Epithelial stratification and placode invagination are separable functions in early morphogenesis of the molar tooth

Ectodermal organs, which include teeth, hair follicles, mammaryducts, and glands such as sweat, mucous and sebaceous glands, areinitiated in development as placodes, which are epithelial thickeningsthat invaginate and bud into the underlying mesenchyme. Theseplacodes are stratified into a basal and several suprabasal layers ofcells. The mechanisms driving stratification and invagination arepoorly understood. Using the mouse molar tooth as a model forectodermal organ morphogenesis, we show here that vertical,stratifying cell divisions are enriched in the forming placode and thatstratification is cell division dependent. Using inhibitor and gain-offunctionexperiments, we show that FGF signalling is necessary andsufficient for stratification but not invagination as such.We show that,instead, Shh signalling is necessary for, and promotes, invaginationonce suprabasal tissue is generated. Shh-dependent suprabasal cellshape suggests convergent migration and intercalation, potentiallyaccounting for post-stratification placode invagination to bud stage.We present a model in which FGF generates suprabasal tissue byasymmetric cell division, while Shh triggers cell rearrangement in thistissue to drive invagination all the way to bud formation.