Enforcement of cytoplasmic Notch pathway implication in epithelio-mesenchymal transition and cell differentiation in chicken embryos

• Main Author: Lebrun, Diane
• Other Authors: Institut NeuroMyoGène (INMG), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Université de Lyon, Christophe Marcelle
• Format: Doctoral or Postdoctoral Thesis
• Language: French
• Published: HAL CCSD 2018
• Subjects: Epithelio-mesenchymal transition (EMT); Electroporation/ immunostaining/ cryosection; Co-immunoprecipitation; Live imaging; Chicken embryo/ embryological development; Induction; Somite/ dorso-medial lip/ ventro-lateral lip; Neural tube/ neural crest; Transition épithélio-mésenchymateuse (EMT); Electroporation/ immunomarquage/ cryosection; Co-immunoprécipitation; Imagerie in vivo; Embryon de poulet; Développement embryonnaire; Somite/ lèvre dorso-médiale (DML); [SDV.BC]Life Sciences [q-bio]/Cellular Biology; DML
• Online Access: https://theses.hal.science/tel-01986211; https://theses.hal.science/tel-01986211/document; https://theses.hal.science/tel-01986211/file/TH2018LEBRUNDIANE.pdf

The epithelio-mesenchymal transition (EMT) is a well-known mechanism by which epithelial cells lose their adherent connections and gain migratory properties, associated with a gain of a mesenchymal phenotype. This EMT is required in numerous processes as gastrulation, organogenesis, fibrosis and cancers. Various molecular pathways orchestrate the EMT depending on the EMT biological context. Recently, our laboratory highlighted the implication of the cytoplasmic Notch pathway in the dorso-medial lip (DML) EMT. In the DML tissue, theEMT is synchronized with differentiation pathways, to generate cells forming the primary myotome. Our laboratory showed that neural crests cells expressing DLL1 activate NOTCH receptor of the DML cells, via a “kiss and run” model. This leads to NOTCH cleavage, releasing an activated intra-cytoplasmic NOTCH domain (NICD). In the cytoplasm, NICD inhibits the GSK3ß kinase, leading to the stabilization of SNAIL and the free cytoplasmic ßcatenin. These molecules translocate into the nucleus and lead to the activation of MRF as Myf5 (ß-catenin) and to the repression of adherent genes (SNAIL). Therefore, Notch cytoplasmic pathway allows a synergized induction of both, the EMT and myogenic programs. This pathway remains controversial and the precise mechanism how NICD inhibits GSK3ß needs to be elucidated. Therefore, the aim of my thesis project was to clarify how NICD inhibits GSK3ß activity. First, I confirmed that NICD and GSK3ß physically interact by CoIP. Moreover, I demonstrated that the serin-threonin kinase AKT, known to inhibit GSK3ß by phosphorylation and also to mediate EMT in cancer, can physically interact with NICD in the cytoplasm. I have also shown that AKT mediates the induction of the myogenic program through the inhibitory phosphorylation of GSK3ß and that SNAIL is downstream of AKT. Together, these experiments indicate that AKT mediates, through phosphorylation, the cytoplasmic NICD inhibition of GSK3ß leading to myogenesis. A comparison of the chicken NICD1 and the 4 ...