Stern, MichaelWarmflash, Aryeh2024-01-242024-01-242023-122023-11-28December 2Ortiz Salazar, Miguel Ángel . "Endogenous Nodal diverges Wnt signaling interpretation from definitive endoderm to posterior mesoderm in geometrically constrained human pluripotent cells." PhD diss., Rice University. https://hdl.handle.net/1911/115409https://hdl.handle.net/1911/115409Embryonic development is a dynamic and highly complex process. It demands the intricate coordination of several mechanisms and signaling pathways to regulate cell fate determination, tissue patterning, and functional integrity, ensuring the embryo’s proper formation until birth. One crucial signaling cascade in development is the Wnt pathway. It is essential during the early stages, as it directs gastrulation, the phase in which the embryo starts forming the germ layers, particularly the endoderm and mesoderm. The Wnt pathway also plays a role in establishing the anterior-to-posterior body axis and helps in elongating the body. However, the precise mechanisms of how Wnt coordinates these diverse stages yielding different outcomes remain not fully elucidated. In this work, we aimed to study the Wnt pathway and its role in cell fate decisions. We uncovered that a posterior-progenitor-like state is induced when the posterior signals, WNT3a and FGF8 are presented to human embryonic stem cell colonies in standard culture. However, when these colonies are geometrically constrained, colonies self- organize themselves into a structure composed of a definitive endoderm cell population surrounding and lifting an epiblast disk-like structure. These unprecedented colonies do not change cell fate when Wnt levels are increased, challenging the classic concentration- dependent morphogen mechanism. While studying this discrepancy, we were able to determine that TGF-ß pathway is responsible for maintaining the definitive endoderm cell fates. Only when this pathway was perturbed, cells were able to respond in a concentration-dependent manner to Wnt and acquire posterior mesoderm cell fates. Upon further inquiry, we were able to determine that specifically, Nodal is the main driver for endoderm cell fates. Finally, we determined that CHIR, a commonly used chemical Wnt activator, induces posterior cell fates in a concentration-dependent manner qualitatively different from induced WNT signaling dynamics and Nodal ligand production. Collectively, we demonstrate that cell fate decision-making is determined by the interplay between multiple pathways and not by the levels of a single pathway, highlighting the dynamic nature of development.application/pdfengCopyright is held by the author, unless otherwise indicated. Permission to reuse, publish, or reproduce the work beyond the bounds of fair use or other exemptions to copyright law must be obtained from the copyright holder.signaling dynamicshuman embryonic stem cellsWntFgfmicropatternsThesis2024-01-24