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Imaging nuclear organization and DNA repair at the single molecule level

Thursday, October 21, 2021 - 11:00
Judith Miné-Hattab
Participer à la réunion Zoom ID de réunion : 892 640 6558 Code secret : 7mdSKr
UMR3664 / Nuclear Dynamics, Institut Curie, PSL University, Sorbonne Université, CNRS

Our genome is constantly damaged by a variety of exogenous and
endogenous agents. Among the various forms of DNA damage, double-strand
breaks (DSBs) are the most cytotoxic and genotoxic for the cell.
Eukaryotic organisms use several mechanisms to repair DSBs among them
non-homologous end-joining (NHEJ) and homologous recombination (HR).

             Here, here, I will present our recent work to investigate
the molecular mechanisms of HR proteins inside cells at the single
molecule level. In response to DSB, repair proteins colocalize from a
diffuse distribution to repair foci located at the damaged DNA site. An
enduring question in the DNA damage field is how do repair proteins find
their correct target, accumulate within repair foci and disassemble with
the proper time-window? Despite their functional importance, the
physical nature of repair sub-compartments remains unclear. To answer
these questions, we use single particle tracking (SPT) and PALM (Photo
Activable Localization Microscopy) to assess the physical properties
underlying repair foci formation and the internal dynamics of these
membrane-less sub-compartments. Using this approach in Saccharomyces
cerevisiae yeast, we found that Rad52 share many properties
characterizing Liquid Liquid Phase Separation including: sharp change in
diffusion coefficient while entering or escaping foci, fusion of
multiple foci, existence of a potential attracting molecules to the
center of foci. Furthermore, I will present some recently developed
set-ups combining single molecule microscopy with laser-micro
irradiation or with micro-fluidic devices.

1.     Heltberg Mathias, Miné-Hattab Judith, Taddei Angela, Walczak
Aleksandra M., Mora Thierry. Physical observables to determine the
nature of membrane-less cellular sub-compartments. BioRxiv (2021).

2.     Judith Miné-Hattab*, Mathias Heltberg, Marie Villemeur, Chloé
Guedj, Aleksandra M. Walczak, Thierry Mora, Maxime Dahan, Angela
Taddei*, * corresponding authors. Single molecule microscopy reveals key
physical features of repair foci in living cells eLife, 10: e60577,

3.     Judith Miné-Hattab* & Irène Chiolo, * corresponding authors.
Complex Chromatin Motions for DNA Repair. Frontiers in Genetics, 11:
800, (2020).

4.     Camille Clément, Guillermo Orsi, Alberto Gatto, Ekaterina
Boyarchuk, Audrey Forest, Bassam Hajj, Judith Miné-Hattab, Mickaël
Garnier, Zachary Gurard-Levin, Jean-Pierre Quivy, and Geneviève
Almouzni. High-resolution visualization of H3 variants during
replication reveals their controlled recycling", Nature Communication,
9;9(1):3181, (2018).

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