Notch Signaling Promotes the Generation of EphrinB1-Positive Intestinal Epithelial Cells

BACKGROUND & AIMS: The intestinal epithelium consists of EphB2-positive proliferative basal cryptic cells and Ephrin B1-positive, postmitotic differentiated cells. We investigated the effects of Notch signaling on formation of the EphB2-EphrinB1 boundary using Mouse and tissue culture models. METHODS: We created mice in which Mind bomb-1 (Mib1), an essential E3 ubiquitin ligase that activates Notch ligands, was inactivated specifically in the intestinal epithelia (Vil-Cre;Mib1(f/f)); Notch is, therefore, inactivated in this tissue. We also studied the effects of different inhibitors on intestinal epithelial cells (IEC-6) that express activated Notch. Tissues and cells were analyzed by immunohistochemical and immunoblot analyses. RESULTS: The intestinal epithelia of Vil-Cre;Mib1(f/f) mice had reduced numbers of EphrinB1-positive cells, compared with controls, but increases in EphB2-positive cells; beta-catenin was activated in these cells. These phenotypes were reversed by expression of a constitutively active form of Notch1. In the IEC-6 cells, Notch signaling activated the expression of EphrinB1 in an Hes1-independent manner, but down-regulated the expression of EphB2 through the GSK3 beta-mediated inhibition of beta-catenin. CONCLUSIONS: Notch signaling regulates formation of the EphB2-EphrinB1 boundary in the mouse intestinal epithelium.