The unlimited potential of stem cells stokes the imagination with promises of eternal life and cures for disease. But we have just begun the quest to unwrap the stem cell mystery. In the intestine, the key to the identity of the stem cell is locked away in the complexity of networked signaling pathways, leaving the intestinal stem cell enigmatic. Previous studies that impact the intestinal stem cell environment (niche) provided a snapshot of the dynamic process of stem cell selection and subsequent differentiation. The signaling network that drives this process most certainly acts in a temporal fashion. However, a temporal analysis of this process is lacking. We will use an inducible Cre recombinase system to abrogate beta-catenin-mediated signaling in the stem cell compartment during gut morphogenesis and for the first time in the adult stem cell niche. We hypothesize that beta-catenin mediated signaling is central in coordinating the signaling network that selects the stem cell and allows differentiation of the lineage progenitors. To test our hypothesis, we propose three specific aims. First we will establish a mouse line that harbors an inducible dominant negative Tcf-4 cDNA (DNTcf-4). This dominant negative mutant inhibits beta-catenin-mediated signaling. The DNTcf-4 mouse will be integrated into our existing inducible Cre system to temporally control beta-catenin mediated signaling in the mouse intestinal epithelium at distinct developmental periods during gut morphogenesis and adulthood. Second, we will exploit the comparison between development and maintenance of the stem cell niche, as it represents a seldom tapped resource. In addition, we will determine if altering beta-catenin mediated signaling at defined developmental periods affects clown stream differentiation of intestinal cell lineages. Third, we will explore the temporal relationships between Wnt signaling and the Sonic Hedgehog (Shh) and Notch signaling pathways. These studies will determine if a signaling cascade is responsible for defining """"""""stemness"""""""" or if a yet to be defined master regulator molecule directs synergy of the multiple pathways.
