Pathological mechanisms underlying microcephaly Role of centrosomes and primary cilia in cerebral cortical development – CentrosomeCiliaMicroceph

Malformations of Cortical development (MCD) constitute a wide spectrum of brain disorders representing a major cause of intellectual deficiency and epilepsy. Among them, microcephaly (MIC) is characterized by prenatal reduced brain size occurring both isolated and in association with other MCD including gyration defects, which will be the focus of this proposal. Brain architecture is determined by various subtypes of neural stem and progenitor cells (NSPC) contributing differently in the regulation of thickness or folding of the cerebral cortex during development.
Centrosomes have been largely involved in the pathological mechanisms underlying MIC with almost 28 genes identified so far. They function as microtubule-organizing centers and are formed by a pair of centrioles that are microtubule-based structures. In cycling cells, centrosomes coordinate spindle pole formation, while in quiescent or interphase (G1) cells, they migrate to the cell surface where the mother centriole forms a basal body that nucleates a primary cilia. Stringent regulations govern switching between those dual roles of centrosome during cell cycle. Consequences of centrosomal proteins dysfunction on mitotic spindle formation have long been regarded as an essential mechanism underlying MIC. However, consequences of centrosomal dysfunction on cilia biogenesis and function have only recently emerged. Primary cilia are present on all NSPC types and have been shown crucial for both NSPC expansion and fate determination; and cilia defects have been reported in MIC patients harboring mutations in a growing number of MIC genes. In addition, several genes encoding centromal proteins have been found causal in ciliopathy diseases. This dual function of centrosome in forming both cilia and mitotic spindle ask the question of how disruption of such functionally linked proteins can lead to so diverse cortical phenotypes ranging from IRM undetectable and thus understudied in ciliopathies to isolated or gyration defects associated MIC.
Although human genetics has identified mutations in several genes encoding centrosomal proteins as responsible for MIC and ciliopathies, unanswered questions still remain: 1/ Many cases remain without any molecular basis implying the need for further MIC causal genes investigations. Preliminary exome analyses, performed in a unique cohort of both fetal and postnatal MIC cases with or without any gyration defects, have shed light on two novel candidate genes, LRRTM4 and MTOR. Their functions in cerebral cortical development will be dissected thinly to identify the mechanisms responsible for NSPC pool reduction. 2/ In view of the emerging roles of primary cilia in cortical development, we propose to dissect mechanisms associated to specific centrosomal gene disruption leading to distinct cortical malformations: ASPM in which mutations lead to isolated MIC, WDR62 linked to MIC with gyration defects both being major MIC genes and CEP290 involved in ciliopathy phenotypes. Focusing on the consequences of centriolar defects on both mitotic spindle formation and cilia dynamics and function in specific NSPC subtypes, should allow us to give new insight into the cellular mechanisms underlying different cortical phenotypes.
In addition to immunohistological analysis in human cerebral tissues, we propose to generate highly innovative and relevant models by deriving 2D and 3D cell based cortical models from patient fibroblasts derived induced pluripotent stem cells. Those complementary approaches should allow us to dissect the pathophysiological mechanisms underlying cortical size and folding and to enhance our understanding on the role of primary cilia during both pathological and normal cerebral cortical development.