The Dystrophin Associated Protein Complex (DAPC) is a key actor of the cell – extracellular matrix (ECM) interface, as revealed by its implication in human genetic disorders. However, its molecular and cellular functions are still poorly understood because tractable model systems allowing state-of-the-art in vivo cell biology and genetic approaches are lacking. The three partners of this project have developed the first transgenic dystrophin reporters that have revealed remarkably compartmentalized membrane distributions of the DAPC in epithelia and muscle cells in C. elegans, Drosophila, and mouse. The project aims to elucidate the organization and dynamics of the DAPC and to characterize its new functions in relation to specific cell cortical compartments.

Our project proposes to study the differentiation of enteric neurons, derived from the neural crest, whose roles are constantly expanding as our understanding of the brain-gut axis is improved. A large window of the ontogeny of enteric nervous circuits remains enigmatic, during which neurons organize themselves into ganglia and develop axonal projections whose precise and stereotyped orientation is the basis of the architecture of enteric circuits. Some childhood pathologies result in dysfunctions of gastrointestinal motility whose still unknown etiology could be rooted in early developmental alterations. A better understanding of the formation of enteric circuits could thus shed light on these pathologies. We will explore the mechanisms by which enteric neurons and their axons orient in the space of the gastrointestinal tract to construct the enteric circuits. We will take advantage of the chicken embryo model, which is particularly suitable for prenatal manipulations and the study of the neural crest. We have defined a paradigm to map the trajectories of different types of developing enteric axons using light sheet microscopy, and correlate them, by single nuclei RNA sequencing of equivalent tissue samples at the same stages, to the transcriptomic program of their neurons. We have extracted a first list of genes coding for guidance and cell adhesion ligands and receptors that we will study by loss of function. We will enrich these first data with new transcriptomic analyzes to cover a wider time window and capture the transcriptional dynamics that characterize the emergence of different neuronal subtypes and their own pattern of connections. Using a similar methodology, we will also study the development of the projection of the nodose ganglia, still little described and whose function is to relay information to the brain from specialized cells responsible for perceiving signals from the intestinal lumen. In a second axis of the project, we will focus on pathological contexts such as Hirschsprung's disease, characterized by impaired neural crest cell migration leading to distal aganglionosis. We will investigate whether neurons generated in regions that were populated by the neural crest are capable of forming intact enteric circuits. We have developed a model in the chick embryo to alter the expression of the RET receptor and mimic the effects of mutations associated with Hirschsprung's disease. Our first results attest to the effectiveness of the approach. This model will allow us to look for defects in the organization of enteric neurons and the establishment of their circuits in the areas colonized by the neural crest. Finally, we will also study samples from children with gastrointestinal dysfunctions to look for enteric circuit malformations using the data and methodologies developed with our model and the molecular data collected. Our project should thus provide new fundamental knowledge and open up new clinical perspectives.