Biosynthesis and functional evaluation of novel antimicrobial peptides from mammalian gut microbiomes

The WHO has identified antibiotic resistance as one of the major threats to human health. According to predictions, the number of deaths related to antibiotic resistance is estimated at 10 million a year by 2050. This situation is prompting scientists to find new molecules, ideally natural ones, whose structures and mode of action differ from those of conventional antibiotics, to overcome the phenomena of resistance. One promising alternative concerns antimicrobial peptides of the RiPPs family (ribosomally synthesized and post-translationally modified peptides) produced by bacteria. Numerous studies show that the intestinal microbiome plays a very important role in the health of the host. Among the mechanisms involved, the production of antimicrobial peptides appears to be of particular importance. Part of our collaborative work aims to identify new antimicrobial peptides from complex biological ecosystems using metagenomic methods. To date, we have identified ten potentially interesting sequences. In this project, we will focus on the biosynthesis as well as the biochemical and structural characterizations of the antimicrobial peptides. An important part of the project will be devoted to the biological activity of these compounds on resistant and multi-resistant pathogens to conventional antibiotics. Moreover, the mode of action and the toxicity of the most effective peptides will be addressed.

Involvement of Rad51 paralogs in Rad51 filament formation in DNA repair

Homologous recombination (HR) is a major repair mechanism for DNA double-strand breaks induced by ionizing radiation. A key step in HR is the formation of Rad51 nucleoprotein filaments on the single-stranded DNA generated from these breaks. We have shown that strict control of these filaments is essential, so that HR does not itself induce chromosomal rearrangements (eLife 2018, Cells 2021). In humans, functional homologs of control proteins are tumor suppressors. Thus, the control of HR appears to be as important as the HR mechanism itself. Our project involves the use of new molecular tools enabling a real breakthrough in the study of these controls. We will be using a functional fluorescent version of the Rad51 protein developed for the first time by our collaborators A. Taddei (Institut Curie), R. Guérois and F. Ochsenbein (I2BC, Joliot, CEA). This major advance will enable us to observe the influence of control proteins on DNA repair by microscopy in living cells. We have also developed highly accurate structural models of control protein megacomplexes in association with Rad51 filaments. This study also led to the identification of specific domains for each paralog protein, outside the structurally conserved Rad51-like core, that might define the specificity of each paralog proteins. We will use a multidisciplinary approach based on genetic, molecular biology, biochemistry, protein structure and live microscopy methods and yeast as model organism to study the consequences of the ablation of these specific domains. We will also search for proteins specifically binding these domains. Their identification would be crucial to understand the function of Rad51 paralog complexes and help to develop new therapeutic approaches.

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