SUMMARY The continuously growing mouse incisor serves as a valuable model to study stem cell regulation during organ renewal. Epithelial stem cells are localized in the proximal end of the incisor in the labial cervical loop. Here, we show that the transcription factor Sox2 is a specific marker for these stem cells. Sox2+ cells became restricted to the labial cervical loop during tooth morphogenesis, and they contributed to the renewal of enamel-producing ameloblasts as well as all other epithelial cell lineages of the tooth. The early progeny of Sox2-positive stem cells transiently expressed the Wnt inhibitor Sfrp5. Sox2 expression was regulated by the tooth initiation marker FGF8 and specific miRNAs, suggesting a fine-tuning to maintain homeostasis of the dental epithelium. The identification of Sox2 as a marker for the dental epithelial stem cells will facilitate further studies on their lineage segregation and differentiation during tooth renewal.

Heterozygous germline mutations in p63, a transcription factor of the p53 family, result in abnormal morphogenesis of the skin and its associated structures, including hair follicles and teeth. In mice lacking p63, all ectodermal organs fail to develop, and stratification of the epidermis is absent. We show that the ectodermal placodes that mark early tooth and hair follicle morphogenesis do not form in p63-deficient embryos, although the multilayered dental lamina that precedes tooth placode formation develops normally. The N-terminally truncated isoform of p63 (⌬Np63) was expressed at high levels in embryonic ectoderm at all stages of tooth and hair development, and it was already dominant over the transactivating TAp63 isoform prior to epidermal stratification. Bmp7, Fgfr2b, Jag1 and Notch1 transcripts were co-expressed with ⌬Np63 in wild-type embryos, but were not detectable in the ectoderm of p63 mutants. In addition, ␤-catenin and Edar transcripts were significantly reduced in skin ectoderm. We also demonstrate that BMP2, BMP7 and FGF10 are potent inducers of p63 in cultured tissue explants. Hence, we suggest that p63 regulates the morphogenesis of surface ectoderm and its derivatives via multiple signalling pathways.

Most characteristics of tooth shape and pattern can be altered by modulating the signal pathways mediating epithelial-mesenchymal interactions in developing teeth. These regulatory signals function in complex networks, characterized by an abundance of activators or inhibitors. In addition, multiple specific inhibitors of all conserved signal pathways have been identified as modulators in tooth development. The number of teeth as well as molar cusp patterns can be modified by tinkering with several different signal pathways. The inhibition of any of the major conserved signal pathways in knockout mice leads to arrested tooth formation. On the other hand, the stimulation of the Wnt pathway in the oral epithelium in transgenic mice leads to abundant de novo tooth formation. The modulation of some of the signal pathways can rescue the development of vestigial tooth rudiments in the incisor and molar regions resulting in extra premolar-like teeth. The size and the degree of asymmetry of the continuously growing mouse incisor can be modulated by modifying the complex network of FGF, bone morphogenetic protein, and Activin signals, which regulate the proliferation and differentiation of epithelial stem cells. Follistatin, Sprouty, and Sostdc1 are important endogenous inhibitors antagonizing these pathways and they are also involved in regulation of enamel formation, and patterning of teeth in crown and root domains. All these findings support the hypothesis that the diversity of tooth types and dental patterns may have resulted from tinkering with the conserved signal pathways, organized into complex networks, during evolution.

The rodent incisor grows continuously throughout its lifetime. The epithelial stem cell niche is located at the apical end of the tooth and its progeny gives rise to the ameloblasts that form the hard enamel. Previously, mesenchymal FGF10 was shown to support the niche, in conjunction with epithelial Notch signaling. Here we show that in a different continuously growing tooth type, the molar of the sibling vole, a similar regulatory system is in place. Moreover, the identical expression pattern of Bmp4 compared to Fgf10 suggests that BMP4 could also be involved in the regulation of the epithelial stem cell niche.Notch and FGF10 signaling is mainly absent in the mouse molar, which stops growing and develops roots. The regulation of the epithelial stem cell niche seems to be flexible allowing for the existence of different tooth types, such as continuously growing teeth, and high and low crowned molars.