BacNHEJ

DNA double strand breaks (DSBs) are among the most deleterious DNA lesions for living cells. They are repaired either by Homologous Recombination (HR) or by Non-Homologous End Joining (NHEJ). In bacteria, the current NHEJ model involves two proteins: Ku, a DNA end-binding protein and LigD, a multidomain protein with polymerase, ligase and nuclease activities. However, several observations indicate that this model may be simplistic. In particular, many bacterial species encode several copies of Ku and LigD proteins, raising questions about their relative contributions to NHEJ repair: what is the composition/stoichiometry of the NHEJ machinery? Are there multiple NHEJ machineries in a same bacterial cell? One of our working hypotheses is that the variable composition and/or multiplicity of NHEJ machineries could provide bacterial cells the capacity to manage the high chemical diversity of DNA ends present at natural DSBs. The main objective of this project is to better understand NHEJ in bacteria, notably the complex NHEJ pathways. To this end, we will acquire and integrate knowledge on three bacterial models, from one with a simple NHEJ machinery (Bacillus subtilis) to models with more complex or even multiple machineries (Sinorhizobium meliloti and Streptomyces ambofaciens), using a multidisciplinary approach including genetics, interactomics, protein biochemistry and structural biology, with as specific aims: i) revisit the composition/stoichiometry of complex bacterial NHEJ machinerie(s); ii) understand the assembly of components on DNA, and the potential complementarity of individual components and/or full machineries; iii) establish the structure of minimal or complete NHEJ machineries in complex with DNA. We expect from this work to provide new insights into this generic DNA repair mechanism, as well as into other cellular processes it may be involved in or interacting with, including other DNA repair pathways.