Regulation of Neural Stem Cell Reactivation from Quiescence by Maternal Lipids and the Microtubule Binding Protein Toucan

Controlled proliferation of neural stem and progenitor cells (NSPCs) produces various types of neuron and glial cells which eventually form neural circuits and a functional brain. Both cell extrinsic and intrinsic factors are known to play key roles in controlling NSPCs proliferation throughout development. In this thesis, I will investigate how lipid droplets availability and lipid metabolism as cell extrinsic factors affect NSPCs proliferation in the Drosophila freshly hatched larval brain. Next, I will show how a microtubule binding protein affects asymmetric cell division in the mushroom body neuroblasts (MB NBs) during early stages of Drosophila larval brain development.

Here I show that lipid metabolism plays an important role during neuroblasts (NBs) reactivation from quiescence. Drosophila females deposit a large number of lipid droplets in the egg during oogenesis which become depleted at the end of the embryonic stage. Recent studies have shown that these LDs in the embryonic core are distributed to the peripheral tissues during the very early embryonic developmental stages and interference with this process leads to developmental delay or embryonic lethality. We have discovered that the maternally deposited lipids droplets are required for the timely reactivation of the Drosophila neural stem cells. Either knockdown of the Brummer lipase in the NBs or mutation in the genes that are involved in maternal lipid droplets homeostasis in the freshly hatched brain leads to a significant delay in the NBs reactivation from quiescence. We have also identified that the evolutionarily conserved growth signaling PI3K pathway is attenuated when the NBs either lack or fail to break down maternally deposited lipid droplets. Freshly hatched Drosophila larvae start feeding immediately once they hatch out of the embryo and have access to food. They can access lipids from the food and use carbohydrates to synthesize lipids through the de novo lipogenesis. We have determined that dietary lipids are not essential for the quiescent NBs timely reactivation. However, lipids produced via de novo lipogenesis plays a role in the quiescent NBs reactivation. Overall, our data indicate that lipid metabolism plays an important role in the quiescent NBs reactivation.

I also show that a microtubule associated protein (MAP), called Toucan (Toc), regulates NBs asymmetric cell division. The function of MAPs in cell division is evolutionarily conserved across a wide range of species. MAPs play critical roles in the organization, stabilization, and regulation of microtubules during cell division, ensuring proper chromosome alignment, segregation, and cytokinesis. Earlier studies showed that Toc is provided maternally in the Drosophila embryo and plays a critical role in nuclear division during the syncytial blastoderm stage. The maternal pool of Toc is depleted by the end of the embryonic stage. We have discovered that Toc is also expressed in the Drosophila larval brain, presumably another isoform of Toc as there are ten unique polypeptides of Toc in Drosophila. In Toc mutant larval brains, quiescent NBs fail to reactivate, and the proliferating MB NBs exit cell cycle by 24 hours after larval hatching (ALH). Further studies revealed that Toc mutant MB NBs had defects in their microtubule network and most of them underwent cytokinesis failure. We have also found that Toc mutant MB NBs have defects in asymmetric segregation of the basal polarity proteins. By 40 hours ALH, MB NBs were terminally differentiated due to nuclear accumulation of Prospero. Our findings strongly suggest that Toc is a microtubule associated protein required for the asymmetric cell division of the NBs in the Drosophila brain.