Single molecule microscopy reveals key physical features of repair foci in living cells

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Abstract

In response to double strand breaks (DSB), repair proteins accumulate at damaged sites, forming membrane-less sub-compartments or foci. Here we explored the physical nature of these foci, using single molecule microscopy in living cells. Rad52, the functional homolog of BRCA2 in yeast, accumulates at DSB sites and diffuses ~6 times faster within repair foci than the focus itself, exhibiting confined motion. The Rad52 confinement radius coincides with the focus size: foci resulting from 2 DSBs are twice larger in volume that the ones induced by a unique DSB and the Rad52 confinement radius scales accordingly. In contrast, molecules of the single strand binding protein Rfa1 follow anomalous diffusion similar to the focus itself or damaged chromatin. We conclude that while most Rfa1 molecules are bound to the ssDNA, Rad52 molecules are free to explore the entire focus reflecting the existence of a liquid droplet around damaged DNA.

Data availability

All data generated or analysed during this study are included in the manuscript and supporting files. Source data files are available on zenodo using the following link: https://zenodo.org/record/4495116.

The following data sets were generated

(2021) Single molecule microscopy reveals key physical features of repair foci in living cells
Zenodo, doi: 10.5281/zenodo.4495116.

https://zenodo.org/record/4495116

Article and author information

Author details

Judith Miné-Hattab

UMR 3664 - Nuclear Dynamics, Institut Curie, Paris, France

For correspondence
judith.Mine@curie.fr

Competing interests
No competing interests declared.

"This ORCID iD identifies the author of this article:" 0000-0001-9986-4092
Mathias Heltberg

UMR 3664 - Nuclear Dynamics, Institut Curie, paris, France

Competing interests
No competing interests declared.
Marie Villemeur

UMR3664 - Nuclear Dynamics, Institut Curie, Paris, France

Competing interests
No competing interests declared.
Chloé Guedj

UMR3664 - Nuclear Dynamics, Institut Curie, Paris, France

Competing interests
No competing interests declared.
Thierry Mora

Laboratoire de physique, Ecole Normale Supérieure, Paris, France

Competing interests
No competing interests declared.

"This ORCID iD identifies the author of this article:" 0000-0002-5456-9361
Aleksandra M Walczak

Laboratoire de Physique Theorique, École Normale Supérieure, Paris, France

Competing interests
Aleksandra M Walczak, Reviewing editor, eLife.

"This ORCID iD identifies the author of this article:" 0000-0002-2686-5702
Maxime Dahan

Division of Genetics, Genomics & Development, Department of Molecular and Cell Biology, Institut Curie, Paris, France

Competing interests
No competing interests declared.
Angela Taddei

UMR3664, Institut Curie, Paris, France

For correspondence
angela.taddei@curie.fr

Competing interests
No competing interests declared.

"This ORCID iD identifies the author of this article:" 0000-0002-3217-0739

Funding

Agence Nationale de la Recherche (ANR-11-LABEX-0044 DEEP)

Angela Taddei

Agence Nationale de la Recherche (ANR-10-IDEX-0001-02 PSL)

Angela Taddei

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Copyright

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.