NEDD4L-mediated ubiquitination activity and AKT-mTOR pathways deregulation in malformations of cortical development – MCD-NEDD4L-mTOR

The development of the human cerebral cortex requires coordinated spatial and temporal regulation of interdependent developmental processes that include proliferation, migration and layering, as well as differentiation of distinct neuronal populations. Disruption of any of these processes can result in a wide range of developmental disorders, many of which are recognized as malformations in studies of brain imaging. Collectively these malformations correspond to a class of disorders designated as malformations of cortical development (MCD) that represent common causes of developmental delay and epilepsy.

During the past decade the classification of MCD and their genetic causes have advanced significantly. In practice clinical and genetic diagnosis of categories such as lissencephaly, pachygyria, polymicrogyria, focal cortical dysplasia (FCD), and periventricular nodular heterotopia (PNH) could nowadays be established for a substantial number of patients.
Within the heterogeneous group of MCD associated with heterotopic positioning of neuronal cells, PNH is a fairly common condition and easily recognizable by brain MRI, whose major causes are mutations of the X-linked gene Filamin A (FLNA) that account for about 50% of the cases.

In a recent collaborative study involving the four partners of this proposal, we showed that missense mutations in the HECT domain of the E3 ubiquitin ligase NEDD4L lead to a developmental disorder characterized by PNH. Cellular and expression data showed a sensitivity of PNH-associated mutants to proteasome degradation that contrasts with the stability of wild type (WT) NEDD4L. Moreover, in utero electroporation approach showed that PNH-related mutants and excess of WT NEDD4L affect neurogenesis, neuronal positioning and terminal translocation. Further investigations, revealed differential deregulations of pathways involved. Excess of WT NEDD4L leads to a disruption of Dab1 and mTORC1 pathways, while PNH-related mutations are associated with a deregulation of mTORC1, Akt and Smad2/3 activities in basal conditions and upon TGF-ß activation.

These initial findings provide an interesting entry point to investigate the role of NEDD4L-mediated ubiquitination in developmental processes. The overall objective of the present proposal is to further understand the role of NEDD4L in the regulation of AKT/mTOR pathways and its contribution to the regulation of neurodevelopmental processes underlying cortical development. Moreover, the proposal will provide insights into pathophysiological mechanisms underlying the large spectrum of neurodevelopmental syndromes, some of which are associated with phenotypic features of MCD, caused by mutations in AKT3, PIK3R2, mTOR and PIK3C leading mainly to an activation of PI3K-AKT-mTORC1 signalling pathway.

In this collaborative project that brings together laboratories with complementary skills, we propose functional studies that combine investigations of a unique conditional knock-in mouse model and iPSC derived from mutant fibroblasts, as well as targeted and large scale in vitro and in vivo studies to identify relevant NEDD4L substrates and interactors critical for the regulation the AKT/mTOR pathways during cortical developmental processes.

This proposal should bring the first cellular and molecular dissection of the pathophysiological consequences of NEDD4L mutations on AKT/mTOR pathways, as well as a better understanding of the role of NEDD4L mediated ubiquitination regulation of AKT/mTOR signalling pathways in normal and diseased brains.