Project Summary Neurulation is a crucial stage in human development when the nervous system is patterned, and the neural tube is shaped. A large fraction of severe birth defects as well as post-implantation spontaneous abortions occur at this stage. Despite its importance, this stage of human development is almost completely inaccessible as IVF embryos cannot be cultured until this point, and recovery of tissue samples from these early stages is rare. Even if such methods were possible, they present ethical dilemmas. A powerful emerging alternative is to create self- organizing models of these events starting from human pluripotent stem cells (hPSCs). We have created a two- dimensional system that mimics the early patterning of the ectodermal medial-lateral axis and is ideal for studying self-organization of ectodermal fates which include the nervous system, skin, neural crest, and sensory organs. We have also built upon this system to create a controlled three-dimensional system that undergoes reproducible morphogenesis reminiscent of neural tube closure. Here we propose to refine these models, as well as to create new models of the anterior-posterior axis, and to use these models together with live cell reporters to probe the interplay between morphogen signaling through the Wnt, BMP, and FGF pathways, patterning along the anterior- posterior (AP) and medial-lateral (ML) axes, and neural tube morphogenesis. We will address key questions about how spatial patterns of morphogen signals are organized and how they are interpreted by cells to give rise to patterns of fates and the physical movements that shape tissues. Taken together, these experiments will provide insight into ectodermal patterning and morphogenesis and establish a system for studying birth defects associated with this stage of development.