Inflation-collapse dynamics drive patterning and morphogenesis in intestinal organoids

How stem cells self-organize to form structured tissues is an unsolved problem. Intestinal organoids offer a model of self-organization as they generate stem cell zones (SCZs) of typical size even without a spatially structured environment. Here we examine processes governing the size of SCZs. We improve the viability and homogeneity of intestinal organoid cultures to enable long-term time-lapse imaging of multiple organoids in parallel. We find that SCZs are shaped by fission events under strong control of ion channel-mediated inflation and mechanosensitive Piezo-family channels. Fission occurs through stereotyped modes of dynamic behavior that differ in their coordination of budding and differentiation. Imaging and single-cell transcriptomics show that inflation drives acute stem cell differentiation and induces a stretch-responsive cell state characterized by large transcriptional changes, including upregulation of Piezo1. Our results reveal an intrinsic capacity of the intestinal epithelium to self-organize by modulating and then responding to its mechanical state. [Display omitted] •Sandwich culture facilitates long-term live imaging of intestinal organoids•Inflation and collapse of the organoid lumen drives stem cell zone (SCZ) fission•Organoid inflation and SCZ fission are mediated by CFTR and Piezo1•Organoid inflation induces a stretch-associated cell state Intestinal organoids contain stem cell zones (SCZs) that resemble the crypt base in vivo. We studied SCZ patterning by live imaging and perturbing organoids in “sandwich” cultures. We found that SCZs undergo Piezo-dependent fission driven by inflation and collapse of the organoid lumen, and inflation induces a stretch-associated cell state.