Analysis of genetic interactions and hierarchies of Wnt-signaling components in vivo

Abstract

In cooperation with other signaling cascades the Wnt/β-catenin signaling pathway is an essential regulator of embryonic development as well as adult tissue homeostasis and stem cell proliferation. Mutations in components of this signal transduction pathway, which lead to aberrant activation of β-catenin, are frequently observed in human cancers including colon cancer and other degenerative diseases. Pygopus proteins have emerged as nuclear co-factors of β-catenin and are supposed to act in a context dependent way by modulating canonical Wnt signaling. Pygo2, one of the mammalian Pygopus homologues, is involved in neoplastic transformation of multiple cell types. Although overexpressed in colon cancer its role in intestinal tumorigenesis is not yet investigated. This study shows for the first time, that Pygo2 is dispensable for the normal intestinal homeostasis, but is essential for intestinal tumor development induced by mutant β-catenin. Conditional Villin-Cre mediated deletion of Pygo2 in mouse intestinal epithelial cells did not affect the normal embryonic development or the homeostasis of adult intestine. Moreover, loss of Pygo2 did not influence the expression of Wnt/β-catenin target genes. However, Pygo2 deficiency in the context of aberrant Wnt signaling mediated by stabilization of β-catenin completely blocked intestinal hyperproliferation and rescued the increased expression of Wnt/β-catenin target genes and of intestinal stem cell marker. Surprisingly, when deleted in ApcMin/+ mice Pygo2 failed to prevent or even reduce tumor development. Comparing both mouse models we found upregulation of BCL9-2 in intestinal adenomas of ApcMin/+ mice but not in the hyperproliferation of β-catenin mutant mice, which suggests that BCL9-2 might be sufficient to promote tumor progression in the context of Apc deficiency. We further demonstrated that both Pygo2 and BCL9-2 were required for the proliferation of human colon cancer cells in vitro. In addition, we found phosphorylation of ERK1/2, a downstream effector of K-ras, in intestinal adenomas of ApcMin/+ mice but not in the early hyperproliferation of β-catenin mutant mice, indicating that Apc deficient tumors acquired additional mutations. Taken together, our data suggest that the function of Pygo2 in the intestine is context dependent. Pygo2 appears to be redundant for the normal intestinal epithelium, but may be required for tumor initiation induced by aberrant Wnt/β-catenin signaling. Therefore, Pygo2 might be an attractive therapeutic target in early intestinal tumors that arise upon mutation of β-catenin.