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Cell signaling associated with vacuolar rupture and autophagy (F Lafont-CIIL CNRS UMR8204-INSERM U1019-Insitut Pasteur de Lille-Univ. Lille Nord de France)

This project is funded by the French Research National Agency (ANR). It is a consortium between 4 partners : F Lafont (coordinator), J Enninga, P Sansonetti and P Cossart (Institut Pasteur, Paris). Partners have identified some of the molecular targets for these modifications and have uncovered the presence of a variety of signaling molecules on membrane remnants of intracytoplasmic replicating bacteria. Moreover, Partners have established that these membranes are targeted to autophagy. The project involves first, high-content/high-throughput microscopy screens (RNAi approach) developed on recent findings (identification of cellular markers and pathways) and tools (specific probes). Second, atomic force microscopy-guided biophotonic measurements at high-resolution will be used to follow the dynamic orchestration of the involvement of the molecules identified with proteomics and functional microscopic assays. Also involving collaborations with chemists and biocomputing experts, this project will unveil mechanisms linking bacteria adhesion and membrane vacuolar rupture to induction of signaling pathways involved in the regulation of inflammatory and immune responses, and cell death.

Modelling of Bacillus spores and Listeria biofilms/material interaction dynamics (F Lafont)

This project is conducted as part of a project funded by the French Research National Agency and grant from the EU and Région Nord-pad-de-Calais. Importantly, this project is also financed by the industrial clusters MAUD and Aquimer. It is a scientific consortium between several partners : C Faille (INRA UR638 Lille, coordinator), and F Lafont, M Mercier-Bonin (INRA LISBP Toulouse)-P Schmitz (INSA Toulouse), Y Guerardel (CNRS UGSF Univ. Lille1-ULNF), C Slomianny (Inserm U1003, Univ. Lille 1-ULNF), G Bourdin (ANSES Boulogne s/mer). Mastering pathogen contamination of food industry equipment surfaces, which is necessary to avoid food contamination risk, requires an understanding of bacteria/material interaction dynamics. To fill the lack of tools to quantify interaction forces and knowledge of bacteriological complexity (physico-chemistry, deformability and morphology) we will use HCS to screen strain with different adhesion features and those selected for their remarkable properties will be analyzed in AFM. Of note, substrata will be similar to those used in food industry, i.e. steel plates. The results will then be applied to L. monocytogenes, a non-sporulated germ and L monocytogenes biofilms that represent a major contamination in fish industry.

(P Brodin-CIIL CNRS UMR8204-INSERM U1019-Insitut Pasteur de Lille-Univ. Lille Nord de France)

(A Baulard-C Locht-CIIL CNRS UMR8204- INSERM U1019-Insitut Pasteur de Lille-Univ. Lille Nord de France)

(C Grangette-CIIL CNRS UMR8204-INSERM U1019-Inst Pasteur de Lille-Univ. Lille Nord de France)

Inflammasome-targeting drug discovery (M. Chamaillard-CIIL CNRS UMR8204-INSERM U1019-Insitut Pasteur de Lille-Univ. Lille Nord de France)

More efficient therapeutic approaches are urgently needed in Crohn’s disease and colorectal cancer. This team wants to identify novel compounds that may have anti-inflammatory and/or anti-tumoral properties that are controlled by certain Nods-like receptors. A screening for modulators will be performed through HCS analysis by using a large library of chemicals in collaboration with B. Déprez, P. Brodin and F. Lafont. Any lead compounds will be further used to screen for potential endogenous regulators by using commercially available panels of siRNAs. After functional biochemical studies, the selected modulators will be tested in experimental mouse models of tissue injury and colitis-associated colorectal cancer and on cells isolated from CD patients.

(N. Prevarskaya- Inserm U1003- Université Lille 1-Univ. Lill Nord de France)

Nucleus-associated or cytoplasmic NMD inhibitors (F Lejeune-INSERM Avenir TEAM-IFR 142-Univ. Lille Nord de France)

Non-sense mediated mRNA decay (NMD) is a eukaryotic mRNA quality control that degrades mRNA harboring a premature termination codon (PTC). NMD can occur either in the nuclear fraction or in the cytoplasmic fraction. According to the identity of the PTC- containing mRNA, it will be degraded by nucleus-associated or cytoplasmic NMD. Among the 33% of inherited genetic diseases that are caused by mutations leading to the apparition of a premature termination codon (PTC), a small fraction could be rescued by blocking NMD. It is the case for example of nonsense mutations located in the exons 70 to 78 of dystrophin gene that lead to Duchenne muscular Dystrophy. The proposed project aims to identify specific NMD inhibitors capable to block exclusively nucleus-associated or cytoplasmic NMD by screening small chemical libraries using cell lines that express a PTC-containing mRNA subject to nucleus-associated NMD (Globin mRNA ; Thermann et al., 1998 ; Zhang et al., 1998) or cytoplasmic NMD (GPx1 mRNA ; Moriaty et al., 1998) and carrying a YFP-tag. PTC-containing mRNAs translation will lead to the absence of YFP expression. In the case, a molecule inhibits either nucleus-associated and/or cytoplasmic NMD, the PTC-containing mRNA will be stabilized and translated into a YFP-protein. HCS will be used to measure the YFP expression and determine where the protein is expressed as some inhibitors of NMD concentrate PTC-containing in cytoplasmic foci called P-bodies (Durand et al., 2007). We would like to know whether translation of mRNAs escaping from nucleus-associated NMD are translated at the same cellular region as PTC-containing mRNAs escaping from cytoplasmic NMD.

(F Foulquier-JC Michalski- CNRS UMR8576-Univ. Lille 1-Univ Lille Nord de France)

(B Staels-INSERM U545- Institut Pasteur de Lille-Univ. Lille 2-Univ Lille Nord de France)

(B Deprez-Inserm U 761– Institut Pasteur de Lille-Univ. Lille2-Univ. lille Nord de France)

(L Héliot-CNRS USR 3078-Univ. Lille 1-Univ Lille Nord de France)