Date of Award


Degree Name

Doctor of Philosophy


Biological Sciences


Anita M. Quintana


Neural precursor cells (NPCs) are the stem-like cells of the developing brain. These cells differentiate into the differentiated cell types of the central nervous system. If disruption occurs in the number of total NPCs formed or their survival, various disorders including learning disabilities, behavioral problems, or epilepsy can occur. This dissertation describes how the HCFC1 gene controls the proliferation and differentiation of NPCs. HCFC1 encodes for a transcriptional co-factor that regulates the growth and metabolism of stem cells, including NPCs. Mutations in HCFC1 cause cblX syndrome, a neural developmental disorder that affects the nervous system and causes microcephaly, epilepsy, and intellectual disability. Since 2013, we have known that missense mutations in HCFC1 cause cblX syndrome with varied neurological impairments. However, since this finding there have been no studies to reveal a mechanism for how this occurs. Collectively, this dissertation describes independent studies using zebrafish as a model to study the function of HCFC1 and reveal a potential mechanism whereby HCFC1 regulates NPC proliferation and differentiation. Our first study describes how a loss of function (LOF) mutation in the hcfc1a gene in zebrafish results in increased numbers of proliferating NPCs and hypomotility. Subsequent RNA sequencing of these LOF mutants identified increased expression of asxl1, a gene that encodes for a transcription factor that modulates cellular proliferation. We show that inhibition of asxl1 expression restored the number of NPCs to normal levels, demonstrating a potential mechanism by which hcfc1a regulates NPC proliferation and brain development. However, several questions remained to be addressed from this work. For example, RNA-sequencing was performed in whole brain homogenates which include a plethora of cells in addition to NPCs. Therefore, we needed to address if Asxl1 increases were directly involved in NPC development or an artifact of other cells such as endothelial or hematopoietic cell populations. Additionally, because of the transcriptional co-factor function of HCFC1, it is plausible that this protein bound to the promoter of Asxl1 to increase mRNA expression, which was not fully investigated. Finally, protein validation of Asxl1 was required to understand if the increases in mRNA also translated to changes in protein. Thus, in our following work, we sought to address these key questions. We used a missense mutant allele of hcfc1a to further investigate the potential mechanism by which Asxl1 might regulate abnormal brain development. We sought to use this allele because it was more representative of cblX patients which inherit hemizygous missense mutations and have little to no protein changes in HCFC1. Using both the missense and nonsense alleles together, we sought to characterize the function of Asxl1 in these two mutant backgrounds. We observed contrasting protein expression of two downstream target genes of HCFC1: Asxl1 and YWHAB, a gene that encodes for the 14-3-3-β/α protein known to regulate AKT phosphorylation and activity. Interestingly, we found contrasting protein expression of Akt and downstream targets of mTor that correlated with contrasting numbers of radial glial cells produced. Based on these results we can conclude that mutations in hcfc1a disrupt Akt/mTOR activity and glial cell development. These findings are the first to suggest a mechanism for how HCFC1 regulates different aspects of neurogenesis. Our data collectively provide a foundational understanding of how mutations in HCFC1 cause neurological impairments in humans and move the cblX field forward by proposing possible therapeutic approaches that can be applied in the future.




Recieved from ProQuest

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Rights Holder

Victoria Lynn Castro