Human development is complex. Countless molecular events guide a single cell and its daughters through processes like cell division, differentiation and even death to produce a free-living organism that comprises trillions of cells, multiple organs and a complex brain capable of adaptation, emotion and invention. To achieve this end, cells have evolved a highly complex system of signals to communicate with each other throughout development and into adulthood. The Wnt signaling pathway is key among such developmental signals, and is required for multicellular animal life. The role of the Wnt signaling pathway in early human development is poorly understood. Using human pluripotent stem cells (hPSCs) and human neural stem cells (hNSCs), I have undertaken a series of studies designed to elucidate the functions of the Wnt pathway in these models of early human development.
In Chapter 1, I introduce hPSCs, their derivatives (such as hNSCs) and their potential as a model system. I discuss the Wnt signaling pathway and its roles in development and disease. Finally, I introduce the transcription factor SP5, which I discovered is an important target of Wnt signaling in hPSCs and hNSCs.
In Chapter 2, I describe high throughput RNA sequencing expression analysis of Wnt3a-treated hPSCs. I identify SP5 as a key transcriptional target and characterize its binding genome-wide by chromatin immunoprecipitation followed by sequencing. I demonstrate that SP5 functions by repressing Wnt antagonist genes and that this activity restricts the formation of definitive endoderm in Wnt-treated hPSCs.
In Chapter 3, I describe the effects of Wnt pathway activation on pluripotency and differentiation to early germ layers. I further discuss the novel role of Wnt signaling as a negative regulator of EGR1 expression.
In Chapter 4, I demonstrate the effects of Wnt signaling on hNSC multipotency and differentiation. I describe high throughput RNA sequencing expression analysis of Wnt3a-treated hNSCs. I identify FGF family members and SP5 as key transcriptional targets and demonstrate their roles promoting hNSC multipotency and proliferation.
In Chapter 5, I discuss the broader implications of my findings and speculate on the subject and results of future work in the field.