Team Leader
Amandine Pelletier
Engineer
Julie Robbe
Doctoral student
Shingo Fujii
Researcher
Titouan Feunteun
Master 2 student
Valérie Corinne Garcia
Researcher
Our NHEJ homologous recombination and genomic integrity team is committed to studying DNA double-strand break repair mechanisms at the molecular level, in particular homologous recombination and the non-homologous end joining (NHEJ) pathway, using a combination of biochemical and cell biology approaches.
A defect in the detection and repair of DNA double-strand breaks can be at the origin of cancer formation. To better understand this process, our team is studying the molecular mechanisms of the two main repair pathways of these genotoxic lesions: Homologous Recombination and NHEJ (Non-Homologous End Joining).
On the other hand, these repair systems enable cancer cells to resist anti-cancer treatments based on radiotherapy or chemotherapy using substances such as mitomycin C or cisplatin. Indeed, the principle of these treatments is to induce the formation of double-strand breaks in the DNA of cancer cells in order to kill them. Homologous recombination and NHEJ are therefore prime targets for devising ways of improving the efficacy of these anti-cancer treatments.
Our team is committed to studying the mechanisms of DNA double-strand break repair at the molecular level, in particular Homologous Recombination and the Non-Homologous End Joining (NHEJ) pathway, using a combination of biochemical and cell biology approaches.
We also use 'single molecule' methods to visualize and monitor the dynamic behavior of proteins involved in the repair process on isolated DNA molecules. For this, we use :
- Optical tweezers for attaching and manipulating isolated DNA molecules
- Fluorescence microscopy for real-time observation of tagged proteins interacting with DNA substrate
The projects
DNA is a molecule that is often damaged. If this damage is not repaired, or is badly repaired, mutations can appear, leading to a state of genomic instability. A cell in such a state may proliferate abnormally, giving rise to a tumor, or simply be doomed to die. Efficient repair of DNA damage is therefore a major necessity for living organisms.
Among these damages, DNA double-strand breaks are particularly dangerous, as they can induce chromosomal aberrations such as translocations or the loss of a chromosome fragment.
To counteract these effects, our cells have several repair pathways:
- Homologous recombination is based on a strand exchange between two identical or nearly identical DNA molecules, catalyzed by the RAD51 recombinase. This mechanism is essential for protecting genome integrity. However, incorrect use or disruption of this pathway can lead to the development of cancers, as in the case of mutations affecting BRCA2 in breast cancer.
- The non-homologous end joining (NHEJ) is based on specific proteins, such as DNA Ligase 4 and its cofactors XRCC4 and XLF. It plays a key role in the repair of double-strand breaks induced by ionizing radiation, protecting the genome against the formation of chromosomal aberrations.
Our research aims to understand precisely how these repair systems function at the molecular level. We study the activities of the proteins involved in vitro using biochemical approaches, but also within living cells. We also use a "single molecule" approach, enabling direct real-time visualization of repair proteins in action on a DNA molecule.
Ultimately, our research should lead to improved medical treatments for cancer and pathologies associated with DNA repair defects.
XLF, XRCC4 and DNA Ligase 4 (XXL) proteins of the non-homologous end joining (NHEJ) DNA double-strand break repair pathway interact in a dynamic multivalent disorder-based network, resulting in in vitro phase separation. These XXL condensates are powerful catalysts for the ligation of broken DNA ends and are capable of selectively recruiting helper proteins.
The ANR XXL project proposes to combine biochemical, biophysical, structural and genetic methods to understand how XXL condensates control NHEJ and its interactions with other repair pathways, in space and time in vitro and at DNA damage sites within cells.
PROJECT MANAGER
Valérie Corinne Garcia
DNA double-strand breaks (DSBs) are a major threat to genome stability, as they can lead to mutations, chromosomal rearrangements, and cancer. Interestingly, during meiosis, these potentially harmful lesions are intentionally introduced to trigger homologous recombination (HR), a repair process essential for accurate chromosome segregation and genetic diversity. Our past work has made significant contributions to the field, particularly by shedding light onto the function of the Mre11 nuclease in DSBs resection, an early step in meiotic HR (Garcia et al, Nature 2011), and by demonstrating the role of the checkpoint kinase Tel1/ATM in regulating meiotic DSB formation (Garcia et al, Nature 2015).
Recent discoveries
Meiotic DSBs are formed by Spo11 in certain areas of the genome called 'hotspots,' which colocalize with gene promoters. It was believed that only one DSB was formed within each hotspot, but our recent study demonstrated that multiple cuts can happen coincidently in these areas. These 'double cuts' create small gaps in the DNA, that are repaired through HR. Importantly, although our work mostly focused on the yeast model S.cerevisiae, our investigations in mouse have also revealed that DCs represent a general feature of meiotic recombination.
“Concerted cutting by Spo11 illuminates meiotic DNA break mechanics”. Johnson D, Crawford M, Cooper T, Claeys Bouuaert C, Keeney S, Llorente B, Garcia V*, Neale MJ*. Nature. 2021 *co-corresponding
To get mechanistic insights onto the role of Tel1 in DSB control, we investigated Tel1 dynamics onto meiotic chromosome and its genetics dependencies.
“Tel1 is recruited at chromosomal loop/axis contact sites to regulate meiotic DNA double-strand breaks interference” Dorme M, Aithal R, Cayrou C, Luciano P, Vernerey J, Borde V, Llorente B, Garcia V. PLoS Genetics 2025 *co-corresponding
Current investigations
In humans, ATM deficiency causes ataxia telangiectasia (A-T), a genetic disorder that is characterized by predisposition to cancer and sterility (amongst other phenotypes). Mutations that affect the kinase domain are particularly harmful in A-T patients and mouse models. However, the reason for the more severe effect of 'kinase dead' ATM mutations remains unknown. Unpublished work of ours demonstrates that inactivation of Tel1/ATM kinase activity (tel1kd) causes a severe deregulation of meiotic DSB formation in S. cerevisiae—something hardly detectable in tel1Δ or wild-type cells. We have identified new interactors of Tel1 involved in this process, and are currently investigating the underlying mechanisms.
