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Abstract

The choroid plexus (ChP) is a specialized secretory tissue located within all brain ventricles, playing diverse crucial roles in brain development, homeostasis, and intercommunication between the brain and the body. Despite growing research interest, the ChP remains underexplored, hindering advancements in understanding and treating associated disorders. One of the pathological conditions of the ChP is choroid plexus tumors (CPT), which are rare tumors that tend to occur with increased frequency and malignancy in young patients. The research and treatment of CPT are challenged by the absence of valid experimental models, insights into the disease mechanism, and consequently, candidates for targeted therapy. Limited literature suggests a potential link between CPT and ChP cilia, but little is known regarding the formation and maintenance of these cilia during normal development, impeding efforts to establish a correlation between cilia and CPT. In my PhD research, I led and conducted two main projects: (i) investigating the tumorigenesis and in vitro modeling of CPT, and (ii) characterizing the biogenesis and maintenance of ChP cilia, aiming to fill crucial gaps in our understanding of these processes. To investigate CPT tumorigenesis, I first analyzed genomic and transcriptomic data from CPT patients and discovered that large-scale chromosomal instability events of the CPT genomes cause a constitutive activation of Wnt/β-catenin signaling in human CPTs. Next, I inhibited Wnt/β-catenin signaling pharmacologically and showed that CPT-derived cells depend on autocrine Wnt/β-catenin for survival. Additionally, constitutive Wnt/β-catenin pathway activation, either through knock-out of the negative regulator APC or overexpression of the ligand WNT3A, induced tumorigenic properties in 2D in vitro models of choroid plexus cells. Systematic hyper-activation of Wnt/β-catenin pathway in ChP organoids led to reduced differentiation of choroid plexus epithelial cells, rendering them increasingly susceptible to tumor development. Remarkably, CRISPR-Cas9 knock-out of APC in choroid plexus organoids was sufficient to induce oncogenic transformation. In summary, this project identifies Wnt/β-catenin signaling as a critical driver of CPT oncogenesis and provides the first 3D in vitro model for future pathological and therapeutic studies of CPT. To study the ChP cilia, I used a combination of super-resolution-microscopy approaches and mouse genetics, tracking cilia progression from embryogenesis when the ChP first emerges until senescence. I was able to provide fundamental understanding on ChP ciliogenesis, showing that their cilia are built on a gradient of events which are spatio-temporally regulated. Unexpectedly, I found that adult ChP cilia undergo axoneme resorption through a tubulin destabilization process, which is partially controlled by the microtubule-severing enzyme spastin and could be mitigated by polyglutamylation levels.