Abstract

Understanding the basic cell biology of neural stem and progenitor cells is fundamental, on the one hand, to know how the large and complex brain of humans has evolved and in the other hand, for their successful application in regenerative medicine. While many central features are shared between different types of neural progenitors from diverse sources, there may be possibly an equal number of differences, and therefore it is of paramount importance to compare them. Towards that end, our laboratory identifies and investigates novel genes and molecular mechanisms regulating homeostasis and fate commitment of neural progenitors common to the developing and the adult brain, in mouse and recently, in humans. This line of experimentation has led to the discovery of new factors and biological phenomena essential for both embryonic and adult neurogenesis, and has been critical in pioneering and further developing direct neuronal reprogramming i.e. forced neurogenesis. My PhD work is the functional characterization of one such common candidate genes called Akna, and the investigation of its molecular regulation. I have discovered that this gene, erroneously annotated as an AT-hook transcription factor, is in fact an integral component of interphase centrosomes in the differentiating subtype of neural stem cells - radial glia - and in basal progenitors of the developing forebrain and in neuronal precursors of the adult brain. It localizes predominantly at subdistal appendages of mother centrioles where it regulates the organization and polymerization of microtubules. Gain- and loss-of-function experiments in the murine developing cerebral cortex show that Akna is necessary and sufficient for the delamination of differentiating neural stem cells in the (apical) ventricular zone towards the adjacent (basal) subventricular zone, where it is highest expressed. There, it is required for the retention of basal progenitors. Its subsequent downregulation allows repolarization and migration of young neurons to the cortical plate, where Akna is not detectable. Notably, cells that express Akna have mostly centrosome-based microtubule nucleation, while those without Akna, i.e. neurons, largely nucleate microtubules from noncentrosomal sources. This is indeed also the case for other cell types including adult brain neural precursors (neuroblasts) and immune cells; both of which have high levels of Akna. Furthermore, Akna’s enrichment in the outer subventricular zone of the folded ferret and macaque brains together with manipulation in human induced pluripotent stem cell-derived cerebral organoids suggest a conserved role in brain ontogeny and phylogeny. The delamination process of epithelial-like neural stem cells is reminiscent of the mesenchymal transition that can occur in true epithelial cells. In fact, many factors and molecular pathways are common to both processes, and so is Akna. We have found that it is upregulated early in mammary gland epithelial cells undergoing epithelial to mesenchymal transition. In its absence, disassembly of cell-cell junctions is impaired because degradation of epithelial adhesion molecules is delayed and hence, the resulting mesenchymal cell scattering is impaired. This, together with the above-mentioned results, supports a mechanistic model in which Akna’s role as microtubule organizer at centrosomes facilitates disassembly of cell-cell contacts and cell polarization in epithelial cells in general. Altogether, the results of my work uncover previously unconsidered, and therefore not observed, roles of centrosomal microtubule nucleation and highlights the key relevance of centrosome composition.