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    Genome stability and instability (SInG)

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    Team Leader:

     

    Genome stability maintenance is essential to prevent aging, cancer, neuronal and/or congenital disorders. The organisms are continually challenged by stresses (radiation, chemical contaminants, free radicals ...) that damaged the DNA and thus that generate genetic instability. Our laboratory is interested in the subtle balances of between the different mechanisms that orchestrate DNA repair, protecting against genetic instability while allowing genetic diversity (antibodies, gametes, molecular evolution).

    Objectives

    The integrity of the genetic material is daily compromised by exogenous (radiation exposure) or endogenous attacks. Endogenous stress can result from reactive oxygen species (ROS) generated by cell metabolism; in addition, replicative stresses also represent important endogenous stressors. Indeed, the progression of the replication forks is inevitably blocked by numerous endogenous structural damages or configurations (DNA / RNA hybrids, DNA structure difficult to replicate, strong protein binding, etc.). It can be noted that oxidative stress and replicative stress are generally considered both tumorigenic and promoting senescence that counteracts tumour development. The transmission of the genetic information therefore requires the control of the DNA damage response (DDR), which coordinates a network of metabolic pathways. It should be emphasized that spontaneous activation of DDR, in response to endogenous replicative stress, has been described in precancerous cells or at early stages of tumour initiation. In addition, DDR defects are also associated with developmental, neuronal or inflammation pathologies. Thus the study of the nature and consequences of endogenous replicative stress is of crucial importance for the understanding of spontaneous genetic instability.

    DNA Double-strand breaks (DSBs) are highly toxic lesions that can be produced by ionizing radiation or arrests of  the replication forks. DSBs can also be physiologically produced by the cell during meiotic differentiation or V (D) J recombination generating the immune repertoire. The study of CBD repair therefore has both fundamental and applied interests. Indeed, on the one hand, strategies using radiotherapy should largely benefit from knowledge on the regulation of DSB repair. On the other hand, if DSBs can be used to generate genetic diversity (meiosis, V (D) J recombination) they can also create genetic instability. Fine and precise regulation of DSB repair is therefore essential to preserve these balances and cellular issues.

    The metabolic network repairing DNA damages must therefore support the maintenance of genome stability, while allowing genetic diversity. For some lesions, several alternative mechanisms can act; it is therefore essential to coordinate the different repair systems between them, but also with other fundamental processes of the cell, such as DNA replication, the cell cycle and apoptosis.

     

    DSB repair uses two main general strategies:

    1. The first strategy uses one DNA partner sharing homologous and refers to homologous recombination (RH). This is a process, conserved in all organisms, and is at the heart of the balance maintaining the balance between stability and genetic diversity. In addition to DSBs repair, RH allows the restart of blocked replication forks, which gives it a protective role against endogenous replicative stress.
    2. The second DSB repair strategy ligates the double-stranded ends without requiring sequence homologies (NHEJ: Non-homologous End-joining).

     

    Our project is based on two main complementary axes.

    Axis 1: Origin of endogenous replicative stress, and consequences of RH defect on mitotic segregation. Given its role in restarting the blocked replication forks, the RH defect thus makes it possible to reveal the endogenous stress. This axis provides on the one hand to identify the mechanisms responsible for the slowing down of the replication forks in the rh- cells (thus the original endogenous stress), on the other hand to analyse the consequences on the G2 phase of the defect of RH, leading to chaotic chromosomal segregations in mitosis.

    In addition, we have developed a postnatal conditional inactivation of HR mouse model. The in vivo consequences of the invalidation of HR  will therefore be analysed.

    Axis 2: Regulation of NHEJ and of the choice of DSB Repair System. We have developed substrates to analyse end-joining in an intra-chromosomal context. The substrates we have developed allowed us to identify and characterize the alternative end-joining pathway (A-EJ), in an intrachromosomal context, in living cells. A-EJ is initiated by single-stranded DNA resection (like HR). The choice of the DSB repair system is therefore two levels: 1- NHEJ vs. Resection; 2-A-EJ vs. RH. Our project plans to analyse the impact of HR proteins, Brca2 and Rad51, in the choice A-EJ vs HR. We are analysing the mechanisms controlling the correct choice of DSB repair system. Finally, we analyse the impact of RAD51 and RAD52 on these different processes.

     

    Main publications

    Flavie Coquel, Maria-Joao Silva, Hervé Técher, Karina Zadorozhny, Sushma Sharma, Jadwiga Nieminuszczy, Clément Mettling, Elodie Dardillac, Antoine Barthe, Anne-Lyne Maurin, Alexy Promonet, Alexandra Cribier, Amélie Sarrazin, Wojciech Niedzwiedz, Bernard Lopez, Vincenzo Costanzo, Lumir Krejci, Andrei Chabes, Monsef Benkirane, Yea-Lih Lin and Philippe Pasero. SAMHD1 acts at stalled replication forks to prevent ssDNA-mediated induction of type I interferons (2018). Nature, 557, 57-61. doi: 10.1038/s41586-018-0050-1

    Camille Gelot, Josée Guirouilh-Barbat, Tangui Le Guen, Elodie Dardillac, Catherine Chailleux, Yvan Canitrot and Bernard S. Lopez (2016). The cohesin complex prevents the end-joining of distant DNA double-strand ends. Mol Cell, 61, 15-26.

    Josée Guirouilh-Barbat, Camille Gelot, Anyong Xie, Elodie Dardillac, Ralph Scully and Bernard S. Lopez (2016) 53BP1 Protects Against CtIP-dependent Capture of Ectopic Chromosomal Sequences at the Junction of Distant Double-Strand Breaks. PLoS Genet. DOI:10.1371/journal.pgen.1006230.

    Therese Wilhelm, Sandrine Ragu, Indiana Magdalou, Christelle Machon, Elodie Dardillac, Hervé Técher, Jérôme Guitton, Michelle Debatisse and Bernard S. Lopez (2016). Slow Replication Fork Velocity of Homologous Recombination-Defective Cells Results from Endogenous Oxidative Stress. PLoS Genet., 12(5): e1006007. doi:10.1371/journal.pgen.1006007

    Therese Wilhelm, Indiana Magdalou, Aurélia Barascu, Hervé Técher, Michelle Debatisse and Bernard S. Lopez (2014). Spontaneous slow replication fork progression elicits mitosis alterations in homologous recombination-deficient mammalian cells. Proc Natl. Acad. Sci. USA. 111.763-768.

     

    Team's news

    • Equipe labélisée par la Ligue Nationale contre le Cancer
    • Contract INCa, 2 contracts ANR
    • 2017: Annual National Prize of the Cuban Academy of Sciences. Category : Biomedical Sciences
    • 2010: Prix de la  Ligue Nationale Française contre le Cancer “