Project description:We use single cell RNA sequencing to describe the transcriptional changes during gastruloid development from 24 to 84 hours with 12 hours intervals.
Project description:Intestinal organoids are complex three-dimensional structures that mimic cell type composition and tissue organization of the intestine by recapitulating the self-organizing capacity of cell populations derived from a single stem cell. Crucial in this process is a first symmetry-breaking event, in which only a fraction of identical cells in a symmetrical cyst differentiate into Paneth cells, which in turn generates the stem cell niche and leads to asymmetric structures such as crypts and villi. We here combine a quantitative single-cell gene expression and imaging approach to characterize the development of intestinal organoids from a single cell. We show that intestinal organoid development follows a regeneration process driven by transient Yap1 activation. Cell-to-cell variability in Yap1, emerging in symmetrical cysts, initiates a Notch/Dll1 lateral inhibition event driving the symmetry-breaking event and the formation of the first Paneth cell. Our findings reveal how single cells exposed to a uniform growth-promoting environment have the intrinsic ability to generate emergent, self-organized behavior resulting in the formation of complex multicellular asymmetric structures.
Project description:Intestinal organoids are complex three-dimensional structures that mimic cell type composition and tissue organization of the intestine by recapitulating the self-organizing capacity of cell populations derived from a single stem cell. Crucial in this process is a first symmetry-breaking event, in which only a fraction of identical cells in a symmetrical cyst differentiate into Paneth cells, which in turn generates the stem cell niche and leads to asymmetric structures such as crypts and villi. We here combine a quantitative single-cell gene expression and imaging approach to characterize the development of intestinal organoids from a single cell. We show that intestinal organoid development follows a regeneration process driven by transient Yap1 activation. Cell-to-cell variability in Yap1, emerging in symmetrical cysts, initiates a Notch/Dll1 lateral inhibition event driving the symmetry-breaking event and the formation of the first Paneth cell. Our findings reveal how single cells exposed to a uniform growth-promoting environment have the intrinsic ability to generate emergent, self-organized behavior resulting in the formation of complex multicellular asymmetric structures.
Project description:Gastruloids are highly scalable, three-dimensional assemblies generated from pluripotent stem cells that recapitulate fundamental principles of embryonic pattern formation in vitro. Using single cell RNA and multiome sequencing we provide a comprehensive resource mapping cellular states and cell types found during gastruloid development and compare them to the in vivo embryo. We further develop a high throughput gastruloid handling and imaging pipeline to spatially monitor cell type emergence and unfolding of symmetry breaking during gastruloid development. We report spatial variability of pluripotency states in early gastruloids that determines a binary cell response to Wnt activation. While cells situated in the core of the gastruloid revert to an ectopic pluripotent state, peripheral cells differentiate to a primitive streak like state. These two populations then cause gastruloids to break radial symmetry, allowing axial elongation and commitment to the three embryonic germ layers. Finally by performing a phenotypic compound screen, we perturb thousands of gastruloids at relevant developmental time points deriving a phenotypic landscape and inferring molecular regulator networks underlying gastruloid development. Employing this resource, we improve the formation of anterior structures in the existing gastruloid model, using a dual Wnt modulation approach to differentiate an anterior ectopic pluripotent core to anterior ecto- and endodermal structures. This work gives is a resource to understand how gastruloids develop and, more generally, how homogenous cell populations can generate complex patterns in vitro.
Project description:The human embryo breaks symmetry to form the anterior-posterior axis of the body. As the embryo elongates along this axis, progenitors in the tailbud give rise to tissues that generate the spinal cord, skeleton, and musculature. This raises the question of how the embryo achieves axial elongation and patterning. While ethics necessitate in vitro studies, the variability of organoid systems has hindered mechanistic insights. Here we developed a bioengineering and machine learning framework that optimizes symmetry breaking by tuning the spatial coupling between human stem cell-derived organoids. This framework enabled the reproducible generation of axially elongating organoids, each possessing a tailbud and neural tube. We discovered that an excitable system composed of WNT/FGF signaling drives elongation through induction of a neuromesodermal progenitor (NMP)-like signaling center. We discovered that instabilities in the excitable system are suppressed by secreted WNT inhibitors. Absence of these inhibitors led to ectopic tailbuds and branches. Our results identify mechanisms governing stable human axial elongation.