Switching genes to silent mode near
            <scp>DNA</scp>
            double‐strand breaks

Polo, Sophie E.

doi:10.15252/embr.201744052

Cited by 11 publications

(7 citation statements)

References 12 publications

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“…We also predict the compacted chromatin border likely contributes to DDR-dependent transcriptional inhibition, which is predicted to halt passage of RNA polymerases through a DDR locus to enable efficient repair [14,20]. DDR-dependent transcriptional silencing occurs in cis and is dependent upon ATM and RNF8 (11,61). This is consistent with our observation that formation of the compact chromatin border surrounding the damage focus is ATM and RNF8-dependent.…”

Section: Discussionsupporting

confidence: 82%

Phasor histone FLIM-FRET microscopy quantifies spatiotemporal rearrangement of chromatin architecture during the DNA damage response

Lou

Scipioni

Wright

et al. 2018

Preprint

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To investigate how chromatin architecture is spatiotemporally organised at a double strand break (DSB) repair locus, we established a biophysical method to quantify chromatin compaction at the nucleosome level during the DNA damage response (DDR). The method is based on phasor image correlation spectroscopy (ICS) of histone FLIM-FRET microscopy data acquired in live cells co-expressing H2B-eGFP and H2B-mCherry. This multiplexed approach generates spatiotemporal maps of nuclear-wide chromatin compaction that when coupled with laser micro-irradiation induced DSBs, quantify the size, stability, and spacing between compact chromatin foci throughout the DDR. Using this technology, we identify that ATM and RNF8 regulate rapid chromatin decompaction at DSBs and formation of a compact chromatin ring surrounding the repair locus. This chromatin architecture serves to demarcate the repair locus from the surrounding nuclear environment and modulate 53BP1 mobility. SIGNIFICANCE STATEMENTChromatin dynamics play a central role in the DNA damage response (DDR). A long-standing obstacle in the DDR field was the lack of technology capable of visualising chromatin dynamics at double strand break (DSB) sites. Here we describe novel biophysical methods that quantify spatiotemporal chromatin compaction dynamics in living cells. Using these novel tools, we identify how chromatin architecture is reorganised at a DSB locus to enable repair factor access and demarcate the lesion from the surrounding nuclear environment. Further, we identify novel regulatory roles for key DDR enzymes in this process. Finally, we demonstrate method utility with physical, pharmacological and genetic manipulation of the chromatin environment, identifying method potential for use in future studies of chromatin biology.

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Section: Discussionsupporting

confidence: 82%

Phasor histone FLIM-FRET microscopy quantifies spatiotemporal rearrangement of chromatin architecture during the DNA damage response

Lou

Scipioni

Wright

et al. 2018

Preprint

View full text Add to dashboard Cite

show abstract

“…Conversely, the compacted chromatin border excludes 53BP1 retention and may also inhibit the spurious binding of other DNA-repair factors. We also predict that the compacted chromatin border likely contributes to DDR-dependent transcriptional inhibition, which is predicted to halt passage of RNA polymerases through a DDR locus to enable efficient repair (45). DDR-dependent transcriptional silencing occurs in cis and is dependent upon ATM and RNF8 (5).…”

Section: Discussionmentioning

confidence: 99%

Phasor histone FLIM-FRET microscopy quantifies spatiotemporal rearrangement of chromatin architecture during the DNA damage response

Lou

Scipioni

Wright

et al. 2019

Proc. Natl. Acad. Sci. U.S.A.

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To investigate how chromatin architecture is spatiotemporally organized at a double-strand break (DSB) repair locus, we established a biophysical method to quantify chromatin compaction at the nucleosome level during the DNA damage response (DDR). The method is based on phasor image-correlation spectroscopy of histone fluorescence lifetime imaging microscopy (FLIM)-Förster resonance energy transfer (FRET) microscopy data acquired in live cells coexpressing H2B-eGFP and H2B-mCherry. This multiplexed approach generates spatiotemporal maps of nuclear-wide chromatin compaction that, when coupled with laser microirradiation-induced DSBs, quantify the size, stability, and spacing between compact chromatin foci throughout the DDR. Using this technology, we identify that ataxia–telangiectasia mutated (ATM) and RNF8 regulate rapid chromatin decompaction at DSBs and formation of compact chromatin foci surrounding the repair locus. This chromatin architecture serves to demarcate the repair locus from the surrounding nuclear environment and modulate 53BP1 mobility.

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“…Consistently, animals lacking HP1 (Luijsterburg et al 2009) or Polycomb components (Hong et al 2008) are hypersensitive to genotoxic stress. Besides a potential role of these factors in the recruitment of the repair machinery, they are also thought to promote silencing around sites of damage to prevent conflicts or interference between the repair and transcription machineries (Ui et al 2015;Vissers et al 2012) reviewed in Polo (2017).…”

Section: The Two-faced Role Of Heterochromatin In Dna Damage and Repairmentioning

confidence: 99%

The Importance of Satellite Sequence Repression for Genome Stability

Zeller

Gasser

2017

Cold Spring Harb Symp Quant Biol

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Up to two-thirds of eukaryotic genomes consist of repetitive sequences, which include both transposable elements and tandemly arranged simple or satellite repeats. Whereas extensive progress has been made toward understanding the danger of and control over transposon expression, only recently has it been recognized that DNA damage can arise from satellite sequence transcription. Although the structural role of satellite repeats in kinetochore function and end protection has long been appreciated, it has now become clear that it is not only these functions that are compromised by elevated levels of transcription. RNA from simple repeat sequences can compromise replication fork stability and genome integrity, thus compromising germline viability. Here we summarize recent discoveries on how cells control the transcription of repeat sequence and the dangers that arise from their expression. We propose that the link between the DNA damage response and the transcriptional silencing machinery may help a cell or organism recognize foreign DNA insertions into an evolving genome.

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Switching genes to silent mode near DNA double‐strand breaks

Cited by 11 publications

References 12 publications

Phasor histone FLIM-FRET microscopy quantifies spatiotemporal rearrangement of chromatin architecture during the DNA damage response

Phasor histone FLIM-FRET microscopy quantifies spatiotemporal rearrangement of chromatin architecture during the DNA damage response

Phasor histone FLIM-FRET microscopy quantifies spatiotemporal rearrangement of chromatin architecture during the DNA damage response

The Importance of Satellite Sequence Repression for Genome Stability

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