Modulating Mesendoderm Competence During Human Germ Layer Differentiation

Abstract

As pluripotent human embryonic stem cells progress towards one germ layer fate, they lose the ability to adopt alternative fates. It is unknown how the cells’ competence for these alternative fates changes along their developmental trajectory or if this competence can be modulated. Here, we show that a differentiating stem cell’s probability of adopting a mesendodermal fate given appropriate signals falls sharply at a point along the ectodermal differentiation trajectory, and demonstrate that this point can be moved using genetic perturbations. Using a low-dimensional reaction coordinate to monitor progression towards ectoderm, we can determine the probability that individual cells at different points along this path are competent to adopt mesendodermal fate upon BMP4 and Activin A signal exposure. Knowing this probability allows us to prospectively isolate and profile differentiating cells based on their mesendoderm competence. We then analyze RNA-seq and ATAC-seq profiles of these different populations to identify transcription factors that control the cell’s mesendoderm competence. By modulating these key transcription factors, we can expand the window of competence to adopt the mesendodermal fate along the ectodermal differentiation trajectory. The ability of the underlying gene regulatory network to modulate competence is essential for understanding human development and controlling the fate choices of stem cells in vitro.