Genetic vulnerability to Crohn's disease reveals a spatially resolved epithelial restitution program.

Effective tissue repair requires coordinated intercellular communication to sense damage, remodel the tissue, and restore function. Here, we dissected the healing response in the intestinal mucosa by mapping intercellular communication at single-cell resolution and integrating with spatial transcriptomics. We demonstrated that a risk variant for Crohn's disease, hepatocyte growth factor activator (HGFAC) Arg509His (R509H), disrupted a damage-sensing pathway connecting the coagulation cascade to growth factors that drive the differentiation of wound-associated epithelial (WAE) cells and production of a localized retinoic acid (RA) gradient to promote fibroblast-mediated tissue remodeling. Specifically, we showed that HGFAC R509H was activated by thrombin protease activity but exhibited impaired proteolytic activation of the growth factor macrophage-stimulating protein (MSP). In Hgfac R509H mice, reduced MSP activation in response to wounding of the colon resulted in impaired WAE cell induction and delayed healing. Through integration of single-cell transcriptomics and spatial transcriptomics, we demonstrated that WAE cells generated RA in a spatially restricted region of the wound site and that mucosal fibroblasts responded to this signal by producing extracellular matrix and growth factors. We further dissected this WAE cell-fibroblast signaling circuit in vitro using a genetically tractable organoid coculture model. Collectively, these studies exploited a genetic perturbation associated with human disease to disrupt a fundamental biological process and then reconstructed a spatially resolved mechanistic model of tissue healing.

Stepwise-edited, human melanoma models reveal mutations' effect on tumor and microenvironment.

Establishing causal relationships between genetic alterations of human cancers and specific phenotypes of malignancy remains a challenge. We sequentially introduced mutations into healthy human melanocytes in up to five genes spanning six commonly disrupted melanoma pathways, forming nine genetically distinct cellular models of melanoma. We connected mutant melanocyte genotypes to malignant cell expression programs in vitro and in vivo, replicative immortality, malignancy, rapid tumor growth, pigmentation, metastasis, and histopathology. Mutations in malignant cells also affected tumor microenvironment composition and cell states. Our melanoma models shared genotype-associated expression programs with patient melanomas, and a deep learning model showed that these models partially recapitulated genotype-associated histopathological features as well. Thus, a progressive series of genome-edited human cancer models can causally connect genotypes carrying multiple mutations to phenotype.