UCSF

In the developing vertebrate brain, Radial Glia Progenitor (RGP), the principal neural stem cells, undergo symmetric and asymmetric cell division (ACD), giving rise to both RGPs (self-renewal) and differentiated cell types (e.g., neurons, oligodendrocytes, and astrocytes) that will ultimately form the central nervous system. Perturbations of RGPs divisions will result in neurodevelopmental disorders and brain tumors. Understanding the basic biology of RGPs divisions will potentially have a significant impact on understanding brain development and may provide insights into treatments for developmental disorders or brain tumors. During ACD, the mother cell must establish the proper axis of polarity with respect to asymmetrically localized cell fate determinants. These processes are regulated by the evolutionarily conserved Partition defective protein (Par) complexes. The Par complexes have been extensively characterized in invertebrates, but much remains to be understood in vertebrates, especially in the context of RGPs ACD. Previous studies in invertebrates and vertebrates have demonstrated that Par-3 is essential for regulating ACD by localizing asymmetrically along the division axis. Par-3 has long been thought to function exclusively at the cell cortex. Recent findings have led us to investigate the role of Par-3 during ACD in RGPs. Therefore, one approach to gain insights into this question is the utilization of zebrafish (Danio rerio), a powerful model organism used in biological research. Proteins of interest can be easily targeted by both pharmacological agents and genetic alterations within zebrafish, allowing for examination of complex phenotypes resulting from desired perturbations. Unlike mammalian models, development is relatively fast, and breeding produces a large amount of progeny, allowing for quick generation progression and large sample sizes. Also, unlike mammals, development occurs externally, and the brain is transparent, allowing for accessible and clear visualization of neurodevelopment, neuronal activity, and fluorescent markers of interest throughout the whole brain. These attributes make zebrafish an excellent model for studying the in vivo the process of RGPs undergoing ACD in the developing forebrain. In this dissertation, we explore how polarity plays a role during ACD in RGPs. In Chapter 1, we delve into Par-3's role in regulating polarity along the cell cortex in RGPs, influencing Notch activity in daughter cell nuclei. We explore techniques, such as the antibody uptake assay and in vivo time-lapse imaging in zebrafish, revealing the dynamics of internalized Notch ligand DeltaD during RGP ACD. Our findings uncover the role of cytoplasmic Par-3 in localizing intracellular determinants. Next, in Chapter 2, we delve deeper into the role of Par-3 during active neurogenesis, particularly in ACD in RGPs. Using various techniques including in vivo time-lapse imaging, biochemical assays, and pharmacological studies, we investigate the interplay between cortical and cytoplasmic Par-3. Our focus is on understanding how Aurora Kinase A (AurkA) phosphorylation of Par-3 influences ACD in RGPs. Our findings highlight AurkA's role in regulating Par-3 dynamics between the cytoplasm and cortex during mitotic RGPs, ultimately impacting neural progenitor fate. In chapter 3, we pivot away from lab bench research and discuss a diversity, equity, and inclusion (DEI) project, DE-SILO (Diversity, Equity, and Sociology training in Laboratory Organizations), that I worked on with Dr. Melaine Jeske. We designed a course module that was tailored for laboratory meetings at academic institutions. Our aim was to intersect DEI topics and facilitate discussions in the context of lab meetings. Therefore, by designing core curriculum modules and community building modules, it would assist our “facilitators” (individuals tasked with leading DE-SILO modules at respective institutions) in leading discussions within in their lab. We piloted our project at four universities: UCSF, CalTech, University of Washington, and University of Washington, Saint Louis. In conclusion, this dissertation sheds light on the intricate processes of RGP during ACD in the developing vertebrate brain. By elucidating the role of Par-3 in regulating polarity and Notch activity during ACD, we contribute to a deeper understanding of regulation of cell polarity. Our findings underscore the significance of proper RGP divisions for normal brain development and highlight the potential implications for addressing neurodevelopmental disorders and brain tumors. Through our investigations using zebrafish as a model organism, we uncover novel insights into the dynamic interplay between cortical and cytoplasmic Par-3 localization, orchestrated by AurkA-mediated phosphorylation. This regulatory mechanism governs the fate of neural progenitors, offering new avenues for therapeutic exploration. Furthermore, our commitment to DEI is evident in our DE-SILO project, which aims to foster inclusive environments within academic laboratory settings. By integrating DEI principles into laboratory meetings, we promote meaningful discussions and cultivate a more equitable scientific community. In essence, this dissertation not only advances our understanding of fundamental neurobiological processes but also underscores the importance of inclusivity in scientific research and academia.