Fig 2: Jagged-1 mediates signal from ECM-sensing basal cells to drive luminal cell differentiation.a JAG1 expression in basal cells from control, DDR1i, and DDR1i Rel tissues. P(DDR1iR vs Ctrl) < 2.5e−14, p(DDR1i vs Ctrl) < 7e−20, p(DDR1iR vs DDR1i) = 0.38. The likelihood ratio test (FDR) was used, not corrected for multiple testing. b JAG1 expression among epithelial clusters. c Immunofluorescent staining of Jagged-1 (green) and E-cadherin (red) in 3D-cultured patient-derived tissues. Results are representative of three independent repeats of this experiment. d, e Flow cytometry analysis of primary tissue in 3D hydrogels treated with DDR1i for 12 days. Basal cells were identified based on EpCAM and CD49f expression and further analyzed for Jagged-1 expression (d). The percentage of Jagged-1+ cells (e) was compared between control and treated groups. Data were derived from n = 3 independent patient samples. P-value = 0.035, two-sided Student’s t-test. * Indicates P ≤ 0.05. Data are presented as mean values ± SD. f–g Development of MCF10A cells (WT, overexpressing LacZ or overexpressing JAG1) in 3D collagen with or without DDR1i. Representative images shown in (f), diameter of organoids shown in (g). Organoid diameter was measured for each treatment group. n = 206 and n = 145 organoids were measured in the WT group cultured under active or inactive DDR1, respectively. n = 166 and n = 92 organoids were measured in the LacZ group cultured under active or inactive DDR1, respectively. n = 183 and n = 92 organoids were measured in the JAG1 group cultured under active or inactive DDR1, respectively. Letters over graph bars indicate statistically different groups, P = 3 × 10−13 (One-way ANOVA with Tukey’s multiple comparisons test). Data are presented as mean values ± SD. h Quantification of secondary organoid development by cells isolated from structures formed by WT, LacZ-overexpressing, or JAG1-overexpressing MCF10A cells with or without DDR1i treatment. Data were derived from n = 10 fields analyzed for each treatment group. Letters over graph bars indicate statistically different groups, P ≤ 0.001 (two-way ANOVA with Tukey’s multiple comparisons test). Data are presented as mean values ± SD. i Immunofluorescent staining of Jagged-1 (green), Notch1 (red) and Hoechst (gray) in 3D-cultured patient-derived tissues. (I) 3D reconstruction of a Z-stack captured by confocal microscopy. (II) a single 2D plane across lobule framed by rectangle in (I). (III) high-power (×60) image of lobule framed by rectangle in (I) and (II). (IV) Enlargement of area framed by rectangle in (III). j Model for the role of DDR1, Jagged-1, and Notch1 within the mammary differentiation hierarchy (left) and visual depiction of how tissue regeneration and morphogenesis is controlled by these factors through spatially regulated cell fate decisions (right). Scale bars = 50 µm. * Indicates p < 0.05; ** indicates p < 0.01. Source data are provided as a Source data file.