Gene targeting was used to create a null allele at the epidermal growth factor receptor locus (Egfr). The phenotype was dependent on genetic background. EGFR deficiency on a CF-1 background resulted in peri-implantation death due to degeneration of the inner cell mass. On a 129/Sv background, homozygous mutants died at mid-gestation due to placental defects; on a CD-1 background, the mutants lived for up to 3 weeks and showed abnormalities in skin, kidney, brain, liver, and gastrointestinal tract. The multiple abnormalities associated with EGFR deficiency indicate that the receptor is involved in a wide range of cellular activities.

Wnt/beta-catenin signaling plays key roles in tooth development, but how this pathway intersects with the complex interplay of signaling factors regulating dental morphogenesis has been unclear. We demonstrate that Wnt/beta-catenin signaling is active at multiple stages of tooth development. Mutation of beta-catenin to a constitutively active form in oral epithelium causes formation of large, misshapen tooth buds and ectopic teeth, and expanded expression of signaling molecules important for tooth development. Conversely, expression of key morphogenetic regulators including Bmp4, Msx1, and Msx2 is downregulated in embryos expressing the secreted Wnt inhibitor Dkk1 which blocks signaling in epithelial and underlying mesenchymal cells. Similar phenotypes are observed in embryos lacking epithelial beta-catenin, demonstrating a requirement for Wnt signaling within the epithelium. Inducible Dkk1 expression after the bud stage causes formation of blunted molar cusps, downregulation of the enamel knot marker p21, and loss of restricted ectodin expression, revealing requirements for Wnt activity in maintaining secondary enamel knots. These data place Wnt/beta-catenin signaling upstream of key morphogenetic signaling pathways at multiple stages of tooth development and indicate that tight regulation of this pathway is essential both for patterning tooth development in the dental lamina, and for controlling the shape of individual teeth.

Primary cilia are present on most mammalian cells and are implicated in transducing Hedgehog (Hh) signals during development; however, the prevalence of cilia on human tumors remains unclear, and the role of cilia in cancer has not been examined. Here we show that human basal cell carcinomas (BCCs) are frequently ciliated, and we test the role of cilia in BCC by conditionally deleting Kif3a (encoding kinesin family member 3A) or Ift88 (encoding intraflagellar transport protein 88), genes required for ciliogenesis, in two Hh pathway-dependent mouse tumor models. Ciliary ablation strongly inhibited BCC-like tumors induced by an activated form of Smoothened. In contrast, removal of cilia accelerated tumors induced by activated Gli2, a transcriptional effector of Hh signaling. These seemingly paradoxical effects are consistent with a dual role for cilia in mediating both the activation and the repression of the Hh signaling pathway. Our findings demonstrate that cilia function as unique signaling organelles that can either mediate or suppress tumorigenesis depending on the nature of the oncogenic initiating event.Elevated Hh pathway activity is associated with diverse tumors, including basal cell carcinoma, the most commonly diagnosed cancer in North America 1,2 . Hh signaling is normally restrained by the tumor suppressor Patched (Ptch1), which inhibits the function of the protooncogene Smoothened (Smo), a central activator of the pathway 3 . Binding of Hh ligand to Ptch1 relieves its inhibition of Smo, allowing Smo to induce downstream Gli activators and inhibit the formation of Gli repressors. In BCCs, loss of function mutations in PTCH1, gain of function mutations in SMO, and upregulation of GLI1 and GLI2 are frequently observed, suggesting that dysregulated Hh signaling is the underlying cause of this disease [4][5][6][7][8] .Recent studies have shown that the primary cilium has a prominent role in modulating mammalian Hh signaling 9 . Ptch1 suppresses the Hh pathway, at least in part, by preventing the trafficking of Smo into the primary cilium 10 In addition to its essential role in transducing Hh signals, the cilium also negatively regulates the pathway. Genetic analyses have shown that the cilium is required for the proteolytic processing of Gli3 into a form that represses the Hh transcriptional program (Gli3-R) [12][13][14][15] . Cilium-dependent formation of Gli3-R occurs in the absence of Hh and is suppressed upon Smo movement to the cilium. Thus, the cilium exerts both positive and negative control over the Hh pathway.Although cilia are present on many vertebrate cells, their involvement in cancer has not been explored. Early ultrastructural studies have indicated that individual cells from human BCCs can be ciliated 16 ; however, the prevalence of these ciliated cells has remained unclear. To determine whether cancer cells frequently possess cilia, we examined clinical biopsies from eight human BCCs. We observed that five BCCs contained numerous ciliated cells ( Fig. 1a and Supplementary Fig. ...

SUMMARY Smoothened (SMO) inhibitors are under clinical investigation for the treatment of several cancers. Vismodegib is approved for the treatment of locally advanced and metastatic basal cell carcinoma (BCC). Most BCC patients experience significant clinical benefit on vismodegib, however, some develop resistance. Genomic analysis of tumor biopsies revealed vismodegib resistance is associated with Hedgehog (Hh) pathway reactivation, predominantly through mutation of the drug target SMO and to a lesser extent through concurrent copy number changes in SUFU and GLI2. SMO mutations either directly impaired drug binding or activated SMO to varying levels. Furthermore, we found evidence for intra-tumor heterogeneity, suggesting that a combination of therapies targeting components at multiple levels of the Hh pathway is required to overcome resistance.