Tissue-scale mechanics controls differentiation strategy and dynamics of epithelial multilayering

Abstract Generating and maintaining multilayered epithelia requires coordinated cell division, differentiation, and tissue architecture, yet the precise mechanisms of multilayering remain unclear. Using the developing mouse epidermis, we show that basal stem cells adopt distinct multilayering strategies depending on tissue mechanics. Combining quantitative morphometry, embryo live imaging and physical modeling, we observe that early in development, the epidermis is fluid-like, allowing undifferentiated cells to move suprabasally through perpendicular divisions or basal detachment before differentiating. As the tissue matures and rigidifies, a mechanical barrier is established that only allows upward movement of basal cells that have committed to differentiation. The final step of this commitment is delamination that requires Notch signaling, triggered by increased tissue stiffness and jamming. This mechanical regulation orchestrates a feedback loop that induces cell upward motion precisely when the basal layer becomes crowded. Together, our findings identify tissue mechanics as the key determinant of how tissues drive multilayering and reveal mechanically regulated Notch signaling as a driver of epidermal delamination.