To assess the critical role of Wnt signals in intestinal crypts, we generated transgenic mice ectopically expressing Dickkopf1 (Dkk1), a secreted Wnt inhibitor. We find that epithelial proliferation is greatly reduced coincidentally with the loss of crypts. Although enterocyte differentiation appears unaffected, secretory cell lineages are largely absent. Disrupted intestinal homeostasis is reflected by an absence of nuclear ␤-catenin, inhibition of c-myc expression, and subsequent up-regulation of p21. Thus, our data are the first to establish a direct requirement for Wnt ligands in driving proliferation in the intestinal epithelium, and also define an unexpected role for Wnts in controlling secretory cell differentiation.Supplemental material is available at http://www.genesdev.org.

Lgr5 stem cells reside at small intestinal crypt bottoms, generating both the enterocyte and secretory lineage. Entry into the latter epithelial lineage requires silencing of Notch signaling. The Notch ligand Dll1 is strongly up-regulated in a small subset of immediate stem cell daughters. Lineage tracing utilizing a novel Dll1GFP-ires-CreERT2 knock-in mouse reveals that single Dll1high cells generate small, short-lived clones of all four secretory cell types. In culture, sorted Dll1high cells can form long-lived organoids when briefly exposed to Wnt3A. When Dll1 cells are genetically marked prior to tissue damage, significant numbers of stem cell tracing events occur. Lineage specification therefore occurs already in the earliest stem cell daughters through Notch lateral inhibition. Yet, specified secretory progenitors display plasticity and can regain stemness upon tissue damage.

The gut epithelium has remarkable self-renewal capacity that under homeostatic conditions is driven by Wnt signalling in Lgr5(+) intestinal stem cells (ISCs). However, the mechanisms underlying ISC regeneration after injury remain poorly understood. The Hippo signalling pathway mediates tissue growth and is important for regeneration. Here we demonstrate in mice that Yap, a downstream transcriptional effector of Hippo, is critical for recovery of intestinal epithelium after exposure to ionizing radiation. Yap transiently reprograms Lgr5(+) ISCs by suppressing Wnt signalling and excessive Paneth cell differentiation, while promoting cell survival and inducing a regenerative program that includes Egf pathway activation. Accordingly, growth of Yap-deficient organoids is rescued by the Egfr ligand epiregulin, and we find that non-cell-autonomous production of stromal epiregulin may compensate for Yap loss in vivo. Consistent with key roles for regenerative signalling in tumorigenesis, we further demonstrate that Yap inactivation abolishes adenomas in the Apc(Min) mouse model of colon cancer, and that Yap-driven expansion of Apc(-/-) organoids requires the Egfr module of the Yap regenerative program. Finally, we show that in vivo Yap is required for progression of early Apc mutant tumour-initiating cells, suppresses their differentiation into Paneth cells, and induces a regenerative program and Egfr signalling. Our studies reveal that upon tissue injury, Yap reprograms Lgr5(+) ISCs by inhibiting the Wnt homeostatic program, while inducing a regenerative program that includes activation of Egfr signalling. Moreover, our findings reveal a key role for the Yap regenerative pathway in driving cancer initiation.