The initiation knot is a signaling center required for molar tooth development

Signaling centers, or organizers, regulate many aspects of embryonic morphogenesis. In the mammalian molar tooth, reiterative signaling in specialized centers called enamel knots (EKs) determine tooth patterning. Preceding the first, primary EK, a transient epithelial thickening appears whose significance remains debated. Using tissue confocal fluorescence imaging with laser ablation experiments, we show that this transient thickening is an earlier signaling center, the molar initiation knot (IK), required for the progression of tooth development. IK cell dynamics manifest signaling center hallmarks; cell cycle exit, condensation, and eventual apoptotic silencing. IK initiation and maturation are defined by the juxtaposition of high Wnt activity cells to Shh-expressing non-proliferating cells, the combination of which drives the growth of the tooth bud, leading to the formation of the primary EK as an independent cell cluster. Overall, the whole development of the tooth, from initiation to patterning, is driven by the iterative use of signaling centers.