Figure S3.

Notch1 regulates epithelial architecture, adherens junctions, cortical actin organization, and proliferation in human epithelia. (A) Immunofluorescence micrographs of Scramble, NOTCH1^KO, and ICD^KO MCF7 cells immunostained for E-cadherin (black) and Notch1 (black) and labeled with phalloidin (black). Scale bar, 20 µm. (B) YZ orthogonal projections from micrographs of Scramble, NOTCH1^KO, and ICD^KO MCF7 cells labeled with Hoechst (white). (C) Quantification of the frequency of focal adherens junctions in scramble control (SCR), NOTCH1^KO, and ICD^KO MCF7 cells. n ≥ 12 fields of view from three independent experiments. (D) Relative number of SCR, NOTCH1^KO, and ICD^KO MCF7 cells measured at passage. n = 3 independent experiments. (E) Top: Immunofluorescence micrographs of Scramble (top row) and NOTCH1^KO (bottom row) Caco-2 cells immunostained for E-cadherin (black) and Notch1 (black) and labeled with phalloidin (black). Scale bar, 20 µm. Bottom: YZ orthogonal projections from micrographs of Scramble and NOTCH1^KO Caco-2 cells labeled with phalloidin (black). (F) Top: Fluorescence micrographs of Scramble (top row) and NOTCH1^KO (bottom row) 16hbe14o- cells immunostained for E-cadherin (black) and Notch1 (black) and labeled with phalloidin (black). Scale bar, 20 µm. Bottom: YZ orthogonal projections from micrographs of Scramble and NOTCH1^KO 16hbe14o- cells labeled with phalloidin (black). (G) Relative number of SCR and NOTCH1^KO Caco-2 cells measured at passage. n = 2 independent experiments. (H) Relative number of SCR and NOTCH1^KO 16hbe14o- cells measured at passage. n = 3 independent experiments. For plots C and D, mean ± SEM; one-way ANOVA with Tukey’s post-hoc test, *P < 0.05, ****P < 0.0001, ns denotes non-significant. For plot H, mean ± SEM; two-tailed unpaired t test, **P < 0.01.