Motile Cilia Formation in Brain Ventricles
Internship title: Motile Cilia Formation in Brain Ventricles
Affiliation: CNRS 8197
Address: 46 rue d’ulm 75005 Paris
Name: Pierre Paoeletti
Phone number: 0144323727
Name: Alice Meunier
Phone number: 0144323727
Subject Keywords: Cilia, Centrioles, brain, Chromosome Conformation, Phase separation
Tools and methodologies: Chromosome Capture, Single cell RNAseq, Cryo-electron tomography, Microrhéologie
Summary of lab’s interests: Motile cilia cover the brain ventricles, the respiratory and the reproductive tracts. They are necessary to propel the cerebrospinal fluid, to expel microorganisms/toxins and to direct the egg toward the uterus. Motile cilia are nucleated from centriolar structures at the apical surface of epithelial cells called multiciliated cells. The lab is interested in how this specialized cell type controls the production of up to 300 centriolar organelles necessary to build its ciliary tuft.
Project summary: Radial glial cells (RG) are neural stem cells located in the ventricular zone (VZ) from which glial and neuronal diversity arise during brain development1. By the end of the embryonic period, RG disappears while differentiating into several cell types, including adult stem cells and multiciliated ependymal cells. Ependymal cells line the subventricular zone (SVZ), the main neurogenic site located in the brain lateral ventricles. During their differentiation, they undergo massive subcellular changes driving the formation of up to 100 motile cilia. Cilium-driven cerebrospinal fluid circulation represents a cell-extrinsic route of regulating brain functions and ciliary dysfunction results in altered neurogenesis and neurodevelopmental defects. We have developed a RG cell culture assay to decipher the molecular and cellular processes underpinning motile ciliation with a focus on the amplification of centrioles, the structures nucleating motile cilia.
We have recently shown that multiciliated progenitors, while post-mitotic, hijack the robust mitotic program to orchestrate the massive production of centrioles. Here we propose a PhD project combining high-resolution imaging, chromosome capture experiments, biophysics and cell biology to further assess how complex events of centriole amplification/MCC differentiation are orchestrated in space and in time by the co-optation of the cell cycle machinery. Understanding how centrioles are amplified is particularly important in the context of cilia-related diseases as well as cancer biology. The student will benefit at ENS from an interdisciplinary environment specialized in cell and neurodevelopmental biology and will have access to state-of-the-art equipment and facilities. He will also benefit from national and international collaborations for chromosome capture and high resolution imaging experiments.
Interdisciplinary aspect of the project: The project relies on a combination of high-resolution imaging of MCC differentiation, HiC chromosome capture experiments, single cell RNA seq and biophysics (collaborations with Romain Koszul and Zoher Gueroui). The MCC culture assay developed for live cell observation of fluorescent proteins during centriole amplification will provide a robust basis to integrate multi-scale data both in the nucleus and in the cytoplasm. This culture system will provide with homogeneous cell populations undergoing differentiation to explore, using Hi-C and single cell RNAseq, changes in the higher order chromosome architecture / transcription during the progression of MCC differentiation. In parallel, it will allow assessing changes in the biophysical properties of the cytoplasm and featuring organelles (synthetic biology, microrheology, FRAP…) and to allow monitoring how centriole amplification is affected by modulation of these properties after siRNA depletions, drug treatments or production of tunable synthetic organelles.