Developmental mechanisms underlying human structural cerebellum defects – SCD-Mec

The goal of this project application is to investigate the key mechanisms of human structural cerebellar defects (SCD), utilizing the technical revolution in genomic sequencing and modeling. This program is designed to make a high impact on our understanding of the genetic and mechanistic basis of SCD. Disruption of the cerebellum development can cause structural cerebellar defects that are identified with MR imaging. SCD are generally associated with both motor coordination and cognitive deficits and can be separated into 2 main categories, degenerative and malformative patterns. The etiology remains unknown in more than 40% of the cases and therapeutic interventions are almost never possible.
Based on the recruitment at the Necker hospital, we previously built a cohort of undiagnosed patients with SCD. Using cutting-edge sequencing techniques, we propose to continue our effort to identify new disease genes using a new gene panel dedicated to SCD, whole exome and whole genome sequencing in order to improve etiological diagnosis. This application will then focus on dissecting the developmental processes and pathology mechanisms involved in a group of disorders thought to result from excessive cell death during fetal development or in the very first postnatal years. These disorders include some early-onset cerebellar atrophies, cerebellar hypoplasia and pontocerebellar hypoplasia. In order to have a broad impact, preliminary results are used to investigate new gene defects involving 3 biological processes affected in many other SCD, RNA processing defects (i), Calcium homeostasis (ii) and cell metabolic defects caused by disruption of protein N-glycosylation (iii). Additionally, we will investigate a completely new developmental mechanism disrupted in a new type of pontocerebellar hypoplasia.
The central models of this application are human Neural Stem Cells (NSCs) that will be CRISPR/Cas9-genome edited in order to reproduce the identified genetic defects. Proliferation, differentiation and cell survival of these models will be characterized. In parallel, in vivo modeling will be used to study the impact of patients’ mutations on zebrafish brain structure and function. Finally, we will use a recently developed mouse model for a SCD caused by a defect in protein N-glycosylation. Preliminary results obtained with this model show a defect in neuronal differentiation in the developing cerebellum. Using this mouse and a human NSC model, we will investigate cellular and proteomic consequences underlying the defect.
Correlations of in vivo and in vitro results obtained in this project will be used to understand cellular and molecular mechanisms involved in SCD and will set the stage for improved diagnosis and treatment.