal translocations (CHT) form. Therefore, we compared the DSB repair kinetics and formation of γH2AX/p53BP1 repair clusters upon the action of γ-rays [ 2 , 3 ], protons (15 and 30 MeV) [ 4 ], and 20 Ne ions (preliminary data). Consequently, we discuss biological impacts of these clusters.…”

mentioning

confidence: 99%

Primary and secondary clustering of DSB repair foci and repair kinetics compared for -rays, protons of different energies and high-LET 20Ne ions

Falk

Lukášová

Falková

et al. 2014

Journal of Radiation Research

View full text Add to dashboard Cite

Purpose: Ionizing radiations of different qualities (e.g. high-LET and low-LET) might differently interact with structurally and functionally distinct higher order chromatin domains (discussed in [ 1] and citations therein); this might be reflected by DNA double strand break (DSB) repair efficiency and the mechanism of how cancerogenous chromosomal translocations (CHT) form. Therefore, we compared the DSB repair kinetics and formation of γH2AX/p53BP1 repair clusters upon the action of γ-rays [ 2, 3], protons (15 and 30 MeV) [ 4], and 20Ne ions (preliminary data). Consequently, we discuss biological impacts of these clusters.Material and methods: Immunostaining methods in combination with high-resolution confocal microscopy, performed on 3D-fixed normal human skin fibroblasts [ 2– 4], were used to study initial distributions of γH2AX and p53BP1 repair foci and their changes during the post-irradiation (PI) time, with a special concern on foci clustering. Irradiations with γ-rays, protons of different energies (15 and 30 MeV), and high-LET 20Ne ions was performed in IBP ASCR Brno (CR), NPI AVCR Řež (CR) and JINR Dubna (Russia), respectively.Results: Upon irradiating cells with 20Ne ions, tracks of multiple clustered γH2AX and p53BP1 repair foci appeared immediately after the irradiation; these clusters, called here as the ‘primary clusters’, were rare in cells irradiated with γ-rays or protons (submitted). Though γH2AX/p53BP1 foci were positionally quite stable [ 2], ‘secondary clusters’ occasionally appeared after all kinds of irradiation during about 30 min PI. The formation of secondary clusters usually appeared due to the heterochromatin decondensation at the sites of heterochromatic DNA double-strand breaks (hcDSBs), followed by their protrusion into a limited space of nuclear subdomains of low density-chromatin (discussed in [ 1, 2, 5]).Conclusions: Primary clusters appear in cell nuclei immediately PI as the consequence of highly localized energy deposition, while secondary clusters develop during (and because of) DSB repair. Primary DSB clusters probably represent the main cause of chromosomal translocations induced with high-LET radiations while secondary clusters seem to be more important for low-LET γ-rays and protons. Secondary clusters of primary clusters (higher-order clusters) observed for 20Ne ions might explain frequent formation of complex translocations upon the action of high-LET radiations. Finally, we suggest [ 1, 2, 4] a model that describes the relationship between the higher order chromatin structure, DSB formation, repair and misrepair.

show abstract

Local changes of higher-order chromatin structure during DSB-repair

Cited by 14 publications

References 41 publications

Elucidation of the Clustered Nano-Architecture of Radiation-Induced DNA Damage Sites and Surrounding Chromatin in Cancer Cells: A Single Molecule Localization Microscopy Approach

Elucidation of the Clustered Nano-Architecture of Radiation-Induced DNA Damage Sites and Surrounding Chromatin in Cancer Cells: A Single Molecule Localization Microscopy Approach

Repair of DNA Strand Breaks in a Minichromosome In Vivo: Kinetics, Modeling, and Effects of Inhibitors

Primary and secondary clustering of DSB repair foci and repair kinetics compared for -rays, protons of different energies and high-LET 20Ne ions

Contact Info

Product

Resources

About